laser scanning confocal microscope  (Carl Zeiss)

 
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Name:
    ZEN Module Confocal Topography Hardware License Key
    Description:
    ZEN Module Confocal Topography Hardware License Key Module for analysis of surface data and visualisation of measument results
    Catalog Number:
    410136-1038-230
    Price:
    None
    Category:
    ZEN Software Software ZEN ZEN Modules Material
    Buy from Supplier


    Structured Review

    Carl Zeiss laser scanning confocal microscope
    Cell surface β1,4-galactosyltransferase V (B4GalT5) is not responsible for the stemness of breast cancer. (A) Immunofluorescence analysis of B4GalT5 expression in MCF-7ADR cells. Cells were stained with IgG or anti-B4GalT5 antibody followed by fluorescein isothiocyanate (FITC)–conjugated anti-Rabbit antibodies as described and analyzed by a <t>laser</t> <t>scanning</t> <t>confocal</t> <t>microscope.</t> (B) Cell surface biotinylation assay to compare B4GalT5 localization in plasma membrane and cytoplasmic fractions of MCF-7ADR cells. (C) Construction of MCF-7ADR cells that stably expresses B4GalT5 with a C-terminal 6×His tag. MCF-7ADR cells were transfected with pBABE-B4GalT5-His-IRES-puro plasmid for 48 hours and then selected with 150 μg/mL puromycin. Expression of corresponding proteins was examined by western blotting. (D) Representative images of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. After digested and resuspended, the cells were stained with anti-His antibody at room temperature for 1 hour followed by FITC-conjugated anti-rabbit antibody at room temperature for 1 hour. Pictures were taken by a laser scanning confocal microscope. (E, F) Flow cytometry (FCM) analysis of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. Cells were stained with phycoerythrin (PE)-conjugated anti-His antibody at room temperature for 1 hour and analyzed by Moflo XDP. Error bars represent standard error of the mean (n=3, ***p
    ZEN Module Confocal Topography Hardware License Key Module for analysis of surface data and visualisation of measument results
    https://www.bioz.com/result/laser scanning confocal microscope/product/Carl Zeiss
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    laser scanning confocal microscope - by Bioz Stars, 2021-06
    99/100 stars

    Images

    1) Product Images from "β1,4-Galactosyltransferase V Modulates Breast Cancer Stem Cells through Wnt/β-catenin Signaling Pathway"

    Article Title: β1,4-Galactosyltransferase V Modulates Breast Cancer Stem Cells through Wnt/β-catenin Signaling Pathway

    Journal: Cancer Research and Treatment : Official Journal of Korean Cancer Association

    doi: 10.4143/crt.2020.093

    Cell surface β1,4-galactosyltransferase V (B4GalT5) is not responsible for the stemness of breast cancer. (A) Immunofluorescence analysis of B4GalT5 expression in MCF-7ADR cells. Cells were stained with IgG or anti-B4GalT5 antibody followed by fluorescein isothiocyanate (FITC)–conjugated anti-Rabbit antibodies as described and analyzed by a laser scanning confocal microscope. (B) Cell surface biotinylation assay to compare B4GalT5 localization in plasma membrane and cytoplasmic fractions of MCF-7ADR cells. (C) Construction of MCF-7ADR cells that stably expresses B4GalT5 with a C-terminal 6×His tag. MCF-7ADR cells were transfected with pBABE-B4GalT5-His-IRES-puro plasmid for 48 hours and then selected with 150 μg/mL puromycin. Expression of corresponding proteins was examined by western blotting. (D) Representative images of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. After digested and resuspended, the cells were stained with anti-His antibody at room temperature for 1 hour followed by FITC-conjugated anti-rabbit antibody at room temperature for 1 hour. Pictures were taken by a laser scanning confocal microscope. (E, F) Flow cytometry (FCM) analysis of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. Cells were stained with phycoerythrin (PE)-conjugated anti-His antibody at room temperature for 1 hour and analyzed by Moflo XDP. Error bars represent standard error of the mean (n=3, ***p
    Figure Legend Snippet: Cell surface β1,4-galactosyltransferase V (B4GalT5) is not responsible for the stemness of breast cancer. (A) Immunofluorescence analysis of B4GalT5 expression in MCF-7ADR cells. Cells were stained with IgG or anti-B4GalT5 antibody followed by fluorescein isothiocyanate (FITC)–conjugated anti-Rabbit antibodies as described and analyzed by a laser scanning confocal microscope. (B) Cell surface biotinylation assay to compare B4GalT5 localization in plasma membrane and cytoplasmic fractions of MCF-7ADR cells. (C) Construction of MCF-7ADR cells that stably expresses B4GalT5 with a C-terminal 6×His tag. MCF-7ADR cells were transfected with pBABE-B4GalT5-His-IRES-puro plasmid for 48 hours and then selected with 150 μg/mL puromycin. Expression of corresponding proteins was examined by western blotting. (D) Representative images of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. After digested and resuspended, the cells were stained with anti-His antibody at room temperature for 1 hour followed by FITC-conjugated anti-rabbit antibody at room temperature for 1 hour. Pictures were taken by a laser scanning confocal microscope. (E, F) Flow cytometry (FCM) analysis of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. Cells were stained with phycoerythrin (PE)-conjugated anti-His antibody at room temperature for 1 hour and analyzed by Moflo XDP. Error bars represent standard error of the mean (n=3, ***p

    Techniques Used: Immunofluorescence, Expressing, Staining, Microscopy, Cell Surface Biotinylation Assay, Stable Transfection, Transfection, Plasmid Preparation, Western Blot, Flow Cytometry

    β1,4-Galactosyltransferase V (B4GalT5) promotes Wnt/β-catenin signaling that is hyperactivated in breast cancer stem cells (BCSCs). (A) Immunofluorescence analysis of phosphorylated-GSK3β (Ser 9) expression in BCSCs and non-BCSCs. MCF-7ADR cells were sorted into BCSCs and non-BCSCs, then stained with antibodies and DAPI as described and analyzed by a laser scanning confocal microscope. (B, C) Western blotting analysis of β-catenin and phosphorylated-β-catenin (Ser 45) expression levels in BCSCs and non-BCSCs. Protein band densities were quantified by normalizing to β-actin. (D, E) The effect of B4GalT5 knockdown on Frizzled-1, β-catenin, and B4GalT5 expression levels in MCF-7ADR, MCF-7, and MDA-MB-231 cell lines by western blotting analysis. Protein band densities were quantified by normalizing to β-actin. (F, G) The effect of B4GalT5 overexpression on Frizzled-1, β-catenin, and B4GalT5 expression levels in MCF-7ADR, MCF-7, and MDA-MB-231 cell lines by western blotting analysis. Protein band densities were quantified by normalizing to β-Actin. (H) The effect of B4GalT5 on wnt 3α induced Wnt/β-catenin signaling by western blotting (WB). After transfected using B4GalT5 siRNA for 12 hours, MCF-7ADR cells were starved for 36 hours followed by adding wnt 3α, and subjected to western blotting. (I) The effect of B4GalT5 on Wnt 3α induced ALDH1A1 expression by western blotting. MCF-7ADR/shNC and MCF-7ADR/shB4GalT5 cells were starved for 36 hours followed by adding Wnt 3α, and subjected to western blotting. (J) The degradation pathway of β-catenin due to B4GalT5 knockdown. After transfected using B4GalT5 siRNA for 41 hours, MCF-7ADR cells were treated with MG132 (protease inhibitor) for 7 hours. Protein levels were examined by western blotting. (K) Western blotting analysis of membrane and cytosol proteins in MCF-7ADR cells. MCF-7ADR cells were transfected using B4GalT5 siRNA for 48 hours, extracted into membrane and cytosol proteins and then subjected to western blotting. Erk was used for examining purity of membrane proteins. (L) The degradation pathway of Frizzled-1 on the cell surface of MCF-7ADR cells due to B4GalT5 knockdown. After transfected using B4GalT5 siRNA for 40 hours, MCF-7ADR cells were treated with 20 μM leupeptin (lysosome inhibitor) for 8 hours. The membrane proteins were extracted as described and then subjected to western blotting. Erk was used for examining purity of membrane proteins. (M) The effect of B4GalT5 on biosynthesis of galactosyl-oligosaccharides in glycans of Frizzled-1 by RCA-I lectin pull-down assay. MCF-7ADR/shNC, MCF-7ADR/shB4GalT5, MCF-7ADR/NC, and MCF-7ADR/oe-B4GalT5 cells were lysed and incubated with agarose-bound RCA-I overnight. Total Frizzled-1 was used as loading control. All experiments were performed in three replicates. Data are presented as means±standard error of the mean. *p
    Figure Legend Snippet: β1,4-Galactosyltransferase V (B4GalT5) promotes Wnt/β-catenin signaling that is hyperactivated in breast cancer stem cells (BCSCs). (A) Immunofluorescence analysis of phosphorylated-GSK3β (Ser 9) expression in BCSCs and non-BCSCs. MCF-7ADR cells were sorted into BCSCs and non-BCSCs, then stained with antibodies and DAPI as described and analyzed by a laser scanning confocal microscope. (B, C) Western blotting analysis of β-catenin and phosphorylated-β-catenin (Ser 45) expression levels in BCSCs and non-BCSCs. Protein band densities were quantified by normalizing to β-actin. (D, E) The effect of B4GalT5 knockdown on Frizzled-1, β-catenin, and B4GalT5 expression levels in MCF-7ADR, MCF-7, and MDA-MB-231 cell lines by western blotting analysis. Protein band densities were quantified by normalizing to β-actin. (F, G) The effect of B4GalT5 overexpression on Frizzled-1, β-catenin, and B4GalT5 expression levels in MCF-7ADR, MCF-7, and MDA-MB-231 cell lines by western blotting analysis. Protein band densities were quantified by normalizing to β-Actin. (H) The effect of B4GalT5 on wnt 3α induced Wnt/β-catenin signaling by western blotting (WB). After transfected using B4GalT5 siRNA for 12 hours, MCF-7ADR cells were starved for 36 hours followed by adding wnt 3α, and subjected to western blotting. (I) The effect of B4GalT5 on Wnt 3α induced ALDH1A1 expression by western blotting. MCF-7ADR/shNC and MCF-7ADR/shB4GalT5 cells were starved for 36 hours followed by adding Wnt 3α, and subjected to western blotting. (J) The degradation pathway of β-catenin due to B4GalT5 knockdown. After transfected using B4GalT5 siRNA for 41 hours, MCF-7ADR cells were treated with MG132 (protease inhibitor) for 7 hours. Protein levels were examined by western blotting. (K) Western blotting analysis of membrane and cytosol proteins in MCF-7ADR cells. MCF-7ADR cells were transfected using B4GalT5 siRNA for 48 hours, extracted into membrane and cytosol proteins and then subjected to western blotting. Erk was used for examining purity of membrane proteins. (L) The degradation pathway of Frizzled-1 on the cell surface of MCF-7ADR cells due to B4GalT5 knockdown. After transfected using B4GalT5 siRNA for 40 hours, MCF-7ADR cells were treated with 20 μM leupeptin (lysosome inhibitor) for 8 hours. The membrane proteins were extracted as described and then subjected to western blotting. Erk was used for examining purity of membrane proteins. (M) The effect of B4GalT5 on biosynthesis of galactosyl-oligosaccharides in glycans of Frizzled-1 by RCA-I lectin pull-down assay. MCF-7ADR/shNC, MCF-7ADR/shB4GalT5, MCF-7ADR/NC, and MCF-7ADR/oe-B4GalT5 cells were lysed and incubated with agarose-bound RCA-I overnight. Total Frizzled-1 was used as loading control. All experiments were performed in three replicates. Data are presented as means±standard error of the mean. *p

    Techniques Used: Immunofluorescence, Expressing, Staining, Microscopy, Western Blot, Multiple Displacement Amplification, Over Expression, Transfection, Protease Inhibitor, Pull Down Assay, Incubation

    2) Product Images from "Effect of Astaxanthin on Activation of Autophagy and Inhibition of Apoptosis in Helicobacter pylori-Infected Gastric Epithelial Cell Line AGS"

    Article Title: Effect of Astaxanthin on Activation of Autophagy and Inhibition of Apoptosis in Helicobacter pylori-Infected Gastric Epithelial Cell Line AGS

    Journal: Nutrients

    doi: 10.3390/nu12061750

    Effect of astaxanthin on autophagy activation in H. pylori -stimulated AGS cells. The cells were pre-treated with 50 nM astaxanthin for 3 h and then stimulated with H. pylori (cell to H. pylori ratio of 1:50) for 24 h. ( A ) Cells were stained with acridine orange (AO) dye and visualized under a confocal laser scanning microscope (left panel). AO-positive cells were quantified and expressed as % of cells with AO-positive cells/total number of cells (right panel). ( B ) The cells were stained with anti-LC3B antibody and rhodamine-labeled mouse anti-rabbit IgG antibody. Immunocytochemical staining for LC3B (red) and DNA counterstaining with DAPI (blue) are shown in the left panel. Each sample was analyzed using a threshold of > 7 dots/cell. LC3B puncta-positive cells were quantified and expressed as % of cells with > 7 LC3B puncta/total number of cells (right panel). * p
    Figure Legend Snippet: Effect of astaxanthin on autophagy activation in H. pylori -stimulated AGS cells. The cells were pre-treated with 50 nM astaxanthin for 3 h and then stimulated with H. pylori (cell to H. pylori ratio of 1:50) for 24 h. ( A ) Cells were stained with acridine orange (AO) dye and visualized under a confocal laser scanning microscope (left panel). AO-positive cells were quantified and expressed as % of cells with AO-positive cells/total number of cells (right panel). ( B ) The cells were stained with anti-LC3B antibody and rhodamine-labeled mouse anti-rabbit IgG antibody. Immunocytochemical staining for LC3B (red) and DNA counterstaining with DAPI (blue) are shown in the left panel. Each sample was analyzed using a threshold of > 7 dots/cell. LC3B puncta-positive cells were quantified and expressed as % of cells with > 7 LC3B puncta/total number of cells (right panel). * p

    Techniques Used: Activation Assay, Staining, Laser-Scanning Microscopy, Labeling

    3) Product Images from "20(S)-Ginsenoside Rh2 displays efficacy against T-cell acute lymphoblastic leukemia through the PI3K/Akt/mTOR signal pathway"

    Article Title: 20(S)-Ginsenoside Rh2 displays efficacy against T-cell acute lymphoblastic leukemia through the PI3K/Akt/mTOR signal pathway

    Journal: Journal of Ginseng Research

    doi: 10.1016/j.jgr.2019.07.003

    20(S)-GRh2 promoted autophagy by the PI3K/Akt/mTOR pathway. Jurkat cells were treated with 20(S)-GRh2 alone or combined with the PI3K/Akt/mTOR inhibitor for 24 h. (A) Detection of pEGFP-LC3 using a laser scanning confocal microscope (bar = 5 μM). (B) Quantification of GFP-LC3 puncta per cell. Quantitation represents at least 100 cells counted and scored per treatment. (C) The expression levels of Beclin-1, Atg5, and LC3 were analyzed by Western blot. (D) Quantification of Beclin-1 expression. (E) Quantification of Atg5 expression. (F) Quantification of LC3 expression. Data are expressed as mean ± SD. n = 3 for each group. ⁎ p
    Figure Legend Snippet: 20(S)-GRh2 promoted autophagy by the PI3K/Akt/mTOR pathway. Jurkat cells were treated with 20(S)-GRh2 alone or combined with the PI3K/Akt/mTOR inhibitor for 24 h. (A) Detection of pEGFP-LC3 using a laser scanning confocal microscope (bar = 5 μM). (B) Quantification of GFP-LC3 puncta per cell. Quantitation represents at least 100 cells counted and scored per treatment. (C) The expression levels of Beclin-1, Atg5, and LC3 were analyzed by Western blot. (D) Quantification of Beclin-1 expression. (E) Quantification of Atg5 expression. (F) Quantification of LC3 expression. Data are expressed as mean ± SD. n = 3 for each group. ⁎ p

    Techniques Used: Microscopy, Quantitation Assay, Expressing, Western Blot

    20(S)-GRh2 induced autophagy in Jurkat cells. Cells were treated with different concentrations of 20(S)-GRh2 (0, 17.5, and 35 μM) for 24 h. (A) Cells were stained with MDC, and autophagic vacuoles were detected using a flow cytometer. (B) Quantification of autophagic vacuoles in Jurkat cells. (C) The fluorescence of GFP-LC3 was observed using a laser scanning confocal microscope (bar = 5 μM). (D) The number of GFP-LC3 puncta was quantified in each cell. Quantitation represents at least 100 cells counted and scored per treatment. (E) The ultrastructure of Jurkat cells was visualized using a transmission electron microscope (bar = 5 μM). ⁎ p
    Figure Legend Snippet: 20(S)-GRh2 induced autophagy in Jurkat cells. Cells were treated with different concentrations of 20(S)-GRh2 (0, 17.5, and 35 μM) for 24 h. (A) Cells were stained with MDC, and autophagic vacuoles were detected using a flow cytometer. (B) Quantification of autophagic vacuoles in Jurkat cells. (C) The fluorescence of GFP-LC3 was observed using a laser scanning confocal microscope (bar = 5 μM). (D) The number of GFP-LC3 puncta was quantified in each cell. Quantitation represents at least 100 cells counted and scored per treatment. (E) The ultrastructure of Jurkat cells was visualized using a transmission electron microscope (bar = 5 μM). ⁎ p

    Techniques Used: Staining, Flow Cytometry, Fluorescence, Microscopy, Quantitation Assay, Transmission Assay

    4) Product Images from "Axotrophin/MARCH7 acts as an E3 ubiquitin ligase and ubiquitinates tau protein in vitro impairing microtubule binding"

    Article Title: Axotrophin/MARCH7 acts as an E3 ubiquitin ligase and ubiquitinates tau protein in vitro impairing microtubule binding

    Journal: Biochimica et Biophysica Acta

    doi: 10.1016/j.bbadis.2014.05.029

    Subcellular localization of EGFP–axotrophin fusion proteins. N2A cells were transfected with constructs encoding different axotrophin domains and full length axotrophin as EGFP-fusion proteins. Confocal images of EGFP fluorescence (A1–A7), nuclear DAPI fluorescence (B1–B7) and merge EGFP- and DAPI-fluorescence (M1–M7) were obtained by laser scanning microscopy. Axotrophin expressed as full length protein has a cytoplasmic and nuclear localization (A1, B1). In the cytoplasm axotrophin resides in granular structures in close proximity to the Golgi apparatus. In addition picture A1 shows two cells (marked with asterisk) with more prominent nuclear localization. Upon deletion of aa 626–704 axotrophin exhibits a predominant nuclear localization in a nuclear speckle-like appearance (A2, B2). This nuclear speckle-like localization does change when further amino acids were removed from the C-terminal end such as deleting the RING-variant domain leading to a more heterochromatin associated localization (A3, B3), visible by the large overlap with DAPI stained nuclear foci. After deletion the complete C-terminal half yielding a 344aa construct comprising only the first of two putative DNA/RNA binding domains the nuclear axotrophin localization is confined to smaller nuclear subcompartments (A4, B4). In contrast deletion of aa 1–332, removing the first of two putative DNA/RNA binding domains (A5, B5) results in a predominant cytoplasmic localization without the granular appearance observed with full-length axotrophin (A1, B1). Further deletion of the second putative DNA/RNA binding domain resulting in a construct comprising the RING-variant domain and the C-terminus leads to a cytoplasmic localization of the EGFP-fusion protein with a reticular occurrence (A6, B6). For comparison the dual appearance of solely EGFP in the nuclear and cytoplasmic compartments is shown (A7, B7).
    Figure Legend Snippet: Subcellular localization of EGFP–axotrophin fusion proteins. N2A cells were transfected with constructs encoding different axotrophin domains and full length axotrophin as EGFP-fusion proteins. Confocal images of EGFP fluorescence (A1–A7), nuclear DAPI fluorescence (B1–B7) and merge EGFP- and DAPI-fluorescence (M1–M7) were obtained by laser scanning microscopy. Axotrophin expressed as full length protein has a cytoplasmic and nuclear localization (A1, B1). In the cytoplasm axotrophin resides in granular structures in close proximity to the Golgi apparatus. In addition picture A1 shows two cells (marked with asterisk) with more prominent nuclear localization. Upon deletion of aa 626–704 axotrophin exhibits a predominant nuclear localization in a nuclear speckle-like appearance (A2, B2). This nuclear speckle-like localization does change when further amino acids were removed from the C-terminal end such as deleting the RING-variant domain leading to a more heterochromatin associated localization (A3, B3), visible by the large overlap with DAPI stained nuclear foci. After deletion the complete C-terminal half yielding a 344aa construct comprising only the first of two putative DNA/RNA binding domains the nuclear axotrophin localization is confined to smaller nuclear subcompartments (A4, B4). In contrast deletion of aa 1–332, removing the first of two putative DNA/RNA binding domains (A5, B5) results in a predominant cytoplasmic localization without the granular appearance observed with full-length axotrophin (A1, B1). Further deletion of the second putative DNA/RNA binding domain resulting in a construct comprising the RING-variant domain and the C-terminus leads to a cytoplasmic localization of the EGFP-fusion protein with a reticular occurrence (A6, B6). For comparison the dual appearance of solely EGFP in the nuclear and cytoplasmic compartments is shown (A7, B7).

    Techniques Used: Transfection, Construct, Fluorescence, Laser-Scanning Microscopy, Variant Assay, Staining, RNA Binding Assay

    5) Product Images from "Abundant proliferating cells within early chicken taste buds indicate a potentially “built-in” progenitor system for taste bud growth during maturation in hatchlings"

    Article Title: Abundant proliferating cells within early chicken taste buds indicate a potentially “built-in” progenitor system for taste bud growth during maturation in hatchlings

    Journal: Histology and histopathology

    doi: 10.14670/HH-18-055

    Single-plane laser scanning confocal photomicrographs of a representative taste bud from tissue sections of the base of oral cavity at P45. Solid arrowheads point to PCNA + cells also labeled with α-Gustducin. Long arrows point to PCNA + cells outside the taste bud. White dots encircle the taste bud. Scale bar: 20 μ m.
    Figure Legend Snippet: Single-plane laser scanning confocal photomicrographs of a representative taste bud from tissue sections of the base of oral cavity at P45. Solid arrowheads point to PCNA + cells also labeled with α-Gustducin. Long arrows point to PCNA + cells outside the taste bud. White dots encircle the taste bud. Scale bar: 20 μ m.

    Techniques Used: Labeling

    Representative photomicrographs (single-plane laser scanning confocal) from the tissue sections of the base of oral cavity at P3. A. The tissue sections were immunostained with proliferating cell marker PCNA (green) and epithelial cell marker EpCAM (red). Solid arrowheads point to PCNA + cells in EpCAM + epithelial cells. Long arrows point to PCNA + cells in the epithelium outside of the taste bud. B. Distribution of PCNA + (green) cells in the connective tissue labeled with stromal cell marker Vimentin (red). Solid arrowheads point to a representative proliferating cell in the connective tissue immediately beneath the epithelium. Long arrows point to PCNA + cells in the epithelium outside of the taste bud. Insets show high power images of the cell. C. Abundant distribution of PCNA + (green) proliferating cells in taste buds labeled with specific taste cell marker α-Gustducin (red). Long arrows point to PCNA + cells in the epithelium outside of the taste bud. D. proliferating cells labeled by Ki67 were present surrounding mouse taste bud (K8). Solid arrowheads point to PCNA + cells that are also α-Gustducin + . Open arrowheads point to PCNA + cells without α-Gustducin. Dashed lines demarcate epithelium from connective tissue. White dots encircle the taste buds. Scale bars: 20 μ m, applies to images in the same row.
    Figure Legend Snippet: Representative photomicrographs (single-plane laser scanning confocal) from the tissue sections of the base of oral cavity at P3. A. The tissue sections were immunostained with proliferating cell marker PCNA (green) and epithelial cell marker EpCAM (red). Solid arrowheads point to PCNA + cells in EpCAM + epithelial cells. Long arrows point to PCNA + cells in the epithelium outside of the taste bud. B. Distribution of PCNA + (green) cells in the connective tissue labeled with stromal cell marker Vimentin (red). Solid arrowheads point to a representative proliferating cell in the connective tissue immediately beneath the epithelium. Long arrows point to PCNA + cells in the epithelium outside of the taste bud. Insets show high power images of the cell. C. Abundant distribution of PCNA + (green) proliferating cells in taste buds labeled with specific taste cell marker α-Gustducin (red). Long arrows point to PCNA + cells in the epithelium outside of the taste bud. D. proliferating cells labeled by Ki67 were present surrounding mouse taste bud (K8). Solid arrowheads point to PCNA + cells that are also α-Gustducin + . Open arrowheads point to PCNA + cells without α-Gustducin. Dashed lines demarcate epithelium from connective tissue. White dots encircle the taste buds. Scale bars: 20 μ m, applies to images in the same row.

    Techniques Used: Marker, Labeling

    Distribution of S-phase proliferating cells within taste buds in the palate. A. Bright-field image of the palate (left) and fluorescent image of epithelial sheet immunoreacted with Vimentin to illustrate the distribution of taste buds (right). B. Single-plane laser scanning confocal photomicrographs of representative taste bud tissue sections from mgr, ppr and pr regions of the palate at P3. Open arrowheads point to a BrdU + cell negative for α-Gustducin and Vimentin. Solid arrowheads point to BrdU + cells also labeled with α-Gustducin and/or Vimentin. Long arrows point to BrdU + cells in the epithelium outside of the taste bud. Short arrows point to a BrdU + cell in the connective tissue. Dashed lines demarcate epithelium from connective tissue. White dots encircle the taste buds. Scale bars: A, 2 mm; B, 20 μ m.
    Figure Legend Snippet: Distribution of S-phase proliferating cells within taste buds in the palate. A. Bright-field image of the palate (left) and fluorescent image of epithelial sheet immunoreacted with Vimentin to illustrate the distribution of taste buds (right). B. Single-plane laser scanning confocal photomicrographs of representative taste bud tissue sections from mgr, ppr and pr regions of the palate at P3. Open arrowheads point to a BrdU + cell negative for α-Gustducin and Vimentin. Solid arrowheads point to BrdU + cells also labeled with α-Gustducin and/or Vimentin. Long arrows point to BrdU + cells in the epithelium outside of the taste bud. Short arrows point to a BrdU + cell in the connective tissue. Dashed lines demarcate epithelium from connective tissue. White dots encircle the taste buds. Scale bars: A, 2 mm; B, 20 μ m.

    Techniques Used: Labeling

    6) Product Images from "The piperazine compound ASP activates an auxin response in Arabidopsis thaliana"

    Article Title: The piperazine compound ASP activates an auxin response in Arabidopsis thaliana

    Journal: BMC Genomics

    doi: 10.1186/s12864-020-07203-8

    Chemical genetics screening identified a novel compound ASP that produced abnormal leaf-vein pattern. a-b : Leaf vein pattern of 10-day-old seedlings of Q0990 line grown on 1/2 MS medium ( a ) and 1/2 MS medium with 10 μM ASP ( b ). c-d : Observation of 10-day-old seedlings of DR5::GFP reporter line grown on 1/2 MS medium ( c ) and 1/2 MS medium with 10 μM ASP ( d ) by confocal scanning laser microscope. E: Chemical structural formula of endogenous auxin indole-3-acetic acid (IAA), synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D), and small-molecule compound 1-[(4-bromophenoxy) acetyl]-4-[(4-fluorophenyl) sulfonyl] piperazine (ASP)
    Figure Legend Snippet: Chemical genetics screening identified a novel compound ASP that produced abnormal leaf-vein pattern. a-b : Leaf vein pattern of 10-day-old seedlings of Q0990 line grown on 1/2 MS medium ( a ) and 1/2 MS medium with 10 μM ASP ( b ). c-d : Observation of 10-day-old seedlings of DR5::GFP reporter line grown on 1/2 MS medium ( c ) and 1/2 MS medium with 10 μM ASP ( d ) by confocal scanning laser microscope. E: Chemical structural formula of endogenous auxin indole-3-acetic acid (IAA), synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D), and small-molecule compound 1-[(4-bromophenoxy) acetyl]-4-[(4-fluorophenyl) sulfonyl] piperazine (ASP)

    Techniques Used: Produced, Microscopy

    7) Product Images from "Efficient miRNA Inhibitor with GO-PEI Nanosheets for Osteosarcoma Suppression by Targeting PTEN"

    Article Title: Efficient miRNA Inhibitor with GO-PEI Nanosheets for Osteosarcoma Suppression by Targeting PTEN

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S257084

    Uptake of GO-PEI into osteosarcoma cells. ( A ) TEM images of GO-PEI-incubated and non-incubated cells (3 μg/mL GO-PEI in the cell culture medium). The right panel is enlarged images of white squares in the image of GO-PEI-incubated cells. Scale bars: 1 μm. ( B ) FITC-BSA-labeled GO-PEI (green) is visualized in cells through confocal laser scanning microscopy. Fluorescence images of FITC-labeled GO-PEI (green) within MG63 and U2OS cells are shown. The cell cytoskeleton was stained with α-tubulin (red) for MG63 cells and rhodamine-phalloidin (red) for U2OS cells, and the nuclei were stained with DAPI (blue). The right panel is enlarged images of white squares in the image of GO-PEI-incubated cells. Scale bars: 2 μm. ( C ) Cell viability of MG63 and U2OS cells exposed to different concentrations of GO-PEI was measured by CCK8 assays. * p
    Figure Legend Snippet: Uptake of GO-PEI into osteosarcoma cells. ( A ) TEM images of GO-PEI-incubated and non-incubated cells (3 μg/mL GO-PEI in the cell culture medium). The right panel is enlarged images of white squares in the image of GO-PEI-incubated cells. Scale bars: 1 μm. ( B ) FITC-BSA-labeled GO-PEI (green) is visualized in cells through confocal laser scanning microscopy. Fluorescence images of FITC-labeled GO-PEI (green) within MG63 and U2OS cells are shown. The cell cytoskeleton was stained with α-tubulin (red) for MG63 cells and rhodamine-phalloidin (red) for U2OS cells, and the nuclei were stained with DAPI (blue). The right panel is enlarged images of white squares in the image of GO-PEI-incubated cells. Scale bars: 2 μm. ( C ) Cell viability of MG63 and U2OS cells exposed to different concentrations of GO-PEI was measured by CCK8 assays. * p

    Techniques Used: Transmission Electron Microscopy, Incubation, Cell Culture, Labeling, Confocal Laser Scanning Microscopy, Fluorescence, Staining

    8) Product Images from "Tubeimoside I-induced lung cancer cell death and the underlying crosstalk between lysosomes and mitochondria"

    Article Title: Tubeimoside I-induced lung cancer cell death and the underlying crosstalk between lysosomes and mitochondria

    Journal: Cell Death & Disease

    doi: 10.1038/s41419-020-02915-x

    Tub induced blocking of late-stage autophagic flux in lung cancer cells. a Tub induced an increase in the number of GFP-LC3 puncta. GFP-LC3-overexpressing stable cell lines were treated with the vehicle, rapamycin (Rapa, 0.5 μM), bafilomycin A1 (Baf, 0.1 μM) or Tub (20 μM) for 24 h. Images of the cells were captured with a laser-scanning confocal microscope (scale bar = 20 µm). b Tub induced the upregulation of LC3-II and p62. c Lung cancer cells transfected with mCherry-GFP-LC3 tandem plasmids were treated with the vehicle, HBSS, Baf (0.1 μM) or Tub (20 μM) for 24 h. Like Baf treatment, Tub treatment also caused an increase in yellow fluorescence (creating by the merging of red and green fluorescence emitted by mCherry and GFP, respectively). The images were captured by a laser-scanning confocal microscope. The bar chart (right) represents the colocalization rate of GFP and mCherry, which was calculated with the Image J software (Scale bar = 5 µm). *** p
    Figure Legend Snippet: Tub induced blocking of late-stage autophagic flux in lung cancer cells. a Tub induced an increase in the number of GFP-LC3 puncta. GFP-LC3-overexpressing stable cell lines were treated with the vehicle, rapamycin (Rapa, 0.5 μM), bafilomycin A1 (Baf, 0.1 μM) or Tub (20 μM) for 24 h. Images of the cells were captured with a laser-scanning confocal microscope (scale bar = 20 µm). b Tub induced the upregulation of LC3-II and p62. c Lung cancer cells transfected with mCherry-GFP-LC3 tandem plasmids were treated with the vehicle, HBSS, Baf (0.1 μM) or Tub (20 μM) for 24 h. Like Baf treatment, Tub treatment also caused an increase in yellow fluorescence (creating by the merging of red and green fluorescence emitted by mCherry and GFP, respectively). The images were captured by a laser-scanning confocal microscope. The bar chart (right) represents the colocalization rate of GFP and mCherry, which was calculated with the Image J software (Scale bar = 5 µm). *** p

    Techniques Used: Blocking Assay, Stable Transfection, Microscopy, Transfection, Fluorescence, Software

    9) Product Images from "Ionomycin ameliorates hypophosphatasia via rescuing alkaline phosphatase deficiency-mediated L-type Ca2+ channel internalization in mesenchymal stem cells"

    Article Title: Ionomycin ameliorates hypophosphatasia via rescuing alkaline phosphatase deficiency-mediated L-type Ca2+ channel internalization in mesenchymal stem cells

    Journal: Bone Research

    doi: 10.1038/s41413-020-0090-7

    ALPL deficiency promoted the internalization of L-type Ca 2+ channels in HPP patient-derived BMSCs. a The expression of ALPL on the membrane and cytoplasm was decreased in BMSCs from two HPP patients compared with that of normal human BMSCs. β-actin was used as a protein loading control. b Intracellular Ca 2+ imaging analysis showed that KCl-induced Ca 2+ influx was significantly decreased in cultured BMSCs from HPP patients compared with that of normal human BMSCs. c Overexpression of ALPL or transfection with DN-Dyn1 in BMSCs from A1 patients showed elevated KCl-induced Ca 2+ influx. d Overexpression of ALPL or transfection with DN-Dyn1 in BMSCs from A2 patients showed elevated KCl-induced Ca 2+ influx. e Representative images of confocal laser scanning microscopy showing the membrane location of Ca V 1.2 and Ca V 1.3 in control BMSCs, A1 BMSCs, A2 BMSCs, A1 and A2 BMSCs overexpressing ALPL, and A1 and A2 BMSCs transfected with DN-Dyn1. Scale bar, 10 μm. The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. * P
    Figure Legend Snippet: ALPL deficiency promoted the internalization of L-type Ca 2+ channels in HPP patient-derived BMSCs. a The expression of ALPL on the membrane and cytoplasm was decreased in BMSCs from two HPP patients compared with that of normal human BMSCs. β-actin was used as a protein loading control. b Intracellular Ca 2+ imaging analysis showed that KCl-induced Ca 2+ influx was significantly decreased in cultured BMSCs from HPP patients compared with that of normal human BMSCs. c Overexpression of ALPL or transfection with DN-Dyn1 in BMSCs from A1 patients showed elevated KCl-induced Ca 2+ influx. d Overexpression of ALPL or transfection with DN-Dyn1 in BMSCs from A2 patients showed elevated KCl-induced Ca 2+ influx. e Representative images of confocal laser scanning microscopy showing the membrane location of Ca V 1.2 and Ca V 1.3 in control BMSCs, A1 BMSCs, A2 BMSCs, A1 and A2 BMSCs overexpressing ALPL, and A1 and A2 BMSCs transfected with DN-Dyn1. Scale bar, 10 μm. The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. * P

    Techniques Used: Derivative Assay, Expressing, Imaging, Cell Culture, Over Expression, Transfection, Confocal Laser Scanning Microscopy

    ALPL deficiency caused decreased membrane expression of L-type Ca 2+ channels in BMSCs. a Ca 2+ imaging showed decreased Ca 2+ influx in cultured alpl +/− BMSCs and WT BMSCs transfected with shALP (shALP/WT) after they were stimulated with 30 mmol·L −1 KCl for 3 min ( n = 10). b No KCl-induced Ca 2+ influx was detected in cultured WT, alpl +/− , and shALP/WT BMSCs treated with 10 mmol·L −1 EGTA for 3 min ( n = 10). c ALPL overexpression was mediated by a lentivirus in alpl +/− (Lenti-alp/ alpl +/− ) BMSCs and resulted in an elevated Ca 2+ influx following stimulation with 30 mmol·L −1 KCl for 3 min ( n = 10). d , e The expression of Ca V 1.1, Ca V 1.2, and Ca V 1.3 was assessed. alpl +/− BMSCs showed decreases in total cell expression ( d ) and membrane expression of Ca V 1.2 and Ca V 1.3 ( e ) and no significant change in the levels of cytoplasmic Ca V 1.2 and Ca V 1.3 ( e ). Total Ca V 1.1 protein expression was not changed ( d ), and the expression of membrane and cytoplasmic Ca V 1.1 was not altered in alpl +/− BMSCs ( e ). f Cell-surface biotinylation assay. Left two lanes: western blot for Ca V 1.2 and Ca V 1.3 following neutravidin pull down from WT and alpl +/− BMSCs; right two lanes: input, not biotinylated cells. g Lenti-alp/ alpl +/− BMSCs showed elevated membrane expression of ALP, Ca V 1.2, and Ca V 1.3. h , i Representative images of confocal laser scanning microscopy showing the membrane location of Ca V 1.2 and Ca V 1.3 (green) in WT and Lenti-alp/ alpl +/− BMSCs. The plasma membrane was stained with the marker CellMask™ Deep Red Plasma Membrane Stain (red) ( h ). Quantification of the membrane florescence was performed with NIH ImageJ ( i ). Scale bar, 10 μm. The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. * P
    Figure Legend Snippet: ALPL deficiency caused decreased membrane expression of L-type Ca 2+ channels in BMSCs. a Ca 2+ imaging showed decreased Ca 2+ influx in cultured alpl +/− BMSCs and WT BMSCs transfected with shALP (shALP/WT) after they were stimulated with 30 mmol·L −1 KCl for 3 min ( n = 10). b No KCl-induced Ca 2+ influx was detected in cultured WT, alpl +/− , and shALP/WT BMSCs treated with 10 mmol·L −1 EGTA for 3 min ( n = 10). c ALPL overexpression was mediated by a lentivirus in alpl +/− (Lenti-alp/ alpl +/− ) BMSCs and resulted in an elevated Ca 2+ influx following stimulation with 30 mmol·L −1 KCl for 3 min ( n = 10). d , e The expression of Ca V 1.1, Ca V 1.2, and Ca V 1.3 was assessed. alpl +/− BMSCs showed decreases in total cell expression ( d ) and membrane expression of Ca V 1.2 and Ca V 1.3 ( e ) and no significant change in the levels of cytoplasmic Ca V 1.2 and Ca V 1.3 ( e ). Total Ca V 1.1 protein expression was not changed ( d ), and the expression of membrane and cytoplasmic Ca V 1.1 was not altered in alpl +/− BMSCs ( e ). f Cell-surface biotinylation assay. Left two lanes: western blot for Ca V 1.2 and Ca V 1.3 following neutravidin pull down from WT and alpl +/− BMSCs; right two lanes: input, not biotinylated cells. g Lenti-alp/ alpl +/− BMSCs showed elevated membrane expression of ALP, Ca V 1.2, and Ca V 1.3. h , i Representative images of confocal laser scanning microscopy showing the membrane location of Ca V 1.2 and Ca V 1.3 (green) in WT and Lenti-alp/ alpl +/− BMSCs. The plasma membrane was stained with the marker CellMask™ Deep Red Plasma Membrane Stain (red) ( h ). Quantification of the membrane florescence was performed with NIH ImageJ ( i ). Scale bar, 10 μm. The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. * P

    Techniques Used: Expressing, Imaging, Cell Culture, Transfection, Over Expression, Cell Surface Biotinylation Assay, Western Blot, Confocal Laser Scanning Microscopy, Staining, Marker

    ALPL-maintained MSC osteogenic/adipogenic lineage differentiation ability via the L-type Ca 2+ channel. a Ca 2+ imaging showed elevated Ca 2+ influx in alpl +/− BMSCs transfected with oeCa V 1.2 or oeCa V 1.3 following stimulation with 30 mmol·L −1 KCl for 3 min ( n = 10). b , c Representative images of confocal laser scanning microscopy showing the membrane location of Ca V 1.2 and Ca V 1.3 (green) in alpl +/− BMSCs transfected with oeCa V 1.2 or oeCa V 1.3. The plasma membrane was stained with the marker CellMask™ Deep Red Plasma Membrane Stain (red) ( b ). Quantification of the membrane florescence was performed with NIH ImageJ ( c ). Scale bar, 10 μm. Alizarin red staining showed that alpl +/− BMSCs transfected with oeCa V 1.2 or oeCa V 1.3 had an increased capacity to form mineralized nodules when cultured under osteoinductive conditions ( d ) and they exhibited an upregulation of the osteogenic-related proteins RUNX2 and Sp7 ( e ). oeCa V 1.2- or oeCa V 1.3-treated alpl +/− BMSCs showed a decreased number of oil red O-positive adipocytes when cultured under adipo-inductive conditions ( f ) and there was a downregulation of the adipogenic-related proteins PPARγ2 and LPL, as assessed by western blot ( g ). h Alizarin red staining showed that alpl +/− BMSCs transfected with siCa V 1.2 or siCa V 1.3 had a decreased capacity to form mineralized nodules when cultured under osteoinductive conditions. i Western blot analysis showed that BMSCs transfected with siCa V 1.2 or siCa V 1.3 expressed decreased levels of the osteogenic-related proteins RUNX2 and Sp7. β-actin was used as a protein loading control. BMSCs transfected with siCa V 1.2 or siCa V 1.3 showed an increased number of oil red O-positive adipocytes when cultured under adipo-inductive conditions ( j ) and upregulation of the adipogenic-related proteins PPARγ2 and LPL, as assessed by western blot ( k ). The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. * P
    Figure Legend Snippet: ALPL-maintained MSC osteogenic/adipogenic lineage differentiation ability via the L-type Ca 2+ channel. a Ca 2+ imaging showed elevated Ca 2+ influx in alpl +/− BMSCs transfected with oeCa V 1.2 or oeCa V 1.3 following stimulation with 30 mmol·L −1 KCl for 3 min ( n = 10). b , c Representative images of confocal laser scanning microscopy showing the membrane location of Ca V 1.2 and Ca V 1.3 (green) in alpl +/− BMSCs transfected with oeCa V 1.2 or oeCa V 1.3. The plasma membrane was stained with the marker CellMask™ Deep Red Plasma Membrane Stain (red) ( b ). Quantification of the membrane florescence was performed with NIH ImageJ ( c ). Scale bar, 10 μm. Alizarin red staining showed that alpl +/− BMSCs transfected with oeCa V 1.2 or oeCa V 1.3 had an increased capacity to form mineralized nodules when cultured under osteoinductive conditions ( d ) and they exhibited an upregulation of the osteogenic-related proteins RUNX2 and Sp7 ( e ). oeCa V 1.2- or oeCa V 1.3-treated alpl +/− BMSCs showed a decreased number of oil red O-positive adipocytes when cultured under adipo-inductive conditions ( f ) and there was a downregulation of the adipogenic-related proteins PPARγ2 and LPL, as assessed by western blot ( g ). h Alizarin red staining showed that alpl +/− BMSCs transfected with siCa V 1.2 or siCa V 1.3 had a decreased capacity to form mineralized nodules when cultured under osteoinductive conditions. i Western blot analysis showed that BMSCs transfected with siCa V 1.2 or siCa V 1.3 expressed decreased levels of the osteogenic-related proteins RUNX2 and Sp7. β-actin was used as a protein loading control. BMSCs transfected with siCa V 1.2 or siCa V 1.3 showed an increased number of oil red O-positive adipocytes when cultured under adipo-inductive conditions ( j ) and upregulation of the adipogenic-related proteins PPARγ2 and LPL, as assessed by western blot ( k ). The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. * P

    Techniques Used: Imaging, Transfection, Confocal Laser Scanning Microscopy, Staining, Marker, Cell Culture, Western Blot

    ALPL deficiency promoted the internalization of L-type Ca 2+ channels via binding to α2δ subunits. a Representative images of confocal laser scanning microscopy showing a region of membrane colocalization for ALPL (DsRed) and Ca V 1.2 (FITC-labeled) or Ca V 1.3 (FITC-labeled) in WT BMSCs. No region of membrane colocalization was found in alpl +/− BMSCs. Scale bar, 10 μm. b ALPL immunoprecipitated Ca V 1.2 and Ca V 1.3. The left lane shows the expression of Ca V 1.2 and Ca V 1.3, and the right lane shows the levels of Ca V 1.2 and Ca V 1.3 following immunoprecipitation with an anti-ALPL antibody. c Ca V 1.2 and Ca V 1.3 immunoprecipitated ALPL. The left panel shows the expression of ALPL, and the right panel shows the level of ALPL following immunoprecipitation with anti-Ca V 1.2 or anti-Ca V 1.3 antibodies. d Representative images of confocal laser scanning microscopy showing the membrane colocalization region of ALPL (Cy3-labeled) and Ca V 1.2 (FITC-labeled) or Ca V 1.3 (FITC-labeled) in alpl −/− BMSCs overexpressing ALPL and the α2δ subunit. No membrane colocalization region was found in alpl −/− BMSCs and alpl −/− BMSCs overexpressing ALPL or the mutant α2δ subunit. Scale bar, 10 μm. e Western blot analysis showed membrane expression of Ca V 1.2 or Ca V 1.3 in alpl −/− BMSCs overexpressing ALPL and the α2δ subunit. No membrane expression of Ca V 1.2 or Ca V 1.3 was found in alpl −/− BMSCs and alpl −/− BMSCs overexpressing ALPL or the mutant α2δ subunit. No significant change in cytoplasmic Ca V 1.2 or Ca V 1.3 was found in alpl −/− BMSCs, alpl −/− BMSCs overexpressing ALPL and the mutant α2δ subunit, or alpl −/− BMSCs overexpressing ALPL and the α2δ subunit. f α2δ immunoprecipitated ALPL. The left panel shows the expression of ALPL, and the right panel shows the level of ALPL following immunoprecipitation with an anti-α2δ antibody in alp/ f/f and alpl −/− BMSCs. β-actin was used as a protein loading control. The representative results from three independent experiments are shown
    Figure Legend Snippet: ALPL deficiency promoted the internalization of L-type Ca 2+ channels via binding to α2δ subunits. a Representative images of confocal laser scanning microscopy showing a region of membrane colocalization for ALPL (DsRed) and Ca V 1.2 (FITC-labeled) or Ca V 1.3 (FITC-labeled) in WT BMSCs. No region of membrane colocalization was found in alpl +/− BMSCs. Scale bar, 10 μm. b ALPL immunoprecipitated Ca V 1.2 and Ca V 1.3. The left lane shows the expression of Ca V 1.2 and Ca V 1.3, and the right lane shows the levels of Ca V 1.2 and Ca V 1.3 following immunoprecipitation with an anti-ALPL antibody. c Ca V 1.2 and Ca V 1.3 immunoprecipitated ALPL. The left panel shows the expression of ALPL, and the right panel shows the level of ALPL following immunoprecipitation with anti-Ca V 1.2 or anti-Ca V 1.3 antibodies. d Representative images of confocal laser scanning microscopy showing the membrane colocalization region of ALPL (Cy3-labeled) and Ca V 1.2 (FITC-labeled) or Ca V 1.3 (FITC-labeled) in alpl −/− BMSCs overexpressing ALPL and the α2δ subunit. No membrane colocalization region was found in alpl −/− BMSCs and alpl −/− BMSCs overexpressing ALPL or the mutant α2δ subunit. Scale bar, 10 μm. e Western blot analysis showed membrane expression of Ca V 1.2 or Ca V 1.3 in alpl −/− BMSCs overexpressing ALPL and the α2δ subunit. No membrane expression of Ca V 1.2 or Ca V 1.3 was found in alpl −/− BMSCs and alpl −/− BMSCs overexpressing ALPL or the mutant α2δ subunit. No significant change in cytoplasmic Ca V 1.2 or Ca V 1.3 was found in alpl −/− BMSCs, alpl −/− BMSCs overexpressing ALPL and the mutant α2δ subunit, or alpl −/− BMSCs overexpressing ALPL and the α2δ subunit. f α2δ immunoprecipitated ALPL. The left panel shows the expression of ALPL, and the right panel shows the level of ALPL following immunoprecipitation with an anti-α2δ antibody in alp/ f/f and alpl −/− BMSCs. β-actin was used as a protein loading control. The representative results from three independent experiments are shown

    Techniques Used: Binding Assay, Confocal Laser Scanning Microscopy, Labeling, Immunoprecipitation, Expressing, Mutagenesis, Western Blot

    ALPL deficiency promoted the internalization of L-type Ca 2+ channels in BMSCs. a , b Representative images of confocal laser scanning microscopy showing the membrane location of Ca V 1.2 and Ca V 1.3 (green) in alpl +/− BMSCs transfected with DN-Dyn1. The plasma membrane was stained with the marker CellMask™ Deep Red Plasma Membrane Stain (red) ( a ). Quantification of the membrane florescence was performed with NIH ImageJ ( b ). Scale bar, 10 μm. c alpl +/− BMSCs transfected with DN-Dyn1 showed upregulation of Ca V 1.2 and Ca V 1.3 membrane expression and almost no change in the cytoplasmic expression of Ca V 1.2 and Ca V 1.3, as assessed by western blot. β-actin was used as a protein loading control. d 10-min time-lapse confocal laser scanning microscopy images of WT, alpl +/− , DN-Dyn1/ alpl +/− , and Lenti-alp/ alpl +/− BMSCs containing DsRed-CaV1.2. Scale bar, 10 μm. e Ca 2+ imaging showed elevated Ca 2+ influx of cultured alpl +/− BMSCs transfected with DN-Dyn1 after stimulation with 30 mmol·L −1 KCl for 3 min ( n = 10). alpl +/− BMSCs showed a pronounced decrease in DsRed-Ca V 1.2 at the membrane (Dio-labeled ROI, n = 10) ( f ). g Quantification of the florescence in the ROI during the time-course lapse at 0 s, 300 s, and 600 s. h – k Representative images show the colocalization of DsRed-Ca V 1.2 with the Dio-labeled cell membrane of BMSCs. WT, DN-Dyn1/ alpl +/− , and Lenti-alp/ alpl +/− BMSCs had colocalized regions at 0 s and 570 s ( h , j , k ). However, alpl +/− BMSCs showed no colocalization at 0 s and 570 s ( i ). The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. * P
    Figure Legend Snippet: ALPL deficiency promoted the internalization of L-type Ca 2+ channels in BMSCs. a , b Representative images of confocal laser scanning microscopy showing the membrane location of Ca V 1.2 and Ca V 1.3 (green) in alpl +/− BMSCs transfected with DN-Dyn1. The plasma membrane was stained with the marker CellMask™ Deep Red Plasma Membrane Stain (red) ( a ). Quantification of the membrane florescence was performed with NIH ImageJ ( b ). Scale bar, 10 μm. c alpl +/− BMSCs transfected with DN-Dyn1 showed upregulation of Ca V 1.2 and Ca V 1.3 membrane expression and almost no change in the cytoplasmic expression of Ca V 1.2 and Ca V 1.3, as assessed by western blot. β-actin was used as a protein loading control. d 10-min time-lapse confocal laser scanning microscopy images of WT, alpl +/− , DN-Dyn1/ alpl +/− , and Lenti-alp/ alpl +/− BMSCs containing DsRed-CaV1.2. Scale bar, 10 μm. e Ca 2+ imaging showed elevated Ca 2+ influx of cultured alpl +/− BMSCs transfected with DN-Dyn1 after stimulation with 30 mmol·L −1 KCl for 3 min ( n = 10). alpl +/− BMSCs showed a pronounced decrease in DsRed-Ca V 1.2 at the membrane (Dio-labeled ROI, n = 10) ( f ). g Quantification of the florescence in the ROI during the time-course lapse at 0 s, 300 s, and 600 s. h – k Representative images show the colocalization of DsRed-Ca V 1.2 with the Dio-labeled cell membrane of BMSCs. WT, DN-Dyn1/ alpl +/− , and Lenti-alp/ alpl +/− BMSCs had colocalized regions at 0 s and 570 s ( h , j , k ). However, alpl +/− BMSCs showed no colocalization at 0 s and 570 s ( i ). The representative results from three independent experiments are shown. Error bars represent the s.d. from the mean values. * P

    Techniques Used: Confocal Laser Scanning Microscopy, Transfection, Staining, Marker, Expressing, Western Blot, Imaging, Cell Culture, Labeling

    10) Product Images from "β1,4-Galactosyltransferase V Modulates Breast Cancer Stem Cells through Wnt/β-catenin Signaling Pathway"

    Article Title: β1,4-Galactosyltransferase V Modulates Breast Cancer Stem Cells through Wnt/β-catenin Signaling Pathway

    Journal: Cancer Research and Treatment : Official Journal of Korean Cancer Association

    doi: 10.4143/crt.2020.093

    Cell surface β1,4-galactosyltransferase V (B4GalT5) is not responsible for the stemness of breast cancer. (A) Immunofluorescence analysis of B4GalT5 expression in MCF-7ADR cells. Cells were stained with IgG or anti-B4GalT5 antibody followed by fluorescein isothiocyanate (FITC)–conjugated anti-Rabbit antibodies as described and analyzed by a laser scanning confocal microscope. (B) Cell surface biotinylation assay to compare B4GalT5 localization in plasma membrane and cytoplasmic fractions of MCF-7ADR cells. (C) Construction of MCF-7ADR cells that stably expresses B4GalT5 with a C-terminal 6×His tag. MCF-7ADR cells were transfected with pBABE-B4GalT5-His-IRES-puro plasmid for 48 hours and then selected with 150 μg/mL puromycin. Expression of corresponding proteins was examined by western blotting. (D) Representative images of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. After digested and resuspended, the cells were stained with anti-His antibody at room temperature for 1 hour followed by FITC-conjugated anti-rabbit antibody at room temperature for 1 hour. Pictures were taken by a laser scanning confocal microscope. (E, F) Flow cytometry (FCM) analysis of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. Cells were stained with phycoerythrin (PE)-conjugated anti-His antibody at room temperature for 1 hour and analyzed by Moflo XDP. Error bars represent standard error of the mean (n=3, ***p
    Figure Legend Snippet: Cell surface β1,4-galactosyltransferase V (B4GalT5) is not responsible for the stemness of breast cancer. (A) Immunofluorescence analysis of B4GalT5 expression in MCF-7ADR cells. Cells were stained with IgG or anti-B4GalT5 antibody followed by fluorescein isothiocyanate (FITC)–conjugated anti-Rabbit antibodies as described and analyzed by a laser scanning confocal microscope. (B) Cell surface biotinylation assay to compare B4GalT5 localization in plasma membrane and cytoplasmic fractions of MCF-7ADR cells. (C) Construction of MCF-7ADR cells that stably expresses B4GalT5 with a C-terminal 6×His tag. MCF-7ADR cells were transfected with pBABE-B4GalT5-His-IRES-puro plasmid for 48 hours and then selected with 150 μg/mL puromycin. Expression of corresponding proteins was examined by western blotting. (D) Representative images of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. After digested and resuspended, the cells were stained with anti-His antibody at room temperature for 1 hour followed by FITC-conjugated anti-rabbit antibody at room temperature for 1 hour. Pictures were taken by a laser scanning confocal microscope. (E, F) Flow cytometry (FCM) analysis of B4GalT5-His expression on the cell surface of MCF-7ADR/B4GalT5-His and MCF-7ADR/NC cells. Cells were stained with phycoerythrin (PE)-conjugated anti-His antibody at room temperature for 1 hour and analyzed by Moflo XDP. Error bars represent standard error of the mean (n=3, ***p

    Techniques Used: Immunofluorescence, Expressing, Staining, Microscopy, Cell Surface Biotinylation Assay, Stable Transfection, Transfection, Plasmid Preparation, Western Blot, Flow Cytometry

    β1,4-Galactosyltransferase V (B4GalT5) promotes Wnt/β-catenin signaling that is hyperactivated in breast cancer stem cells (BCSCs). (A) Immunofluorescence analysis of phosphorylated-GSK3β (Ser 9) expression in BCSCs and non-BCSCs. MCF-7ADR cells were sorted into BCSCs and non-BCSCs, then stained with antibodies and DAPI as described and analyzed by a laser scanning confocal microscope. (B, C) Western blotting analysis of β-catenin and phosphorylated-β-catenin (Ser 45) expression levels in BCSCs and non-BCSCs. Protein band densities were quantified by normalizing to β-actin. (D, E) The effect of B4GalT5 knockdown on Frizzled-1, β-catenin, and B4GalT5 expression levels in MCF-7ADR, MCF-7, and MDA-MB-231 cell lines by western blotting analysis. Protein band densities were quantified by normalizing to β-actin. (F, G) The effect of B4GalT5 overexpression on Frizzled-1, β-catenin, and B4GalT5 expression levels in MCF-7ADR, MCF-7, and MDA-MB-231 cell lines by western blotting analysis. Protein band densities were quantified by normalizing to β-Actin. (H) The effect of B4GalT5 on wnt 3α induced Wnt/β-catenin signaling by western blotting (WB). After transfected using B4GalT5 siRNA for 12 hours, MCF-7ADR cells were starved for 36 hours followed by adding wnt 3α, and subjected to western blotting. (I) The effect of B4GalT5 on Wnt 3α induced ALDH1A1 expression by western blotting. MCF-7ADR/shNC and MCF-7ADR/shB4GalT5 cells were starved for 36 hours followed by adding Wnt 3α, and subjected to western blotting. (J) The degradation pathway of β-catenin due to B4GalT5 knockdown. After transfected using B4GalT5 siRNA for 41 hours, MCF-7ADR cells were treated with MG132 (protease inhibitor) for 7 hours. Protein levels were examined by western blotting. (K) Western blotting analysis of membrane and cytosol proteins in MCF-7ADR cells. MCF-7ADR cells were transfected using B4GalT5 siRNA for 48 hours, extracted into membrane and cytosol proteins and then subjected to western blotting. Erk was used for examining purity of membrane proteins. (L) The degradation pathway of Frizzled-1 on the cell surface of MCF-7ADR cells due to B4GalT5 knockdown. After transfected using B4GalT5 siRNA for 40 hours, MCF-7ADR cells were treated with 20 μM leupeptin (lysosome inhibitor) for 8 hours. The membrane proteins were extracted as described and then subjected to western blotting. Erk was used for examining purity of membrane proteins. (M) The effect of B4GalT5 on biosynthesis of galactosyl-oligosaccharides in glycans of Frizzled-1 by RCA-I lectin pull-down assay. MCF-7ADR/shNC, MCF-7ADR/shB4GalT5, MCF-7ADR/NC, and MCF-7ADR/oe-B4GalT5 cells were lysed and incubated with agarose-bound RCA-I overnight. Total Frizzled-1 was used as loading control. All experiments were performed in three replicates. Data are presented as means±standard error of the mean. *p
    Figure Legend Snippet: β1,4-Galactosyltransferase V (B4GalT5) promotes Wnt/β-catenin signaling that is hyperactivated in breast cancer stem cells (BCSCs). (A) Immunofluorescence analysis of phosphorylated-GSK3β (Ser 9) expression in BCSCs and non-BCSCs. MCF-7ADR cells were sorted into BCSCs and non-BCSCs, then stained with antibodies and DAPI as described and analyzed by a laser scanning confocal microscope. (B, C) Western blotting analysis of β-catenin and phosphorylated-β-catenin (Ser 45) expression levels in BCSCs and non-BCSCs. Protein band densities were quantified by normalizing to β-actin. (D, E) The effect of B4GalT5 knockdown on Frizzled-1, β-catenin, and B4GalT5 expression levels in MCF-7ADR, MCF-7, and MDA-MB-231 cell lines by western blotting analysis. Protein band densities were quantified by normalizing to β-actin. (F, G) The effect of B4GalT5 overexpression on Frizzled-1, β-catenin, and B4GalT5 expression levels in MCF-7ADR, MCF-7, and MDA-MB-231 cell lines by western blotting analysis. Protein band densities were quantified by normalizing to β-Actin. (H) The effect of B4GalT5 on wnt 3α induced Wnt/β-catenin signaling by western blotting (WB). After transfected using B4GalT5 siRNA for 12 hours, MCF-7ADR cells were starved for 36 hours followed by adding wnt 3α, and subjected to western blotting. (I) The effect of B4GalT5 on Wnt 3α induced ALDH1A1 expression by western blotting. MCF-7ADR/shNC and MCF-7ADR/shB4GalT5 cells were starved for 36 hours followed by adding Wnt 3α, and subjected to western blotting. (J) The degradation pathway of β-catenin due to B4GalT5 knockdown. After transfected using B4GalT5 siRNA for 41 hours, MCF-7ADR cells were treated with MG132 (protease inhibitor) for 7 hours. Protein levels were examined by western blotting. (K) Western blotting analysis of membrane and cytosol proteins in MCF-7ADR cells. MCF-7ADR cells were transfected using B4GalT5 siRNA for 48 hours, extracted into membrane and cytosol proteins and then subjected to western blotting. Erk was used for examining purity of membrane proteins. (L) The degradation pathway of Frizzled-1 on the cell surface of MCF-7ADR cells due to B4GalT5 knockdown. After transfected using B4GalT5 siRNA for 40 hours, MCF-7ADR cells were treated with 20 μM leupeptin (lysosome inhibitor) for 8 hours. The membrane proteins were extracted as described and then subjected to western blotting. Erk was used for examining purity of membrane proteins. (M) The effect of B4GalT5 on biosynthesis of galactosyl-oligosaccharides in glycans of Frizzled-1 by RCA-I lectin pull-down assay. MCF-7ADR/shNC, MCF-7ADR/shB4GalT5, MCF-7ADR/NC, and MCF-7ADR/oe-B4GalT5 cells were lysed and incubated with agarose-bound RCA-I overnight. Total Frizzled-1 was used as loading control. All experiments were performed in three replicates. Data are presented as means±standard error of the mean. *p

    Techniques Used: Immunofluorescence, Expressing, Staining, Microscopy, Western Blot, Multiple Displacement Amplification, Over Expression, Transfection, Protease Inhibitor, Pull Down Assay, Incubation

    11) Product Images from "Transcriptional Control of Brain Tumour Stem Cell Fate by a Carbohydrate Binding Protein"

    Article Title: Transcriptional Control of Brain Tumour Stem Cell Fate by a Carbohydrate Binding Protein

    Journal: bioRxiv

    doi: 10.1101/2021.04.14.439704

    Galectin1 interacts with HOXA5 endogenously in EGFRvIII-expressing BTSCs. ( a ) LGALS1 -differentially genes from RNA-seq analysis were subjected to enrichment analysis of transcription factor (TF) binding motifs using oPOSSUM-3 software. High-scoring or over-represented TF binding site profiles were computed as having z-scores above the mean + 2 x standard deviation (red dotted line). ( b ) Volcano plot representing the HOXA5 targets genes among the LGALS1 -differentially-regulated genes is shown. LGALS1 -differentially regulated genes that possess HOXA5 binding sites (HOXA5 targets) are reported as blue dots, LGALS1 -differentially-regulated genes (DEGs) that do not possess HOXA5 binding sites are reported as red dots and the non-differentially regulated genes (Non DEGs) are represented as black dots. Black lines show RT-qPCR validated cell cycle related genes. ( c ) Relative positions of HOXA5 peaks, obtained from ChIP-seq analysis in human carcinoma cells, to the adjacent TSS of LGALS1 -differentially regulated genes from RNA-seq analysis are shown. The x-axis indicates the distance between peak centers and the TSS of adjacent LGALS1 -differentially regulated genes. The y-axis denotes the expression ratios (log2) of the LGALS1 -differentially regulated gene. Circle size indicates HOXA5 peak height, and color denotes the conservation score of HOXA5 peaks. ( d ) BTSCs were analyzed by immunoblotting using the antibodies indicated on the blots. Wild type EGFR and EGFRvIII bands are marked with * and **, respectively. ( e ) Correlation of HOXA5 expression with galectin1 expression was obtained by running Pearson analyses on the densitometric values of protein expression normalized to tubulin. ( f-g ) BTSCs were electroporated with siCTL or siRNA against HOXA5 (si HOXA5 ). mRNA levels of the LGALS1 -downregulated genes that possess HOXA5 motifs were evaluated by RT-qPCR. BTSC73 ( f ): *** p = 0.0001 for each pairwise comparison; BTSC147 ( g ): *** p = 0.0001 for each pairwise comparison except: ** p E2F7 = 0.0014, ** p NDC80 = 0.0099; One-way ANOVA followed by Dunnett’s test, n = 3. Data are presented as the mean ± SEM. ( h ) Representative phase-contrast images of BTSC73 electroporated with si HOXA5 or siCTL in the absence and presence of 4 Gy are shown. Images were taken following 7 days of plating. Scale bar = 100 µm. ( i ) ELDA was performed following 4 Gy of IR in si HOXA5 vs. siCTL. ( j - m ) Whole cell lysates from BTSC73 ( j ), BTSC147 ( k ) BTSC68 ( l ) and BTSC172 ( m ) were subjected to immunoprecipitation using an antibody against HOXA5 or rabbit IgG control, followed by immunoblotting with galectin1 and HOXA5 antibodies. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC. ( n - q ) Proximity ligation assay (PLA) of galectin1 and HOXA5 were performed in BTSC147 ( n ), BTSC68 ( o ), BTSC172 ( p ) and BTSC73 ( q ). Primary antibodies were omitted for the controls. PLA was performed in LGALS1 CRISPR in ( q ) as an additional control. Nuclei were stained with DAPI. Images were obtained with a 63X objectives on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm. Representative images of three independent experiments are shown. ( r ) LGALS1 CRISPR and CTL BTSC73 were subjected to ChIP using an antibody to HOXA5 or IgG control followed by RT-qPCR for four LGALS1 -downregulated genes that possess HOXA5 binding motifs. HBB locus was used as a negative control. *** p ACSS3 (HOXA5 IP in CTL vs. IgG IP in CTL) = 0.0007, ** p ACSS3 (HOXA5 IP in LGALS1 CRISPR vs. HOXA5 IP in CTL) = 0.0036, *** p ARIDA5B (HOXA5 IP in CTL vs. IgG IP in CTL)
    Figure Legend Snippet: Galectin1 interacts with HOXA5 endogenously in EGFRvIII-expressing BTSCs. ( a ) LGALS1 -differentially genes from RNA-seq analysis were subjected to enrichment analysis of transcription factor (TF) binding motifs using oPOSSUM-3 software. High-scoring or over-represented TF binding site profiles were computed as having z-scores above the mean + 2 x standard deviation (red dotted line). ( b ) Volcano plot representing the HOXA5 targets genes among the LGALS1 -differentially-regulated genes is shown. LGALS1 -differentially regulated genes that possess HOXA5 binding sites (HOXA5 targets) are reported as blue dots, LGALS1 -differentially-regulated genes (DEGs) that do not possess HOXA5 binding sites are reported as red dots and the non-differentially regulated genes (Non DEGs) are represented as black dots. Black lines show RT-qPCR validated cell cycle related genes. ( c ) Relative positions of HOXA5 peaks, obtained from ChIP-seq analysis in human carcinoma cells, to the adjacent TSS of LGALS1 -differentially regulated genes from RNA-seq analysis are shown. The x-axis indicates the distance between peak centers and the TSS of adjacent LGALS1 -differentially regulated genes. The y-axis denotes the expression ratios (log2) of the LGALS1 -differentially regulated gene. Circle size indicates HOXA5 peak height, and color denotes the conservation score of HOXA5 peaks. ( d ) BTSCs were analyzed by immunoblotting using the antibodies indicated on the blots. Wild type EGFR and EGFRvIII bands are marked with * and **, respectively. ( e ) Correlation of HOXA5 expression with galectin1 expression was obtained by running Pearson analyses on the densitometric values of protein expression normalized to tubulin. ( f-g ) BTSCs were electroporated with siCTL or siRNA against HOXA5 (si HOXA5 ). mRNA levels of the LGALS1 -downregulated genes that possess HOXA5 motifs were evaluated by RT-qPCR. BTSC73 ( f ): *** p = 0.0001 for each pairwise comparison; BTSC147 ( g ): *** p = 0.0001 for each pairwise comparison except: ** p E2F7 = 0.0014, ** p NDC80 = 0.0099; One-way ANOVA followed by Dunnett’s test, n = 3. Data are presented as the mean ± SEM. ( h ) Representative phase-contrast images of BTSC73 electroporated with si HOXA5 or siCTL in the absence and presence of 4 Gy are shown. Images were taken following 7 days of plating. Scale bar = 100 µm. ( i ) ELDA was performed following 4 Gy of IR in si HOXA5 vs. siCTL. ( j - m ) Whole cell lysates from BTSC73 ( j ), BTSC147 ( k ) BTSC68 ( l ) and BTSC172 ( m ) were subjected to immunoprecipitation using an antibody against HOXA5 or rabbit IgG control, followed by immunoblotting with galectin1 and HOXA5 antibodies. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC. ( n - q ) Proximity ligation assay (PLA) of galectin1 and HOXA5 were performed in BTSC147 ( n ), BTSC68 ( o ), BTSC172 ( p ) and BTSC73 ( q ). Primary antibodies were omitted for the controls. PLA was performed in LGALS1 CRISPR in ( q ) as an additional control. Nuclei were stained with DAPI. Images were obtained with a 63X objectives on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm. Representative images of three independent experiments are shown. ( r ) LGALS1 CRISPR and CTL BTSC73 were subjected to ChIP using an antibody to HOXA5 or IgG control followed by RT-qPCR for four LGALS1 -downregulated genes that possess HOXA5 binding motifs. HBB locus was used as a negative control. *** p ACSS3 (HOXA5 IP in CTL vs. IgG IP in CTL) = 0.0007, ** p ACSS3 (HOXA5 IP in LGALS1 CRISPR vs. HOXA5 IP in CTL) = 0.0036, *** p ARIDA5B (HOXA5 IP in CTL vs. IgG IP in CTL)

    Techniques Used: Expressing, RNA Sequencing Assay, Binding Assay, Software, Standard Deviation, Quantitative RT-PCR, Chromatin Immunoprecipitation, Immunoprecipitation, Western Blot, Proximity Ligation Assay, CRISPR, Staining, Microscopy, Negative Control

    Galectin1 is a direct transcriptional target of EGFRvIII/STAT3 in patient-derived BTSCs. ( a ) Patient-derived BTSCs that harbour EGFRvIII mutation or lack the mutation were analyzed by immunoblotting using the antibodies indicated on the blots. Wild type EGFR and EGFRvIII bands are marked with * and **, respectively. Tubulin was used as a loading control. ( b-c ) KD of EGFR / EGFRvIII in BTSCs was induced by electroporation using siRNA. KD and control BTSCs (siCTL) were analyzed by immunoblotting as described in a. ( d-g ) BTSCs were treated with 1-5 µM lapatinib and galectin1 expression were assessed by immunoblotting (d-e) and immunostaining (f-g). Nuclei were stained with DAPI. Images were obtained with a 63X objectives on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm. ( h ) Putative STAT3 binding sites at positions -300, -135 and -72 bp upstream of the LGALS1 gene transcriptional start site (TSS) are shown. ( i ) BTSCs were analyzed by immunoblotting using the antibodies indicated on the blots as described in a. ( j-k ) KD of STAT3 was induced in BTSC73 ( j ) and BTSC147 ( k ) via electroporation using siRNAs. STAT3 KD and control BTSCs were analyzed by immunoblottig as described above. ( l-o ) BTSCs were subjected to immunoblotting or immunostaining following treatment with 25-50 µM of the STAT3 inhibitor, S3I-201. Images were captured as described in f-g. Scale bar = 10 μm. ( p-r ) EGFRvIII-expressing BTSCs were subjected to ChIP using an antibody to STAT3 or IgG control followed by RT-qPCR for LGALS1 promotor region using two different pairs of primers ( LGALS1 -a and LGALS1 -b). OSMR , and HPRT loci were used as positive and negative controls, respectively. BTSC73 ( p ): * p OSMR = 0.0288, ** p LGALS1 -a = 0.0079, ** p LGALS1 -b = 0.0013; Unpaired two-tailed t -test, n = 3; BTSC172 ( q ): *** p OSMR = 0.0002, * p LGALS1 -a = 0.0168, * p LGALS1 -b = 0.0102; Unpaired two-tailed t -test, n = 3; BTSC112 ( r ): * p OSMR = 0.0450, *** p LGALS1 -a = 0.0007, * p LGALS1 -b = 0.0256; Unpaired two-tailed t -test, n = 3. ( s ) STAT3 KD and control BTSCs were electroporated with a luciferase reporter plasmid driven by a promoter containing 376 bp region upstream of the LGALS1 gene TSS ( LGALS1 pGL4.23) or the control pGL4.23-basic reporter plasmid (pGL4.23) together with a Renilla expression plasmid and were subjected to dual luciferase assay. *** p
    Figure Legend Snippet: Galectin1 is a direct transcriptional target of EGFRvIII/STAT3 in patient-derived BTSCs. ( a ) Patient-derived BTSCs that harbour EGFRvIII mutation or lack the mutation were analyzed by immunoblotting using the antibodies indicated on the blots. Wild type EGFR and EGFRvIII bands are marked with * and **, respectively. Tubulin was used as a loading control. ( b-c ) KD of EGFR / EGFRvIII in BTSCs was induced by electroporation using siRNA. KD and control BTSCs (siCTL) were analyzed by immunoblotting as described in a. ( d-g ) BTSCs were treated with 1-5 µM lapatinib and galectin1 expression were assessed by immunoblotting (d-e) and immunostaining (f-g). Nuclei were stained with DAPI. Images were obtained with a 63X objectives on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm. ( h ) Putative STAT3 binding sites at positions -300, -135 and -72 bp upstream of the LGALS1 gene transcriptional start site (TSS) are shown. ( i ) BTSCs were analyzed by immunoblotting using the antibodies indicated on the blots as described in a. ( j-k ) KD of STAT3 was induced in BTSC73 ( j ) and BTSC147 ( k ) via electroporation using siRNAs. STAT3 KD and control BTSCs were analyzed by immunoblottig as described above. ( l-o ) BTSCs were subjected to immunoblotting or immunostaining following treatment with 25-50 µM of the STAT3 inhibitor, S3I-201. Images were captured as described in f-g. Scale bar = 10 μm. ( p-r ) EGFRvIII-expressing BTSCs were subjected to ChIP using an antibody to STAT3 or IgG control followed by RT-qPCR for LGALS1 promotor region using two different pairs of primers ( LGALS1 -a and LGALS1 -b). OSMR , and HPRT loci were used as positive and negative controls, respectively. BTSC73 ( p ): * p OSMR = 0.0288, ** p LGALS1 -a = 0.0079, ** p LGALS1 -b = 0.0013; Unpaired two-tailed t -test, n = 3; BTSC172 ( q ): *** p OSMR = 0.0002, * p LGALS1 -a = 0.0168, * p LGALS1 -b = 0.0102; Unpaired two-tailed t -test, n = 3; BTSC112 ( r ): * p OSMR = 0.0450, *** p LGALS1 -a = 0.0007, * p LGALS1 -b = 0.0256; Unpaired two-tailed t -test, n = 3. ( s ) STAT3 KD and control BTSCs were electroporated with a luciferase reporter plasmid driven by a promoter containing 376 bp region upstream of the LGALS1 gene TSS ( LGALS1 pGL4.23) or the control pGL4.23-basic reporter plasmid (pGL4.23) together with a Renilla expression plasmid and were subjected to dual luciferase assay. *** p

    Techniques Used: Derivative Assay, Mutagenesis, Electroporation, Expressing, Immunostaining, Staining, Microscopy, Binding Assay, Chromatin Immunoprecipitation, Quantitative RT-PCR, Two Tailed Test, Luciferase, Plasmid Preparation

    12) Product Images from "A Fish Leukocyte Immune-Type Receptor Uses a Novel Intracytoplasmic Tail Networking Mechanism to Cross-Inhibit the Phagocytic Response"

    Article Title: A Fish Leukocyte Immune-Type Receptor Uses a Novel Intracytoplasmic Tail Networking Mechanism to Cross-Inhibit the Phagocytic Response

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms21145146

    Confocal microscopic analysis of phosphotyrosine staining levels within phagocytic cups. 2.6b ITAM CYT /1.1b WT CYT co-expressing AD293 cells ( A ) were grown on coverslips and incubated with 4.5 µm non-fluorescent (NF) beads opsonized with α-HA mAb and mouse isotype IgG1. After 8 min of incubation at 37 °C, cells were fixed with 4% PFA for 10 min and non-phagocytosed beads were stained using Alexa 647-conjugated goat-α-mouse secondary pAb (red). Cells were then permeabilized and specifically stained for intracellular phosphotyrosine molecules by first incubating them with a rabbit α-phosphotyrosine mAb and then with a secondary goat-α-rabbit pAb conjugated to Alexa 488 (green). Z-stack images were obtained at a magnification of 63X using a Zeiss LSM 710 scanning confocal microscope. Representative images from z-stack acquisitions are shown as images with phosphotyrosine (green) staining, surface-exposed bead (red) staining, and merged-fluorescence images or brightfield-fluorescence merged images. Yellow arrowheads show the positions of representative phagocytic cups and the target bead of interest is indicated by an asterisk (*). ( B ) Qualitative analysis of bead and phosphotyrosine molecule staining intensities was performed using ImageJ software (NIH, Bathesda, Maryland, DC, USA) by calculating the MFI (y-axis) of the bead (red line) and phosphotyrosine molecule staining (green line) across the dash arrowed line. ( C ) To further quantify the phosphotyrosine molecules recruited to phagocytic cups under different activation conditions, 2.6b ITAM CYT /1.1b WT CYT co-expressing cells were incubated with NF beads opsonized with indicated mAbs (’+’ and ‘−’ indicate the presence and absence of corresponding mAbs, respectively). For analyses, a region of interest (ROI, indicated by dashed circles) was first drawn that includes phagocytic cups so phosphotyrosine signal intensities in this area could be calculated. As summarized in ( D ), at least 50 phagocytic cups from three independent experiments were pooled and the data is represented as mean integrated fluorescent intensity ± SEM. Differing letters indicate statistical significance ( p ≤ 0.05) between means. Experimental groups were compared using a one-way ANOVA, followed by the Tukey test using Prism 6 software (GraphPad Software, La Jolla, CA, USA).
    Figure Legend Snippet: Confocal microscopic analysis of phosphotyrosine staining levels within phagocytic cups. 2.6b ITAM CYT /1.1b WT CYT co-expressing AD293 cells ( A ) were grown on coverslips and incubated with 4.5 µm non-fluorescent (NF) beads opsonized with α-HA mAb and mouse isotype IgG1. After 8 min of incubation at 37 °C, cells were fixed with 4% PFA for 10 min and non-phagocytosed beads were stained using Alexa 647-conjugated goat-α-mouse secondary pAb (red). Cells were then permeabilized and specifically stained for intracellular phosphotyrosine molecules by first incubating them with a rabbit α-phosphotyrosine mAb and then with a secondary goat-α-rabbit pAb conjugated to Alexa 488 (green). Z-stack images were obtained at a magnification of 63X using a Zeiss LSM 710 scanning confocal microscope. Representative images from z-stack acquisitions are shown as images with phosphotyrosine (green) staining, surface-exposed bead (red) staining, and merged-fluorescence images or brightfield-fluorescence merged images. Yellow arrowheads show the positions of representative phagocytic cups and the target bead of interest is indicated by an asterisk (*). ( B ) Qualitative analysis of bead and phosphotyrosine molecule staining intensities was performed using ImageJ software (NIH, Bathesda, Maryland, DC, USA) by calculating the MFI (y-axis) of the bead (red line) and phosphotyrosine molecule staining (green line) across the dash arrowed line. ( C ) To further quantify the phosphotyrosine molecules recruited to phagocytic cups under different activation conditions, 2.6b ITAM CYT /1.1b WT CYT co-expressing cells were incubated with NF beads opsonized with indicated mAbs (’+’ and ‘−’ indicate the presence and absence of corresponding mAbs, respectively). For analyses, a region of interest (ROI, indicated by dashed circles) was first drawn that includes phagocytic cups so phosphotyrosine signal intensities in this area could be calculated. As summarized in ( D ), at least 50 phagocytic cups from three independent experiments were pooled and the data is represented as mean integrated fluorescent intensity ± SEM. Differing letters indicate statistical significance ( p ≤ 0.05) between means. Experimental groups were compared using a one-way ANOVA, followed by the Tukey test using Prism 6 software (GraphPad Software, La Jolla, CA, USA).

    Techniques Used: Staining, Expressing, Incubation, Microscopy, Fluorescence, Software, Activation Assay

    13) Product Images from "SOX10-Cre-Labeled Cells Under the Tongue Epithelium Serve as Progenitors for Taste Bud Cells That Are Mainly Type III and Keratin 8-Low"

    Article Title: SOX10-Cre-Labeled Cells Under the Tongue Epithelium Serve as Progenitors for Taste Bud Cells That Are Mainly Type III and Keratin 8-Low

    Journal: Stem Cells and Development

    doi: 10.1089/scd.2020.0022

    SOX10-Cre/tdT -labeled cells in taste buds were rare in newborn and abundant in adult mice. (A–D) Single-plane laser scanning confocal images of tissue sections of the soft palate and fungiform, foliate, and circumvallate papillae in newborn (A) , 2-week- (B) , 4-week- (C) , 16-week- (D) old SOX10-Cre/tdT mice. Taste buds were Krt8 + ( green ). White dashed lines demarcate the lingual epithelium from the underlying connective tissue. Scale bars: 50 μm for all images. (E) Histograms ( X ± SD, n = 3) to illustrate the percentages of SOX10-Cre/tdT -labeled tdT + relative to total taste bud cells in the circumvallate papilla in newborn, 2-, 4-, 8-, and 16-week-old mice. The diamond dots represent the values of individual samples. ** P
    Figure Legend Snippet: SOX10-Cre/tdT -labeled cells in taste buds were rare in newborn and abundant in adult mice. (A–D) Single-plane laser scanning confocal images of tissue sections of the soft palate and fungiform, foliate, and circumvallate papillae in newborn (A) , 2-week- (B) , 4-week- (C) , 16-week- (D) old SOX10-Cre/tdT mice. Taste buds were Krt8 + ( green ). White dashed lines demarcate the lingual epithelium from the underlying connective tissue. Scale bars: 50 μm for all images. (E) Histograms ( X ± SD, n = 3) to illustrate the percentages of SOX10-Cre/tdT -labeled tdT + relative to total taste bud cells in the circumvallate papilla in newborn, 2-, 4-, 8-, and 16-week-old mice. The diamond dots represent the values of individual samples. ** P

    Techniques Used: Labeling, Mouse Assay

    SOX10-Cre/tdT (tdTomato)-labeled taste bud cells in all examined tissue regions, soft palate (A) and taste papillae, that is, fungiform (B) , foliate (C) , and circumvallate (D) , in young adult (8-week old) SOX10-Cre/tdT mice. Krt8 ( green ) indicates presence of taste buds. White dashed lines demarcate lingual epithelium from the underlying connective tissue. Green dashed lines in (A, B) bracket a taste bud. Arrowheads point to tdT + taste bud cells. Scale bars: 50 μm for all images (single-plane laser scanning confocal). Krt8, Keratin 8; SOX10 , SRY-related HMG-box gene 10.
    Figure Legend Snippet: SOX10-Cre/tdT (tdTomato)-labeled taste bud cells in all examined tissue regions, soft palate (A) and taste papillae, that is, fungiform (B) , foliate (C) , and circumvallate (D) , in young adult (8-week old) SOX10-Cre/tdT mice. Krt8 ( green ) indicates presence of taste buds. White dashed lines demarcate lingual epithelium from the underlying connective tissue. Green dashed lines in (A, B) bracket a taste bud. Arrowheads point to tdT + taste bud cells. Scale bars: 50 μm for all images (single-plane laser scanning confocal). Krt8, Keratin 8; SOX10 , SRY-related HMG-box gene 10.

    Techniques Used: Labeling, Mouse Assay

    SOX10-Cre/tdT -labeled distinct proportions of types I, II, and III differentiated taste bud cells. (A–C) Representative images (single-plane laser scanning confocal) of transverse (A 1 , B 1 , C 1 ) and sagittal (A 2 , B 2 , C 2 ) sections of the circumvallate papilla in 8-week-old SOX10-Cre/tdT mice. Specific types of taste bud cells were labeled by NTPDase II for type I (A 1–2 ) , PLCβ2 for type II (B 1–2 ) , and SNAP25 for type III (C 1–2 ) cells. Solid arrowheads point to the cells co-labeled by tdT and markers for specific types of taste bud cells. Open arrowheads point to tdT + cells in taste buds that were negative for specific taste cell type markers. Scale bars: 50 μm in (A 1 , B 1 , C 1 ) and 20 μm for (A 2 , B 2 , C 2 ) . (D, E) Histograms ( X ± SD, n = 3) to illustrate the percentages of tdT + specific type (I, II, and III) versus total tdT + (D) or versus total type II or III (E) taste bud cells in circumvallate taste buds. The diamonds represent data points of individual samples. ** P
    Figure Legend Snippet: SOX10-Cre/tdT -labeled distinct proportions of types I, II, and III differentiated taste bud cells. (A–C) Representative images (single-plane laser scanning confocal) of transverse (A 1 , B 1 , C 1 ) and sagittal (A 2 , B 2 , C 2 ) sections of the circumvallate papilla in 8-week-old SOX10-Cre/tdT mice. Specific types of taste bud cells were labeled by NTPDase II for type I (A 1–2 ) , PLCβ2 for type II (B 1–2 ) , and SNAP25 for type III (C 1–2 ) cells. Solid arrowheads point to the cells co-labeled by tdT and markers for specific types of taste bud cells. Open arrowheads point to tdT + cells in taste buds that were negative for specific taste cell type markers. Scale bars: 50 μm in (A 1 , B 1 , C 1 ) and 20 μm for (A 2 , B 2 , C 2 ) . (D, E) Histograms ( X ± SD, n = 3) to illustrate the percentages of tdT + specific type (I, II, and III) versus total tdT + (D) or versus total type II or III (E) taste bud cells in circumvallate taste buds. The diamonds represent data points of individual samples. ** P

    Techniques Used: Labeling, Mouse Assay

    SOX10-Cre/tdT -labeled taste bud cells had a low intensity of Krt8 immunosignals ( green ). (A) Images of sagittal sections of the circumvallate papilla in an 8-week-old SOX10-Cre/tdT mouse. E-Cadherin (E-cad, cyan ), Krt8 ( green ), and counterstained with DAPI ( blue ). Open arrowheads point to tdT + taste bud cells that were not obvious in Krt8 immunolabeling. Scale bars: 20 μm (single-plane laser scanning confocal images). (B, C) Histograms (B) and box plots (C) to show the CTCF distribution of intensity of Krt8 immunosignals in tdT + and tdT − taste bud cells in SOX10-Cre/tdT mouse circumvallate papilla ( n = 3). The boxes in (C) represent the interquartile range, the whiskers represent the range of normal distribution, and dots represent the outliners. The line and diamond within each box represent the median and average value, respectively. ** P
    Figure Legend Snippet: SOX10-Cre/tdT -labeled taste bud cells had a low intensity of Krt8 immunosignals ( green ). (A) Images of sagittal sections of the circumvallate papilla in an 8-week-old SOX10-Cre/tdT mouse. E-Cadherin (E-cad, cyan ), Krt8 ( green ), and counterstained with DAPI ( blue ). Open arrowheads point to tdT + taste bud cells that were not obvious in Krt8 immunolabeling. Scale bars: 20 μm (single-plane laser scanning confocal images). (B, C) Histograms (B) and box plots (C) to show the CTCF distribution of intensity of Krt8 immunosignals in tdT + and tdT − taste bud cells in SOX10-Cre/tdT mouse circumvallate papilla ( n = 3). The boxes in (C) represent the interquartile range, the whiskers represent the range of normal distribution, and dots represent the outliners. The line and diamond within each box represent the median and average value, respectively. ** P

    Techniques Used: Labeling, Immunolabeling

    14) Product Images from "Oroxindin inhibits macrophage NLRP3 inflammasome activation in DSS-induced ulcerative colitis in mice via suppressing TXNIP-dependent NF-κB pathway"

    Article Title: Oroxindin inhibits macrophage NLRP3 inflammasome activation in DSS-induced ulcerative colitis in mice via suppressing TXNIP-dependent NF-κB pathway

    Journal: Acta Pharmacologica Sinica

    doi: 10.1038/s41401-019-0335-4

    Oroxindin ameliorated DSS-induced acute colitis via inhibition of the NLRP3 inflammasome. a Sections of colonic tissue were immunostained with F4/80 (red) and NLRP3 (green) antibodies and observed under a confocal laser scanning microscope and shown as representative data. b Sections of colonic tissue were immunostained with F4/80 (red) and caspase-1 (green) antibodies and observed under a confocal laser scanning microscope and are shown as representative data. Enlarged areas of interest are shown in merged images. c The concentration of IL-1β in colonic homogenates was determined by an enzyme-linked immunosorbent assay (ELISA) in triplicate. d The concentration of IL-18 in colonic homogenates was determined by ELISA in triplicate. Data are presented as the mean ± SD. ## P
    Figure Legend Snippet: Oroxindin ameliorated DSS-induced acute colitis via inhibition of the NLRP3 inflammasome. a Sections of colonic tissue were immunostained with F4/80 (red) and NLRP3 (green) antibodies and observed under a confocal laser scanning microscope and shown as representative data. b Sections of colonic tissue were immunostained with F4/80 (red) and caspase-1 (green) antibodies and observed under a confocal laser scanning microscope and are shown as representative data. Enlarged areas of interest are shown in merged images. c The concentration of IL-1β in colonic homogenates was determined by an enzyme-linked immunosorbent assay (ELISA) in triplicate. d The concentration of IL-18 in colonic homogenates was determined by ELISA in triplicate. Data are presented as the mean ± SD. ## P

    Techniques Used: Inhibition, Laser-Scanning Microscopy, Concentration Assay, Enzyme-linked Immunosorbent Assay

    15) Product Images from "Changes in Optical Properties upon Dye–Clay Interaction: Experimental Evaluation and Applications"

    Article Title: Changes in Optical Properties upon Dye–Clay Interaction: Experimental Evaluation and Applications

    Journal: Nanomaterials

    doi: 10.3390/nano11010197

    Confocal laser scanning microscopy (CLSM) analysis of FITC and Chi_FITC treated with 1C MMT.
    Figure Legend Snippet: Confocal laser scanning microscopy (CLSM) analysis of FITC and Chi_FITC treated with 1C MMT.

    Techniques Used: Confocal Laser Scanning Microscopy

    16) Product Images from "Oxicam-type nonsteroidal anti-inflammatory drugs inhibit NPR1-mediated salicylic acid pathway"

    Article Title: Oxicam-type nonsteroidal anti-inflammatory drugs inhibit NPR1-mediated salicylic acid pathway

    Journal: bioRxiv

    doi: 10.1101/2020.09.25.311100

    Effect of IBF and IDM on SA-induced NPR1 accumulation. a NPR1p : NPR1-YFP seedlings were treated with 100 μM NSAIDs (IBF or IDM) or 0.5% DMSO for 1 hour before 100 μM SA (+) or water (-) treatment for 24 hours. Fluorescent image was obtained with a confocal laser microscope. YFP fluorescence and chlorophyll autofluorescence were shown in yellow and magenta, respectively. Bar = 20 μm. b NPR1p : NPR1-YFP seedlings were treated with SA and/or NSAIDs as in ( a ). Protein extracts of the seedlings were resolved by SDS-PAGE and analyzed by immunoblotting using anti-NPR1 and anti-PR1 antibodies. CBB staining was shown as a loading control.
    Figure Legend Snippet: Effect of IBF and IDM on SA-induced NPR1 accumulation. a NPR1p : NPR1-YFP seedlings were treated with 100 μM NSAIDs (IBF or IDM) or 0.5% DMSO for 1 hour before 100 μM SA (+) or water (-) treatment for 24 hours. Fluorescent image was obtained with a confocal laser microscope. YFP fluorescence and chlorophyll autofluorescence were shown in yellow and magenta, respectively. Bar = 20 μm. b NPR1p : NPR1-YFP seedlings were treated with SA and/or NSAIDs as in ( a ). Protein extracts of the seedlings were resolved by SDS-PAGE and analyzed by immunoblotting using anti-NPR1 and anti-PR1 antibodies. CBB staining was shown as a loading control.

    Techniques Used: Microscopy, Fluorescence, SDS Page, Staining

    17) Product Images from "Global transcriptional regulation by cell-free supernatant of Salmonella Typhimurium peptide transporter mutant leads to inhibition of intra-species biofilm initiation"

    Article Title: Global transcriptional regulation by cell-free supernatant of Salmonella Typhimurium peptide transporter mutant leads to inhibition of intra-species biofilm initiation

    Journal: bioRxiv

    doi: 10.1101/2020.07.15.204859

    Salmonella ΔyjiY cell free supernatant significantly reduces the biofilm biomass by reducing cell-cell adhesion A. Representative images of biofilm formed on coverslips that were stained with Congo red and imaged using a confocal microscope to generate 3D images and quantify cellulose biomass. Scale is shown on the X- and Y-axes. B. The tensile strength of the biofilm was measured by glass bead assay. Weight of glass beads required to just sink the biofilm to bottom, was plotted (Data are presented as mean + SEM of 5 independent experiments). C. Representative scanning electron micrograph of biofilm formed on a coverslip. Scale bar is 10 μm. D. Cell length of the biofilm inoculated treated or untreated STM WT cells was measured using ImageJ, and plotted (Data are presented as mean + SEM of 1200 cells were measured from 3 independent experiments). E. Representative confocal images of biofilm cells showing a difference in cell length. Scale bar is 5 μm (1000-1200 cells were measured from 3 independent experiments for each treatment). Student’s t-test was used to analyze the data; p values ****
    Figure Legend Snippet: Salmonella ΔyjiY cell free supernatant significantly reduces the biofilm biomass by reducing cell-cell adhesion A. Representative images of biofilm formed on coverslips that were stained with Congo red and imaged using a confocal microscope to generate 3D images and quantify cellulose biomass. Scale is shown on the X- and Y-axes. B. The tensile strength of the biofilm was measured by glass bead assay. Weight of glass beads required to just sink the biofilm to bottom, was plotted (Data are presented as mean + SEM of 5 independent experiments). C. Representative scanning electron micrograph of biofilm formed on a coverslip. Scale bar is 10 μm. D. Cell length of the biofilm inoculated treated or untreated STM WT cells was measured using ImageJ, and plotted (Data are presented as mean + SEM of 1200 cells were measured from 3 independent experiments). E. Representative confocal images of biofilm cells showing a difference in cell length. Scale bar is 5 μm (1000-1200 cells were measured from 3 independent experiments for each treatment). Student’s t-test was used to analyze the data; p values ****

    Techniques Used: Staining, Microscopy

    18) Product Images from "Micro Versus Macro – The Effect of Environmental Confinement on Cellular Nanoparticle Uptake"

    Article Title: Micro Versus Macro – The Effect of Environmental Confinement on Cellular Nanoparticle Uptake

    Journal: Frontiers in Bioengineering and Biotechnology

    doi: 10.3389/fbioe.2020.00869

    Schematic of the experimental methodology. (A) Seeding equal number of cells in the microfluidic device and the petri dish and allowing cells 4–16 h to attach to the glass bottom of the device. (B) FND suspension incubation for 4 h. (C) Fixed and stained macrophages are imaged in a laser scanning confocal microscope. Red dots in the image are FNDs where nucleus and cytoskeleton are indicated with blue and green color respectively. In every experiment, FNDs/cell are quantified in 50 cells per group. Each experiment is repeated three independent times.
    Figure Legend Snippet: Schematic of the experimental methodology. (A) Seeding equal number of cells in the microfluidic device and the petri dish and allowing cells 4–16 h to attach to the glass bottom of the device. (B) FND suspension incubation for 4 h. (C) Fixed and stained macrophages are imaged in a laser scanning confocal microscope. Red dots in the image are FNDs where nucleus and cytoskeleton are indicated with blue and green color respectively. In every experiment, FNDs/cell are quantified in 50 cells per group. Each experiment is repeated three independent times.

    Techniques Used: Incubation, Staining, Microscopy

    19) Product Images from "Capturing Amyloid-β Oligomers by Stirring with Microscaled Iron Oxide Stir Bars into Magnetic Plaques to Reduce Cytotoxicity toward Neuronal Cells"

    Article Title: Capturing Amyloid-β Oligomers by Stirring with Microscaled Iron Oxide Stir Bars into Magnetic Plaques to Reduce Cytotoxicity toward Neuronal Cells

    Journal: Nanomaterials

    doi: 10.3390/nano10071284

    Phagocytic action of BV-2 cells. ( a ) Laser scanning confocal microscope (LSCM) images of BV-2 cells after co-incubation with oAβ 42 , npAβ 42 , and mpAβ 42 , respectively ( n = 8). Aβ 42 in different forms was IHC stained using 6E10 as the primary antibody (λ ex = 565 nm, λ em = 680–730 nm). Scale bar: 50 μm. ( b ) Relative uptake index of Aβ 42 by BV-2 cells. One-way ANOVA was used to examine the mean differences between the end points of the data groups. ** p
    Figure Legend Snippet: Phagocytic action of BV-2 cells. ( a ) Laser scanning confocal microscope (LSCM) images of BV-2 cells after co-incubation with oAβ 42 , npAβ 42 , and mpAβ 42 , respectively ( n = 8). Aβ 42 in different forms was IHC stained using 6E10 as the primary antibody (λ ex = 565 nm, λ em = 680–730 nm). Scale bar: 50 μm. ( b ) Relative uptake index of Aβ 42 by BV-2 cells. One-way ANOVA was used to examine the mean differences between the end points of the data groups. ** p

    Techniques Used: Microscopy, Incubation, Immunohistochemistry, Staining

    20) Product Images from "Effect of Astaxanthin on Activation of Autophagy and Inhibition of Apoptosis in Helicobacter pylori-Infected Gastric Epithelial Cell Line AGS"

    Article Title: Effect of Astaxanthin on Activation of Autophagy and Inhibition of Apoptosis in Helicobacter pylori-Infected Gastric Epithelial Cell Line AGS

    Journal: Nutrients

    doi: 10.3390/nu12061750

    Effect of astaxanthin on autophagy activation in H. pylori -stimulated AGS cells. The cells were pre-treated with 50 nM astaxanthin for 3 h and then stimulated with H. pylori (cell to H. pylori ratio of 1:50) for 24 h. ( A ) Cells were stained with acridine orange (AO) dye and visualized under a confocal laser scanning microscope (left panel). AO-positive cells were quantified and expressed as % of cells with AO-positive cells/total number of cells (right panel). ( B ) The cells were stained with anti-LC3B antibody and rhodamine-labeled mouse anti-rabbit IgG antibody. Immunocytochemical staining for LC3B (red) and DNA counterstaining with DAPI (blue) are shown in the left panel. Each sample was analyzed using a threshold of > 7 dots/cell. LC3B puncta-positive cells were quantified and expressed as % of cells with > 7 LC3B puncta/total number of cells (right panel). * p
    Figure Legend Snippet: Effect of astaxanthin on autophagy activation in H. pylori -stimulated AGS cells. The cells were pre-treated with 50 nM astaxanthin for 3 h and then stimulated with H. pylori (cell to H. pylori ratio of 1:50) for 24 h. ( A ) Cells were stained with acridine orange (AO) dye and visualized under a confocal laser scanning microscope (left panel). AO-positive cells were quantified and expressed as % of cells with AO-positive cells/total number of cells (right panel). ( B ) The cells were stained with anti-LC3B antibody and rhodamine-labeled mouse anti-rabbit IgG antibody. Immunocytochemical staining for LC3B (red) and DNA counterstaining with DAPI (blue) are shown in the left panel. Each sample was analyzed using a threshold of > 7 dots/cell. LC3B puncta-positive cells were quantified and expressed as % of cells with > 7 LC3B puncta/total number of cells (right panel). * p

    Techniques Used: Activation Assay, Staining, Laser-Scanning Microscopy, Labeling

    21) Product Images from "MKP-5 Relieves Lipotoxicity-Induced Islet β-Cell Dysfunction and Apoptosis via Regulation of Autophagy"

    Article Title: MKP-5 Relieves Lipotoxicity-Induced Islet β-Cell Dysfunction and Apoptosis via Regulation of Autophagy

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms21197161

    MKP-5 regulates the PA-induced impairment of autophagic flux in Rin-m5f cells. ( A ) Rin-PC and Rin-MKP-5 cells were transfected with the ptfLC3 plasmid for 48 h, after which they were treated with PA and 3-methyladenine (3-MA) for 9 h, and analyzed by laser scanning confocal fluorescence microscopy (scale bar = 5 µm). ( B ) Quantification of cells positive for red fluorescent protein (mRFP) (red) and mRFP/GFP (green fluorescent protein, yellow) in Rin-PC and Rin-MKP-5 cells transfected with ptfLC3 prior to a 9 h treatment with PA or 3-MA alone or in combination with one another. ( C ) The levels of autophagic proteins in Rin-m5f cells treated with PA ± 3-MA for 9 h were measured by Western blotting (upper) and quantitative analysis was conducted using Image J (lower). *, p
    Figure Legend Snippet: MKP-5 regulates the PA-induced impairment of autophagic flux in Rin-m5f cells. ( A ) Rin-PC and Rin-MKP-5 cells were transfected with the ptfLC3 plasmid for 48 h, after which they were treated with PA and 3-methyladenine (3-MA) for 9 h, and analyzed by laser scanning confocal fluorescence microscopy (scale bar = 5 µm). ( B ) Quantification of cells positive for red fluorescent protein (mRFP) (red) and mRFP/GFP (green fluorescent protein, yellow) in Rin-PC and Rin-MKP-5 cells transfected with ptfLC3 prior to a 9 h treatment with PA or 3-MA alone or in combination with one another. ( C ) The levels of autophagic proteins in Rin-m5f cells treated with PA ± 3-MA for 9 h were measured by Western blotting (upper) and quantitative analysis was conducted using Image J (lower). *, p

    Techniques Used: Transfection, Plasmid Preparation, Fluorescence, Microscopy, Western Blot

    22) Product Images from "Genome-wide Identification and Characterization of FCS-Like Zinc Finger (FLZ) Family Genes in Maize (Zea mays) and Functional Analysis of ZmFLZ25 in Plant Abscisic Acid Response"

    Article Title: Genome-wide Identification and Characterization of FCS-Like Zinc Finger (FLZ) Family Genes in Maize (Zea mays) and Functional Analysis of ZmFLZ25 in Plant Abscisic Acid Response

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms22073529

    Subcellular localization of the eight selected ZmFLZs and ZmFLZs interaction with ZmKIN10 by Y2H assay. ( A ) ZmFLZs-GFP fusion constructs were used to determine the subcellular localization of ZmFLZs in the protoplast cells isolated from maize leaves. NLS-mCherry was used as a nuclear marker. Fluorescent images of GFP and mCherry were captured with a confocal laser scanning microscopy and shown in green and red, respectively. Scale bars = 25 μm. ( B , C ) Yeast two hybrid analysis of the binary interactions between the eight selected typical ZmFLZs and ZmKIN10.
    Figure Legend Snippet: Subcellular localization of the eight selected ZmFLZs and ZmFLZs interaction with ZmKIN10 by Y2H assay. ( A ) ZmFLZs-GFP fusion constructs were used to determine the subcellular localization of ZmFLZs in the protoplast cells isolated from maize leaves. NLS-mCherry was used as a nuclear marker. Fluorescent images of GFP and mCherry were captured with a confocal laser scanning microscopy and shown in green and red, respectively. Scale bars = 25 μm. ( B , C ) Yeast two hybrid analysis of the binary interactions between the eight selected typical ZmFLZs and ZmKIN10.

    Techniques Used: Y2H Assay, Construct, Isolation, Marker, Confocal Laser Scanning Microscopy

    Co-localization analysis of ZmFLZs and ZmKIN10 in maize leaf protoplast cells. Co-expression of mCherry-ZmKIN10 and the GFP fused ZmFLZs in protoplasts followed by confocal imaging. Fluorescent images of GFP and mCherry were captured with a confocal laser scanning microscopy and shown in green and red, respectively. Scale bars = 25 μm.
    Figure Legend Snippet: Co-localization analysis of ZmFLZs and ZmKIN10 in maize leaf protoplast cells. Co-expression of mCherry-ZmKIN10 and the GFP fused ZmFLZs in protoplasts followed by confocal imaging. Fluorescent images of GFP and mCherry were captured with a confocal laser scanning microscopy and shown in green and red, respectively. Scale bars = 25 μm.

    Techniques Used: Expressing, Imaging, Confocal Laser Scanning Microscopy

    23) Product Images from "RepA Promotes the Nucleolar Exclusion of the V2 Protein of Mulberry Mosaic Dwarf-Associated Virus"

    Article Title: RepA Promotes the Nucleolar Exclusion of the V2 Protein of Mulberry Mosaic Dwarf-Associated Virus

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2020.01828

    RepA-dependent nucleolar exclusion of V2. RFP-H2B plant leaves were infiltrated with Agrobacterium tumefaciens cultures carrying constructs to express GFP-V2 and the other six individual ORFs (V1, V3, V4, V5, RepA, and Rep) of MMDaV. Fluorescence was visualized under a Zeiss LSM 880 confocal laser scanning microscope at 36 hpi. The corresponding region in the white box in column 1 is magnified and shown from Column 2 to Column 5. RFP-H2B was used as a nuclear marker. This experiment was done three times and more than 20 cells were observed per sample and replicate. Scale bars correspond to 10 μm.
    Figure Legend Snippet: RepA-dependent nucleolar exclusion of V2. RFP-H2B plant leaves were infiltrated with Agrobacterium tumefaciens cultures carrying constructs to express GFP-V2 and the other six individual ORFs (V1, V3, V4, V5, RepA, and Rep) of MMDaV. Fluorescence was visualized under a Zeiss LSM 880 confocal laser scanning microscope at 36 hpi. The corresponding region in the white box in column 1 is magnified and shown from Column 2 to Column 5. RFP-H2B was used as a nuclear marker. This experiment was done three times and more than 20 cells were observed per sample and replicate. Scale bars correspond to 10 μm.

    Techniques Used: Construct, Fluorescence, Laser-Scanning Microscopy, Marker

    V2 interacts with NbFib2. (A) Yeast two-hybrid assay showing the interaction between V2 and NbFib2 in yeast cells. Full-length NbFib2 was expressed as GAL4 DNA-binding domain fusion (BD, bait) and V2 was expressed as GAL4 activation domain fusion (AD, prey) in yeast cells of the strain Y2H Gold. The interaction of p53 and T was used as a positive control, and cotransformation of Lam and T was used as a negative control. Growth on the plates lacking leucine and tryptophan (SD/-LT) indicates successful transformation of both prey and bait vectors, respectively. Interaction between NbFib2 and V2 is indicated by growth of yeast cells on media also lacking histidine supplementing with 5 mM 3-amino-1,2,4-triazole (SD/-LTH + 3-AT). (B) Bimolecular fluorescence complementation assay (BiFC) assay showing the interaction between NbFib2 and V2 in plant cells. Constructs containing N-terminal YFP fusion (nYFP) and C-terminal YFP fusion (cYFP) fusions were infiltrated into RFP-H2B plant leaves. Combinations of BiFC constructs are shown at the top of each panel. Images were taken using a Zeiss LSM 880 confocal laser scanning microscope at 48 hpi. Reconstituted YFP signals resulting from V2-NbFib2 interaction are displayed as a false-green color. RFP-H2B served as a nuclear marker. Note that deletion of the predicted nuclear localization signal (from amino acid 61–76) of V2 abolishes its interaction with NbFib2. (C) Colocalization analysis of NbFib2 with V2 and V2 mutant in the epidermal cells of N. benthamiana by Zeiss LSM 880 confocal laser scanning microscope at 36 hpi. At least 60 cells from three repeats were examined. Scale bars correspond to 10 μm. (D) Immunoblot of proteins from RFP-H2B plant leaves infiltrated with construct as indicated using anti-GFP antibody. Ponceau staining of the large subunit of Rubisco serves as a loading control.
    Figure Legend Snippet: V2 interacts with NbFib2. (A) Yeast two-hybrid assay showing the interaction between V2 and NbFib2 in yeast cells. Full-length NbFib2 was expressed as GAL4 DNA-binding domain fusion (BD, bait) and V2 was expressed as GAL4 activation domain fusion (AD, prey) in yeast cells of the strain Y2H Gold. The interaction of p53 and T was used as a positive control, and cotransformation of Lam and T was used as a negative control. Growth on the plates lacking leucine and tryptophan (SD/-LT) indicates successful transformation of both prey and bait vectors, respectively. Interaction between NbFib2 and V2 is indicated by growth of yeast cells on media also lacking histidine supplementing with 5 mM 3-amino-1,2,4-triazole (SD/-LTH + 3-AT). (B) Bimolecular fluorescence complementation assay (BiFC) assay showing the interaction between NbFib2 and V2 in plant cells. Constructs containing N-terminal YFP fusion (nYFP) and C-terminal YFP fusion (cYFP) fusions were infiltrated into RFP-H2B plant leaves. Combinations of BiFC constructs are shown at the top of each panel. Images were taken using a Zeiss LSM 880 confocal laser scanning microscope at 48 hpi. Reconstituted YFP signals resulting from V2-NbFib2 interaction are displayed as a false-green color. RFP-H2B served as a nuclear marker. Note that deletion of the predicted nuclear localization signal (from amino acid 61–76) of V2 abolishes its interaction with NbFib2. (C) Colocalization analysis of NbFib2 with V2 and V2 mutant in the epidermal cells of N. benthamiana by Zeiss LSM 880 confocal laser scanning microscope at 36 hpi. At least 60 cells from three repeats were examined. Scale bars correspond to 10 μm. (D) Immunoblot of proteins from RFP-H2B plant leaves infiltrated with construct as indicated using anti-GFP antibody. Ponceau staining of the large subunit of Rubisco serves as a loading control.

    Techniques Used: Y2H Assay, Binding Assay, Activation Assay, Positive Control, Laser Capture Microdissection, Negative Control, Transformation Assay, Bimolecular Fluorescence Complementation Assay, Construct, Laser-Scanning Microscopy, Marker, Mutagenesis, Staining

    RepA interacts with V2 in yeast and plant cells. (A) Yeast two-hybrid assay showing the interaction between RepA and V2 in yeast cells. Growth of yeast cotransformants containing the BD-RepA and AD-V2 fusions on the plates lacking leucine, tryptophan, histidine, and adenine (SD/-LTHA) indicates specific interaction between RepA and V2. (B) BiFC assay showing the interaction between RepA and V2 in plant cells. Combinations of BiFC constructs are shown at the top of each panel. Images were taken using a Zeiss LSM 880 confocal laser scanning microscope at 48 hpi. Reconstituted YFP signals as a consequence of V2-RepA interaction are depicted as a false-green color. RFP-H2B served as a nuclear marker.
    Figure Legend Snippet: RepA interacts with V2 in yeast and plant cells. (A) Yeast two-hybrid assay showing the interaction between RepA and V2 in yeast cells. Growth of yeast cotransformants containing the BD-RepA and AD-V2 fusions on the plates lacking leucine, tryptophan, histidine, and adenine (SD/-LTHA) indicates specific interaction between RepA and V2. (B) BiFC assay showing the interaction between RepA and V2 in plant cells. Combinations of BiFC constructs are shown at the top of each panel. Images were taken using a Zeiss LSM 880 confocal laser scanning microscope at 48 hpi. Reconstituted YFP signals as a consequence of V2-RepA interaction are depicted as a false-green color. RFP-H2B served as a nuclear marker.

    Techniques Used: Y2H Assay, Bimolecular Fluorescence Complementation Assay, Construct, Laser-Scanning Microscopy, Marker

    V2 localization within the nucleolus is modulated in the presence of mulberry mosaic dwarf-associated geminivirus (MMDaV). (A) Subcellular localization of green fluorescent protein (GFP) or GFP-V2 fusion in the absence or presence of MMDaV infection in transgenic Nicotiana benthamian a plants expressing red fluorescent protein (RFP)-tagged histone 2B (RFP-H2B). RFP-H2B was used as a nuclear marker. (B) Colocalization analysis of V2 and fibrillarin 2 (NbFib2) in the absence or presence of MMDaV in N. benthamiana plants. To create an environment mimicking MMDaV infection, the infectious clone of MMDaV was infiltrated into RFP-H2B or N. benthamiana leaves 12 h prior to the infiltration of GFP or GFP-V2. Images were taken using Zeiss LSM 880 confocal laser scanning microscope at 36 hours post infiltration (hpi) of GFP or GFP-V2. This experiment was done three times and more than 20 cells were observed per sample and replicate. A representative image is shown for each set. The corresponding region in the white box in column 1 is magnified and shown from Column 2 to Column 5. Scale bars correspond to 10 μm.
    Figure Legend Snippet: V2 localization within the nucleolus is modulated in the presence of mulberry mosaic dwarf-associated geminivirus (MMDaV). (A) Subcellular localization of green fluorescent protein (GFP) or GFP-V2 fusion in the absence or presence of MMDaV infection in transgenic Nicotiana benthamian a plants expressing red fluorescent protein (RFP)-tagged histone 2B (RFP-H2B). RFP-H2B was used as a nuclear marker. (B) Colocalization analysis of V2 and fibrillarin 2 (NbFib2) in the absence or presence of MMDaV in N. benthamiana plants. To create an environment mimicking MMDaV infection, the infectious clone of MMDaV was infiltrated into RFP-H2B or N. benthamiana leaves 12 h prior to the infiltration of GFP or GFP-V2. Images were taken using Zeiss LSM 880 confocal laser scanning microscope at 36 hours post infiltration (hpi) of GFP or GFP-V2. This experiment was done three times and more than 20 cells were observed per sample and replicate. A representative image is shown for each set. The corresponding region in the white box in column 1 is magnified and shown from Column 2 to Column 5. Scale bars correspond to 10 μm.

    Techniques Used: Infection, Transgenic Assay, Expressing, Marker, Laser-Scanning Microscopy

    24) Product Images from "The Role of Lipase and α-Amylase in the Degradation of Starch/Poly(ɛ-Caprolactone) Fiber Meshes and the Osteogenic Differentiation of Cultured Marrow Stromal Cells"

    Article Title: The Role of Lipase and α-Amylase in the Degradation of Starch/Poly(ɛ-Caprolactone) Fiber Meshes and the Osteogenic Differentiation of Cultured Marrow Stromal Cells

    Journal:

    doi: 10.1089/ten.tea.2008.0025

    Laser scanning confocal microscope images of rat marrow stromal cells stained with calcein-acetoxymethyl ester at the top surface of starch and poly(ɛ-caprolactone) fiber meshes with lipase ( A, D ) or α-amylase ( B, E ) or without enzymes
    Figure Legend Snippet: Laser scanning confocal microscope images of rat marrow stromal cells stained with calcein-acetoxymethyl ester at the top surface of starch and poly(ɛ-caprolactone) fiber meshes with lipase ( A, D ) or α-amylase ( B, E ) or without enzymes

    Techniques Used: Microscopy, Staining

    Laser scanning confocal microscopy images of starch and poly(ɛ-caprolactone) fiber meshes cultured for 16 days with lipase ( A ) or α-amylase ( B ) or without enzymes ( C ) obtained from depth projections. The scale bar is 100 μm
    Figure Legend Snippet: Laser scanning confocal microscopy images of starch and poly(ɛ-caprolactone) fiber meshes cultured for 16 days with lipase ( A ) or α-amylase ( B ) or without enzymes ( C ) obtained from depth projections. The scale bar is 100 μm

    Techniques Used: Confocal Microscopy, Cell Culture

    25) Product Images from "Patchless administration of canine influenza vaccine on dog’s ear using insertion-responsive microneedles (IRMN) without removal of hair and its in vivo efficacy evaluation"

    Article Title: Patchless administration of canine influenza vaccine on dog’s ear using insertion-responsive microneedles (IRMN) without removal of hair and its in vivo efficacy evaluation

    Journal: European Journal of Pharmaceutics and Biopharmaceutics

    doi: 10.1016/j.ejpb.2020.06.006

    Confocal laser scanning microscopy images of DiD-loaded vc378 distribution in the microneedle. Representative confocal images of microneedles coated with DiD-loaded vc378 (left, DiD-loaded vc378; middle, DIC; right, merge). The scale bars are 200 μm.
    Figure Legend Snippet: Confocal laser scanning microscopy images of DiD-loaded vc378 distribution in the microneedle. Representative confocal images of microneedles coated with DiD-loaded vc378 (left, DiD-loaded vc378; middle, DIC; right, merge). The scale bars are 200 μm.

    Techniques Used: Confocal Laser Scanning Microscopy

    26) Product Images from "Whole Transcriptome Analysis Provides Insights Into the Molecular Mechanisms of Chlamydospore-Like Cell Formation in Phanerochaete chrysosporium"

    Article Title: Whole Transcriptome Analysis Provides Insights Into the Molecular Mechanisms of Chlamydospore-Like Cell Formation in Phanerochaete chrysosporium

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2020.527389

    Different morphological characteristics of hyphae and chlamydospores. (A) Light microscopy of the hyphae at 72 h. (B) Light microscopy of the chlamydospores at 72 h. (C) Laser scanning confocal microscope (LSCM) of the hyphae at 72 h. (D) LSCM of the chlamydospores at 72 h. (E) measurement of the thickness of the cell wall between the hyphae and the chlamydospores. Asterisk indicates that the thickness of the cell wall was significantly different between hyphae and chlamydospores (Student’s t -test, ** p
    Figure Legend Snippet: Different morphological characteristics of hyphae and chlamydospores. (A) Light microscopy of the hyphae at 72 h. (B) Light microscopy of the chlamydospores at 72 h. (C) Laser scanning confocal microscope (LSCM) of the hyphae at 72 h. (D) LSCM of the chlamydospores at 72 h. (E) measurement of the thickness of the cell wall between the hyphae and the chlamydospores. Asterisk indicates that the thickness of the cell wall was significantly different between hyphae and chlamydospores (Student’s t -test, ** p

    Techniques Used: Light Microscopy, Microscopy

    27) Product Images from "Bionic Silk Fibroin Film Promotes Tenogenic Differentiation of Tendon Stem/Progenitor Cells by Activating Focal Adhesion Kinase"

    Article Title: Bionic Silk Fibroin Film Promotes Tenogenic Differentiation of Tendon Stem/Progenitor Cells by Activating Focal Adhesion Kinase

    Journal: Stem Cells International

    doi: 10.1155/2020/8857380

    The cell morphology of TSPCs on different matrix surfaces. (a) Cell morphology observation: (A–C) cell morphology under a light microscope; (D–F) the morphology of TSPCs under a confocal laser scanning microscope. The nuclei were stained blue; the cytoskeletons were stained red; (A, D) TSPCs in the cell culture plate; (B, E) TSPCs on the smooth SF film; (C, F) TSPCs on the SF film with a microstructure. (b) Analysis of cell morphology: (A–D) cell body aspects; (E–H) cell body major axis angle (I–L) cell area; group C: TSPCs on the cell culture plate; group S: TSPCs on the smooth SF film; group G: TSPCs on the SF film with a microstructure; ∗∗ P
    Figure Legend Snippet: The cell morphology of TSPCs on different matrix surfaces. (a) Cell morphology observation: (A–C) cell morphology under a light microscope; (D–F) the morphology of TSPCs under a confocal laser scanning microscope. The nuclei were stained blue; the cytoskeletons were stained red; (A, D) TSPCs in the cell culture plate; (B, E) TSPCs on the smooth SF film; (C, F) TSPCs on the SF film with a microstructure. (b) Analysis of cell morphology: (A–D) cell body aspects; (E–H) cell body major axis angle (I–L) cell area; group C: TSPCs on the cell culture plate; group S: TSPCs on the smooth SF film; group G: TSPCs on the SF film with a microstructure; ∗∗ P

    Techniques Used: Light Microscopy, Laser-Scanning Microscopy, Staining, Cell Culture

    28) Product Images from "DDRGK1, a crucial player of ufmylation system, is indispensable for autophagic degradation by regulating lysosomal function"

    Article Title: DDRGK1, a crucial player of ufmylation system, is indispensable for autophagic degradation by regulating lysosomal function

    Journal: Cell Death & Disease

    doi: 10.1038/s41419-021-03694-9

    DDRGK1 loss impairs autophagic flux. a Representative images of control and DDRGK1-deleted MEFs transfected with mCherry-EGFP-LC3 plasmid for 36 h. Enlarged images of the yellow box were indicated on the right side. Laser confocal-scanning microscopy was employed to observe the fluorescent puncta. Bar, 10 μm. b Quantification of only mCherry-labeled and EGFP + mCherry-labeled puncta. Data represent mean ± SEM; n = 3. * P
    Figure Legend Snippet: DDRGK1 loss impairs autophagic flux. a Representative images of control and DDRGK1-deleted MEFs transfected with mCherry-EGFP-LC3 plasmid for 36 h. Enlarged images of the yellow box were indicated on the right side. Laser confocal-scanning microscopy was employed to observe the fluorescent puncta. Bar, 10 μm. b Quantification of only mCherry-labeled and EGFP + mCherry-labeled puncta. Data represent mean ± SEM; n = 3. * P

    Techniques Used: Transfection, Plasmid Preparation, Confocal Laser Scanning Microscopy, Labeling

    29) Product Images from "Lotus seedpod-inspired internal vascularized 3D printed scaffold for bone tissue repair"

    Article Title: Lotus seedpod-inspired internal vascularized 3D printed scaffold for bone tissue repair

    Journal: Bioactive Materials

    doi: 10.1016/j.bioactmat.2020.11.019

    In vitro biocompatibility of 3T3-E1 cells with the scaffolds. (a) Live/Dead analysis of 3T3 -E1 cells cultured on the scaffolds on day 1, 4, and 7. Live cells were stained green, while dead cells appeared red. (b) The laser scanning confocal microscopic images of MC3T3-E1 cells adhered to the scaffolds for 1, 4 and 7 days. The cytoskeleton was stained in red and the nucleus was blue. (c) The survival and proliferation behavior of cells on GML for 1, 4, and 7 days. (d) Statistics of the number of living cells. (e) CCK-8 assay was used to detect the toxicity of scaffolds. (All the above pictures were taken by confocal microscope. Two-way ANOVA was used; NS, no significant difference; # , P
    Figure Legend Snippet: In vitro biocompatibility of 3T3-E1 cells with the scaffolds. (a) Live/Dead analysis of 3T3 -E1 cells cultured on the scaffolds on day 1, 4, and 7. Live cells were stained green, while dead cells appeared red. (b) The laser scanning confocal microscopic images of MC3T3-E1 cells adhered to the scaffolds for 1, 4 and 7 days. The cytoskeleton was stained in red and the nucleus was blue. (c) The survival and proliferation behavior of cells on GML for 1, 4, and 7 days. (d) Statistics of the number of living cells. (e) CCK-8 assay was used to detect the toxicity of scaffolds. (All the above pictures were taken by confocal microscope. Two-way ANOVA was used; NS, no significant difference; # , P

    Techniques Used: In Vitro, Cell Culture, Staining, CCK-8 Assay, Microscopy

    The vascularization studies of β - TCP and TGL scaffolds in vitro . (a) Endothelial network formation of HUVECs at 3 and 6 h after cell culture. (b – e) The differences of total length (b), number of junctions (c), number of meshes (d) and total meshes area per high power field (HPF) were analyzed. (All the above pictures were taken by laser scanning confocal microscope. Two-way ANOVA was used; ***, p
    Figure Legend Snippet: The vascularization studies of β - TCP and TGL scaffolds in vitro . (a) Endothelial network formation of HUVECs at 3 and 6 h after cell culture. (b – e) The differences of total length (b), number of junctions (c), number of meshes (d) and total meshes area per high power field (HPF) were analyzed. (All the above pictures were taken by laser scanning confocal microscope. Two-way ANOVA was used; ***, p

    Techniques Used: In Vitro, Cell Culture, Microscopy

    30) Product Images from "Inhibition of Autophagy Prevents Panax Notoginseng Saponins (PNS) Protection on Cardiac Myocytes Against Endoplasmic Reticulum (ER) Stress-Induced Mitochondrial Injury, Ca2+ Homeostasis and Associated Apoptosis"

    Article Title: Inhibition of Autophagy Prevents Panax Notoginseng Saponins (PNS) Protection on Cardiac Myocytes Against Endoplasmic Reticulum (ER) Stress-Induced Mitochondrial Injury, Ca2+ Homeostasis and Associated Apoptosis

    Journal: Frontiers in Pharmacology

    doi: 10.3389/fphar.2021.620812

    PNS promotes autophagy and autophagic flux. (A) Primary cultured cardiomyocytes, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h (PNS group), before addition of 1 μM thapsigargin (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-LC3B antibody and imaged by a laser scanning confocal microscopy. Scale bar: 30 μm; in box: 10 μm. Bar graph shows the number of LC3 puncta per cell in various groups as indicated (Mean ± SEM; 80–100 cells; * p
    Figure Legend Snippet: PNS promotes autophagy and autophagic flux. (A) Primary cultured cardiomyocytes, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h (PNS group), before addition of 1 μM thapsigargin (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-LC3B antibody and imaged by a laser scanning confocal microscopy. Scale bar: 30 μm; in box: 10 μm. Bar graph shows the number of LC3 puncta per cell in various groups as indicated (Mean ± SEM; 80–100 cells; * p

    Techniques Used: Cell Culture, Confocal Microscopy

    Inhibition of autophagy abolishes PNS protection on TG-induced ER stress response and associated apoptosis. (A) SiNC or siATG7 transfected H9c2 cells, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h before addition of 1 μM TG (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-calnexin antibody and imaged by a laser scanning confocal microscopy. Scale bar: 30 μm; in box: 10 μm. (B) SiNC or siATG7 transfected H9c2 cells, treated as in A, were immunoblotted with the antibodies to BiP, CHOP, Cleaved Caspase-3 and Caspase-12 as well as β -actin. Bands were quantified relative to β -actin by densitometry (Mean ± SEM; ** p
    Figure Legend Snippet: Inhibition of autophagy abolishes PNS protection on TG-induced ER stress response and associated apoptosis. (A) SiNC or siATG7 transfected H9c2 cells, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h before addition of 1 μM TG (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-calnexin antibody and imaged by a laser scanning confocal microscopy. Scale bar: 30 μm; in box: 10 μm. (B) SiNC or siATG7 transfected H9c2 cells, treated as in A, were immunoblotted with the antibodies to BiP, CHOP, Cleaved Caspase-3 and Caspase-12 as well as β -actin. Bands were quantified relative to β -actin by densitometry (Mean ± SEM; ** p

    Techniques Used: Inhibition, Transfection, Confocal Microscopy

    PNS prevents TG-induced mitochondria injury and ROS accumulation. (A) Primary cultured cardiomyocytes, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h (PNS group), before addition of 1 μM thapsigargin (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-TOM20 antibody and imaged by a laser scanning confocal microscopy. Representative images in CN and TG group. Scale bar: 30 μm; in box: 10 μm. Bar graph shows percentage of cardiomyocytes with tubular, intermediate or fragmented mitochondria in various groups as indicated (80–100 cells). (B) Primary cultured cardiomyocytes treated as in A were stained with JC-1 and analyzed by a plate reader. Bar graph shows the ratio of aggregated JC-1 (red)/monomeric JC-1 (green) as the mitochondrial membrane potential (Δ ψ m) (Mean ± SEM; 80–100 cells; * p
    Figure Legend Snippet: PNS prevents TG-induced mitochondria injury and ROS accumulation. (A) Primary cultured cardiomyocytes, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h (PNS group), before addition of 1 μM thapsigargin (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-TOM20 antibody and imaged by a laser scanning confocal microscopy. Representative images in CN and TG group. Scale bar: 30 μm; in box: 10 μm. Bar graph shows percentage of cardiomyocytes with tubular, intermediate or fragmented mitochondria in various groups as indicated (80–100 cells). (B) Primary cultured cardiomyocytes treated as in A were stained with JC-1 and analyzed by a plate reader. Bar graph shows the ratio of aggregated JC-1 (red)/monomeric JC-1 (green) as the mitochondrial membrane potential (Δ ψ m) (Mean ± SEM; 80–100 cells; * p

    Techniques Used: Cell Culture, Confocal Microscopy, Staining

    31) Product Images from "Site-specific Bioconjugation and Convergent Click Chemistry Enhances Antibody–Chromophore Conjugate Binding Efficiency"

    Article Title: Site-specific Bioconjugation and Convergent Click Chemistry Enhances Antibody–Chromophore Conjugate Binding Efficiency

    Journal: Photochemistry and photobiology

    doi: 10.1111/php.13231

    Laser-scanning confocal microscopy of ( a ) Ovcar3 (EGFR high), ( b ) Powder (EGFR medium), and ( c ) T-47D (EGFR (low) cancer cell lines. Cells were incubated with non-specific amine-NHS conjugated (left) or site-specific Glutamine (Gln)-mTGase conjugated (middle) cet–AF647 constructs, or no antibody (right). All cells were stained with Hoechst 33342 nuclear stain. AF647 fluorescence is normalized to the ALR of each conjugate and represents the amount of antibody uptake per cell. AF647 fluorescence intensity for each cell line is indicated via 16-bit color bar. Scale bar, 50 μm.
    Figure Legend Snippet: Laser-scanning confocal microscopy of ( a ) Ovcar3 (EGFR high), ( b ) Powder (EGFR medium), and ( c ) T-47D (EGFR (low) cancer cell lines. Cells were incubated with non-specific amine-NHS conjugated (left) or site-specific Glutamine (Gln)-mTGase conjugated (middle) cet–AF647 constructs, or no antibody (right). All cells were stained with Hoechst 33342 nuclear stain. AF647 fluorescence is normalized to the ALR of each conjugate and represents the amount of antibody uptake per cell. AF647 fluorescence intensity for each cell line is indicated via 16-bit color bar. Scale bar, 50 μm.

    Techniques Used: Confocal Microscopy, Incubation, Construct, Staining, Fluorescence

    32) Product Images from "STAT3 Is an Upstream Regulator of Granzyme G in the Maternal-To-Zygotic Transition of Mouse Embryos"

    Article Title: STAT3 Is an Upstream Regulator of Granzyme G in the Maternal-To-Zygotic Transition of Mouse Embryos

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms22010460

    Abnormal chromosome plate in arrested zygote stage embryos treated with STAT3 inhibitor S3I-201. ( A ) Representative images of abnormal metaphase chromosome plates in arrested zygotes after treatment with 100-μM S3I-201 inhibitor in KSOM-AA embryo culture medium. Chromosomal DNA was stained with DAPI, a blue fluorescent dye. All samples were analyzed under a laser scanning confocal microscope. Scale bar = 20 µm. ( B ) Representative images of the different morphologies of the abnormal metaphase chromosome plates.
    Figure Legend Snippet: Abnormal chromosome plate in arrested zygote stage embryos treated with STAT3 inhibitor S3I-201. ( A ) Representative images of abnormal metaphase chromosome plates in arrested zygotes after treatment with 100-μM S3I-201 inhibitor in KSOM-AA embryo culture medium. Chromosomal DNA was stained with DAPI, a blue fluorescent dye. All samples were analyzed under a laser scanning confocal microscope. Scale bar = 20 µm. ( B ) Representative images of the different morphologies of the abnormal metaphase chromosome plates.

    Techniques Used: Embryo Culture, Staining, Microscopy

    33) Product Images from "Towards explicit regulating-ion-transport: nanochannels with only function-elements at outer-surface"

    Article Title: Towards explicit regulating-ion-transport: nanochannels with only function-elements at outer-surface

    Journal: Nature Communications

    doi: 10.1038/s41467-021-21507-7

    Sensing performances of FE OS as probes. a A scheme showing the working mechanism of the sensor using FE OS as probes. b Capture process of multi-scale targets through designed single-stand DNA. c Sensitivity and selectivity of the DNA@OS (DNA is a designed sequence specifically bonding with targets) for the recognition of ions (Hg 2+ ), small molecules (ATP), protein (Lysozyme) and cells (MCF7). The selective detection of multiscale targets using FE OS was realized based on the change of f rec signal output induced by the surface charge at outer surface. LoD is defined as the limitation of detection for the targets. d Formation of “supersandwich” DNA structure (ssw-DNA) with long concatamers through the successive hybridization of alternating DNA unit. And gradual disassembly of ssw-DNA based on interaction between ATP and the repeat DNA units through increasing ATP concentration. e Agarose gel electrophoresis characterizing of the ssw-DNA: 1) DNA marker; 2) p1; 3) p2 (ATP ampter); 4) p1 + p2; 5) target+p1 + p2. f Depth distribution of the “supersandwich” DNA in nanochannel-system using ToF-SIMS. g Laser scanning confocal microscopy of the OS after the assembly (top) and the disassembly of ssw-DNA (bottom). The scale bar is 20 μm. h The LoD of ATP using ssw-DNA as probe based on nanochannel method and electrochemical method (Fig. S 30 ). i Specificity of ssw-DNA@OS for ATP, in contrast with other NTPs. For the sensing performances part, five sensors using FE@OS were established to obtain each error bar of sensitivity and specificity.
    Figure Legend Snippet: Sensing performances of FE OS as probes. a A scheme showing the working mechanism of the sensor using FE OS as probes. b Capture process of multi-scale targets through designed single-stand DNA. c Sensitivity and selectivity of the DNA@OS (DNA is a designed sequence specifically bonding with targets) for the recognition of ions (Hg 2+ ), small molecules (ATP), protein (Lysozyme) and cells (MCF7). The selective detection of multiscale targets using FE OS was realized based on the change of f rec signal output induced by the surface charge at outer surface. LoD is defined as the limitation of detection for the targets. d Formation of “supersandwich” DNA structure (ssw-DNA) with long concatamers through the successive hybridization of alternating DNA unit. And gradual disassembly of ssw-DNA based on interaction between ATP and the repeat DNA units through increasing ATP concentration. e Agarose gel electrophoresis characterizing of the ssw-DNA: 1) DNA marker; 2) p1; 3) p2 (ATP ampter); 4) p1 + p2; 5) target+p1 + p2. f Depth distribution of the “supersandwich” DNA in nanochannel-system using ToF-SIMS. g Laser scanning confocal microscopy of the OS after the assembly (top) and the disassembly of ssw-DNA (bottom). The scale bar is 20 μm. h The LoD of ATP using ssw-DNA as probe based on nanochannel method and electrochemical method (Fig. S 30 ). i Specificity of ssw-DNA@OS for ATP, in contrast with other NTPs. For the sensing performances part, five sensors using FE@OS were established to obtain each error bar of sensitivity and specificity.

    Techniques Used: Sequencing, Hybridization, Concentration Assay, Agarose Gel Electrophoresis, Marker, Confocal Microscopy

    34) Product Images from "OSMR controls glioma stem cell respiration and confers resistance of glioblastoma to ionizing radiation"

    Article Title: OSMR controls glioma stem cell respiration and confers resistance of glioblastoma to ionizing radiation

    Journal: Nature Communications

    doi: 10.1038/s41467-020-17885-z

    Presence of mitochondrial OSMR in human BTSCs. a – d Four different patient-derived BTSC lines were subjected to subcellular fractionation, and the lysates for each fraction were analyzed by immunoblotting using antibodies to OSMR. α-Tubulin, H3K4me3, BCL2/TOM20, Na+/K+ ATPase, and calnexin. WCL: Whole-cell lysates; Cyto: cytoplasmic; Mito: mitochondrial. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC. e , f BTSC73 and BTSC147 were subjected to immunostaining using antibodies to OSMR (green) and the mitochondrial matrix protein ATP synthase inhibitor F1 (ATPIF1, red). Nuclei were stained with DAPI. White rectangles mark the inset to demonstrate the co-localization of OSMR with ATPIF1. g , h PLA of OSMR and ATPIF1 were performed in BTSC73 ( g ) and BTSC147 ( h ). Primary antibodies were omitted as controls. i Double labeling of the PLA signal (red) and the MitoTracker (green) in BTSC73 is shown. j A FRAP assay was performed on BTSC73 transduced with GFP-OSMR and stained with MitoTracker (red). Different regions of interest (ROIs) containing GFP-OSMR in the mitochondria were defined. ROI1 indicates a non-bleached area and ROI2, a photobleached area. The fluorescence recovery was monitored over time following photobleaching. Images were obtained on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm; Inset scale bar = 1 μm. Representative images of three independent experiments are shown.
    Figure Legend Snippet: Presence of mitochondrial OSMR in human BTSCs. a – d Four different patient-derived BTSC lines were subjected to subcellular fractionation, and the lysates for each fraction were analyzed by immunoblotting using antibodies to OSMR. α-Tubulin, H3K4me3, BCL2/TOM20, Na+/K+ ATPase, and calnexin. WCL: Whole-cell lysates; Cyto: cytoplasmic; Mito: mitochondrial. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC. e , f BTSC73 and BTSC147 were subjected to immunostaining using antibodies to OSMR (green) and the mitochondrial matrix protein ATP synthase inhibitor F1 (ATPIF1, red). Nuclei were stained with DAPI. White rectangles mark the inset to demonstrate the co-localization of OSMR with ATPIF1. g , h PLA of OSMR and ATPIF1 were performed in BTSC73 ( g ) and BTSC147 ( h ). Primary antibodies were omitted as controls. i Double labeling of the PLA signal (red) and the MitoTracker (green) in BTSC73 is shown. j A FRAP assay was performed on BTSC73 transduced with GFP-OSMR and stained with MitoTracker (red). Different regions of interest (ROIs) containing GFP-OSMR in the mitochondria were defined. ROI1 indicates a non-bleached area and ROI2, a photobleached area. The fluorescence recovery was monitored over time following photobleaching. Images were obtained on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm; Inset scale bar = 1 μm. Representative images of three independent experiments are shown.

    Techniques Used: Derivative Assay, Fractionation, Western Blot, Immunostaining, Staining, Proximity Ligation Assay, Labeling, FRAP Assay, Transduction, Fluorescence, Microscopy

    OSMR interacts with different components of ETC in human BTSCs. a , b Mitochondrial fractions from BTSC73 ( a ) and BTSC147 ( b ) were treated with 0.5 mg/mL proteinase K or proteinase K and 1% Triton X-100. Lysates were analyzed by immunoblotting using indicated antibodies. c – f WCL and mitochondrial fractions from BTSC73 ( c , d ) and BTSC147 ( e , f ) were subjected to immunoprecipitation using antibodies to OSMR or mouse IgG control, followed by immunoblotting with mtHSP70 and TIM44 antibodies. g , h PLA of OSMR and mtHSP70 were performed in BTSC73 ( g ) and BTSC147 ( h ). Primary antibodies were omitted for the controls. i Double labeling of the PLA signal (red) from the OSMR/mtHSP70 interaction and MitoTracker (green) is shown. j OSMR protein expression level was assessed in the mitochondrial fractions obtained from BTSC73 electroporated with siRNA control (siCTL) or siRNA against mtHSP70 (si mtHSP70 ). BLC2 was used as a loading control. k OSMR protein expression level was assessed in the mitochondrial fractions obtained from BTSC73 electroporated with siCTL or siRNA against TIM44 (si TIM44 ). BCL2 was used as a loading control. l – o WCL or mitochondrial fractions from BTSC73 ( l , m ) and BTSC147 ( n , o ) were subjected to immunoprecipitation using an antibody to OSMR or mouse IgG control followed by immunoblotting with NDUFS1 and NDUFS2 antibodies. p , q PLA analyses of OSMR/NDUFS1 and OSMR/NDUFS2 were carried out in BTSC73 ( p ) and BTSC147 ( q ). r , s Double labeling of the PLA signal (red) and the MitoTracker (green) is shown. Images were obtained with a 63X objectives on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm. Inset scale bar = 1 μm. Representative images of three independent experiments are shown. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC.
    Figure Legend Snippet: OSMR interacts with different components of ETC in human BTSCs. a , b Mitochondrial fractions from BTSC73 ( a ) and BTSC147 ( b ) were treated with 0.5 mg/mL proteinase K or proteinase K and 1% Triton X-100. Lysates were analyzed by immunoblotting using indicated antibodies. c – f WCL and mitochondrial fractions from BTSC73 ( c , d ) and BTSC147 ( e , f ) were subjected to immunoprecipitation using antibodies to OSMR or mouse IgG control, followed by immunoblotting with mtHSP70 and TIM44 antibodies. g , h PLA of OSMR and mtHSP70 were performed in BTSC73 ( g ) and BTSC147 ( h ). Primary antibodies were omitted for the controls. i Double labeling of the PLA signal (red) from the OSMR/mtHSP70 interaction and MitoTracker (green) is shown. j OSMR protein expression level was assessed in the mitochondrial fractions obtained from BTSC73 electroporated with siRNA control (siCTL) or siRNA against mtHSP70 (si mtHSP70 ). BLC2 was used as a loading control. k OSMR protein expression level was assessed in the mitochondrial fractions obtained from BTSC73 electroporated with siCTL or siRNA against TIM44 (si TIM44 ). BCL2 was used as a loading control. l – o WCL or mitochondrial fractions from BTSC73 ( l , m ) and BTSC147 ( n , o ) were subjected to immunoprecipitation using an antibody to OSMR or mouse IgG control followed by immunoblotting with NDUFS1 and NDUFS2 antibodies. p , q PLA analyses of OSMR/NDUFS1 and OSMR/NDUFS2 were carried out in BTSC73 ( p ) and BTSC147 ( q ). r , s Double labeling of the PLA signal (red) and the MitoTracker (green) is shown. Images were obtained with a 63X objectives on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm. Inset scale bar = 1 μm. Representative images of three independent experiments are shown. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC.

    Techniques Used: Immunoprecipitation, Proximity Ligation Assay, Labeling, Expressing, Microscopy, Western Blot

    35) Product Images from "Inhibition of Autophagy Prevents Panax Notoginseng Saponins (PNS) Protection on Cardiac Myocytes Against Endoplasmic Reticulum (ER) Stress-Induced Mitochondrial Injury, Ca2+ Homeostasis and Associated Apoptosis"

    Article Title: Inhibition of Autophagy Prevents Panax Notoginseng Saponins (PNS) Protection on Cardiac Myocytes Against Endoplasmic Reticulum (ER) Stress-Induced Mitochondrial Injury, Ca2+ Homeostasis and Associated Apoptosis

    Journal: Frontiers in Pharmacology

    doi: 10.3389/fphar.2021.620812

    PNS promotes autophagy and autophagic flux. (A) Primary cultured cardiomyocytes, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h (PNS group), before addition of 1 μM thapsigargin (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-LC3B antibody and imaged by a laser scanning confocal microscopy. Scale bar: 30 μm; in box: 10 μm. Bar graph shows the number of LC3 puncta per cell in various groups as indicated (Mean ± SEM; 80–100 cells; * p
    Figure Legend Snippet: PNS promotes autophagy and autophagic flux. (A) Primary cultured cardiomyocytes, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h (PNS group), before addition of 1 μM thapsigargin (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-LC3B antibody and imaged by a laser scanning confocal microscopy. Scale bar: 30 μm; in box: 10 μm. Bar graph shows the number of LC3 puncta per cell in various groups as indicated (Mean ± SEM; 80–100 cells; * p

    Techniques Used: Cell Culture, Confocal Microscopy

    Inhibition of autophagy abolishes PNS protection on TG-induced ER stress response and associated apoptosis. (A) SiNC or siATG7 transfected H9c2 cells, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h before addition of 1 μM TG (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-calnexin antibody and imaged by a laser scanning confocal microscopy. Scale bar: 30 μm; in box: 10 μm. (B) SiNC or siATG7 transfected H9c2 cells, treated as in A, were immunoblotted with the antibodies to BiP, CHOP, Cleaved Caspase-3 and Caspase-12 as well as β -actin. Bands were quantified relative to β -actin by densitometry (Mean ± SEM; ** p
    Figure Legend Snippet: Inhibition of autophagy abolishes PNS protection on TG-induced ER stress response and associated apoptosis. (A) SiNC or siATG7 transfected H9c2 cells, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h before addition of 1 μM TG (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-calnexin antibody and imaged by a laser scanning confocal microscopy. Scale bar: 30 μm; in box: 10 μm. (B) SiNC or siATG7 transfected H9c2 cells, treated as in A, were immunoblotted with the antibodies to BiP, CHOP, Cleaved Caspase-3 and Caspase-12 as well as β -actin. Bands were quantified relative to β -actin by densitometry (Mean ± SEM; ** p

    Techniques Used: Inhibition, Transfection, Confocal Microscopy

    PNS prevents TG-induced mitochondria injury and ROS accumulation. (A) Primary cultured cardiomyocytes, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h (PNS group), before addition of 1 μM thapsigargin (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-TOM20 antibody and imaged by a laser scanning confocal microscopy. Representative images in CN and TG group. Scale bar: 30 μm; in box: 10 μm. Bar graph shows percentage of cardiomyocytes with tubular, intermediate or fragmented mitochondria in various groups as indicated (80–100 cells). (B) Primary cultured cardiomyocytes treated as in A were stained with JC-1 and analyzed by a plate reader. Bar graph shows the ratio of aggregated JC-1 (red)/monomeric JC-1 (green) as the mitochondrial membrane potential (Δ ψ m) (Mean ± SEM; 80–100 cells; * p
    Figure Legend Snippet: PNS prevents TG-induced mitochondria injury and ROS accumulation. (A) Primary cultured cardiomyocytes, either untreated (CN group) or pretreated with 40 μg/ml PNS for 12 h (PNS group), before addition of 1 μM thapsigargin (TG group or PNS plus TG group) for 12 h were immunofluorescenced with the primary anti-TOM20 antibody and imaged by a laser scanning confocal microscopy. Representative images in CN and TG group. Scale bar: 30 μm; in box: 10 μm. Bar graph shows percentage of cardiomyocytes with tubular, intermediate or fragmented mitochondria in various groups as indicated (80–100 cells). (B) Primary cultured cardiomyocytes treated as in A were stained with JC-1 and analyzed by a plate reader. Bar graph shows the ratio of aggregated JC-1 (red)/monomeric JC-1 (green) as the mitochondrial membrane potential (Δ ψ m) (Mean ± SEM; 80–100 cells; * p

    Techniques Used: Cell Culture, Confocal Microscopy, Staining

    36) Product Images from "The Effect of a TLR4 Agonist/Cationic Liposome Adjuvant on Varicella-Zoster Virus Glycoprotein E Vaccine Efficacy: Antigen Presentation, Uptake, and Delivery to Lymph Nodes"

    Article Title: The Effect of a TLR4 Agonist/Cationic Liposome Adjuvant on Varicella-Zoster Virus Glycoprotein E Vaccine Efficacy: Antigen Presentation, Uptake, and Delivery to Lymph Nodes

    Journal: Pharmaceutics

    doi: 10.3390/pharmaceutics13030390

    Cooperative effects of liposomes (LP) and dLOS on the cellular uptake of gE antigen. ( a ) DC2.4 cells were treated with AF488-labeled gE antigen (green) in combination with DiR-labeled liposomes (red) or CIA09 and cultured at 37 °C. Cumulative cellular fluorescence was acquired over 30 min using a laser scanning confocal microscope. The data presented are representative of two experiments with similar results. Scale bar = 20 µm. ( b ) DC2.4 cells were treated with AF647-labeled gE antigen, alone or in combination with dLOS (1 µg/mL), liposomes (50 µg/mL), or both, and cultured at 37 °C for 1, 2, or 3 h followed by flow cytometry. The data presented are representative of three independent experiments with similar results.
    Figure Legend Snippet: Cooperative effects of liposomes (LP) and dLOS on the cellular uptake of gE antigen. ( a ) DC2.4 cells were treated with AF488-labeled gE antigen (green) in combination with DiR-labeled liposomes (red) or CIA09 and cultured at 37 °C. Cumulative cellular fluorescence was acquired over 30 min using a laser scanning confocal microscope. The data presented are representative of two experiments with similar results. Scale bar = 20 µm. ( b ) DC2.4 cells were treated with AF647-labeled gE antigen, alone or in combination with dLOS (1 µg/mL), liposomes (50 µg/mL), or both, and cultured at 37 °C for 1, 2, or 3 h followed by flow cytometry. The data presented are representative of three independent experiments with similar results.

    Techniques Used: Labeling, Cell Culture, Fluorescence, Microscopy, Flow Cytometry

    37) Product Images from "OSMR controls glioma stem cell respiration and confers resistance of glioblastoma to ionizing radiation"

    Article Title: OSMR controls glioma stem cell respiration and confers resistance of glioblastoma to ionizing radiation

    Journal: Nature Communications

    doi: 10.1038/s41467-020-17885-z

    Presence of mitochondrial OSMR in human BTSCs. a – d Four different patient-derived BTSC lines were subjected to subcellular fractionation, and the lysates for each fraction were analyzed by immunoblotting using antibodies to OSMR. α-Tubulin, H3K4me3, BCL2/TOM20, Na+/K+ ATPase, and calnexin. WCL: Whole-cell lysates; Cyto: cytoplasmic; Mito: mitochondrial. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC. e , f BTSC73 and BTSC147 were subjected to immunostaining using antibodies to OSMR (green) and the mitochondrial matrix protein ATP synthase inhibitor F1 (ATPIF1, red). Nuclei were stained with DAPI. White rectangles mark the inset to demonstrate the co-localization of OSMR with ATPIF1. g , h PLA of OSMR and ATPIF1 were performed in BTSC73 ( g ) and BTSC147 ( h ). Primary antibodies were omitted as controls. i Double labeling of the PLA signal (red) and the MitoTracker (green) in BTSC73 is shown. j A FRAP assay was performed on BTSC73 transduced with GFP-OSMR and stained with MitoTracker (red). Different regions of interest (ROIs) containing GFP-OSMR in the mitochondria were defined. ROI1 indicates a non-bleached area and ROI2, a photobleached area. The fluorescence recovery was monitored over time following photobleaching. Images were obtained on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm; Inset scale bar = 1 μm. Representative images of three independent experiments are shown.
    Figure Legend Snippet: Presence of mitochondrial OSMR in human BTSCs. a – d Four different patient-derived BTSC lines were subjected to subcellular fractionation, and the lysates for each fraction were analyzed by immunoblotting using antibodies to OSMR. α-Tubulin, H3K4me3, BCL2/TOM20, Na+/K+ ATPase, and calnexin. WCL: Whole-cell lysates; Cyto: cytoplasmic; Mito: mitochondrial. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC. e , f BTSC73 and BTSC147 were subjected to immunostaining using antibodies to OSMR (green) and the mitochondrial matrix protein ATP synthase inhibitor F1 (ATPIF1, red). Nuclei were stained with DAPI. White rectangles mark the inset to demonstrate the co-localization of OSMR with ATPIF1. g , h PLA of OSMR and ATPIF1 were performed in BTSC73 ( g ) and BTSC147 ( h ). Primary antibodies were omitted as controls. i Double labeling of the PLA signal (red) and the MitoTracker (green) in BTSC73 is shown. j A FRAP assay was performed on BTSC73 transduced with GFP-OSMR and stained with MitoTracker (red). Different regions of interest (ROIs) containing GFP-OSMR in the mitochondria were defined. ROI1 indicates a non-bleached area and ROI2, a photobleached area. The fluorescence recovery was monitored over time following photobleaching. Images were obtained on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm; Inset scale bar = 1 μm. Representative images of three independent experiments are shown.

    Techniques Used: Derivative Assay, Fractionation, Western Blot, Immunostaining, Staining, Proximity Ligation Assay, Labeling, FRAP Assay, Transduction, Fluorescence, Microscopy

    OSMR interacts with different components of ETC in human BTSCs. a , b Mitochondrial fractions from BTSC73 ( a ) and BTSC147 ( b ) were treated with 0.5 mg/mL proteinase K or proteinase K and 1% Triton X-100. Lysates were analyzed by immunoblotting using indicated antibodies. c – f WCL and mitochondrial fractions from BTSC73 ( c , d ) and BTSC147 ( e , f ) were subjected to immunoprecipitation using antibodies to OSMR or mouse IgG control, followed by immunoblotting with mtHSP70 and TIM44 antibodies. g , h PLA of OSMR and mtHSP70 were performed in BTSC73 ( g ) and BTSC147 ( h ). Primary antibodies were omitted for the controls. i Double labeling of the PLA signal (red) from the OSMR/mtHSP70 interaction and MitoTracker (green) is shown. j OSMR protein expression level was assessed in the mitochondrial fractions obtained from BTSC73 electroporated with siRNA control (siCTL) or siRNA against mtHSP70 (si mtHSP70 ). BLC2 was used as a loading control. k OSMR protein expression level was assessed in the mitochondrial fractions obtained from BTSC73 electroporated with siCTL or siRNA against TIM44 (si TIM44 ). BCL2 was used as a loading control. l – o WCL or mitochondrial fractions from BTSC73 ( l , m ) and BTSC147 ( n , o ) were subjected to immunoprecipitation using an antibody to OSMR or mouse IgG control followed by immunoblotting with NDUFS1 and NDUFS2 antibodies. p , q PLA analyses of OSMR/NDUFS1 and OSMR/NDUFS2 were carried out in BTSC73 ( p ) and BTSC147 ( q ). r , s Double labeling of the PLA signal (red) and the MitoTracker (green) is shown. Images were obtained with a 63X objectives on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm. Inset scale bar = 1 μm. Representative images of three independent experiments are shown. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC.
    Figure Legend Snippet: OSMR interacts with different components of ETC in human BTSCs. a , b Mitochondrial fractions from BTSC73 ( a ) and BTSC147 ( b ) were treated with 0.5 mg/mL proteinase K or proteinase K and 1% Triton X-100. Lysates were analyzed by immunoblotting using indicated antibodies. c – f WCL and mitochondrial fractions from BTSC73 ( c , d ) and BTSC147 ( e , f ) were subjected to immunoprecipitation using antibodies to OSMR or mouse IgG control, followed by immunoblotting with mtHSP70 and TIM44 antibodies. g , h PLA of OSMR and mtHSP70 were performed in BTSC73 ( g ) and BTSC147 ( h ). Primary antibodies were omitted for the controls. i Double labeling of the PLA signal (red) from the OSMR/mtHSP70 interaction and MitoTracker (green) is shown. j OSMR protein expression level was assessed in the mitochondrial fractions obtained from BTSC73 electroporated with siRNA control (siCTL) or siRNA against mtHSP70 (si mtHSP70 ). BLC2 was used as a loading control. k OSMR protein expression level was assessed in the mitochondrial fractions obtained from BTSC73 electroporated with siCTL or siRNA against TIM44 (si TIM44 ). BCL2 was used as a loading control. l – o WCL or mitochondrial fractions from BTSC73 ( l , m ) and BTSC147 ( n , o ) were subjected to immunoprecipitation using an antibody to OSMR or mouse IgG control followed by immunoblotting with NDUFS1 and NDUFS2 antibodies. p , q PLA analyses of OSMR/NDUFS1 and OSMR/NDUFS2 were carried out in BTSC73 ( p ) and BTSC147 ( q ). r , s Double labeling of the PLA signal (red) and the MitoTracker (green) is shown. Images were obtained with a 63X objectives on a laser scanning confocal microscope (ZEISS LSM 800). Scale bar = 10 μm. Inset scale bar = 1 μm. Representative images of three independent experiments are shown. The Western blots represent a minimum of three replicates from different passage numbers for each BTSC.

    Techniques Used: Immunoprecipitation, Proximity Ligation Assay, Labeling, Expressing, Microscopy, Western Blot

    38) Product Images from "Inhibition of laser induced rats choroidal neovascularization by intravitreous injection of sEphB4-HSA"

    Article Title: Inhibition of laser induced rats choroidal neovascularization by intravitreous injection of sEphB4-HSA

    Journal: Annals of Translational Medicine

    doi: 10.21037/atm-20-3810

    Effects of the sEphB4 injection on fluorescein leakage and CNV (choroidal neovascularization) lesion in rat. Fluorescein angiogram (FA) was performed 14 days after laser photocoagulation and intravitreal injection of the sEphB4. (A) Representative FA images in laser-induced rat (B) FA score: sEphB4 injection resulted in a dose dependent inhibition of fluorescein leakage CNV (20 FA images were pooled in each group). (C) Representative CNV lesions from PBS or different concentrations of sEphB4-HSA injection measured in FITC-labeled isolectin-B4 stained flat retinal mounts on day 14. (D) After being stained with isolectin B4, retinal flatmounts were visualized with a laser scanning confocal microscope Z stack images of CNV lesion were taken. The image stacks were rendered in 3D using velocity imaging software (Improvision Inc., Waltham, USA) and processed to digitally extract the fluorescent lesion volume. CNV volume was measured in micrometers cubed (14 images/each group), A dose dependent significant reduction of CNV volume was seen in EphB4-HSA treated rats compared with PBS (phosphate-buffered saline) injection.
    Figure Legend Snippet: Effects of the sEphB4 injection on fluorescein leakage and CNV (choroidal neovascularization) lesion in rat. Fluorescein angiogram (FA) was performed 14 days after laser photocoagulation and intravitreal injection of the sEphB4. (A) Representative FA images in laser-induced rat (B) FA score: sEphB4 injection resulted in a dose dependent inhibition of fluorescein leakage CNV (20 FA images were pooled in each group). (C) Representative CNV lesions from PBS or different concentrations of sEphB4-HSA injection measured in FITC-labeled isolectin-B4 stained flat retinal mounts on day 14. (D) After being stained with isolectin B4, retinal flatmounts were visualized with a laser scanning confocal microscope Z stack images of CNV lesion were taken. The image stacks were rendered in 3D using velocity imaging software (Improvision Inc., Waltham, USA) and processed to digitally extract the fluorescent lesion volume. CNV volume was measured in micrometers cubed (14 images/each group), A dose dependent significant reduction of CNV volume was seen in EphB4-HSA treated rats compared with PBS (phosphate-buffered saline) injection.

    Techniques Used: Injection, Inhibition, Labeling, Staining, Microscopy, Imaging, Software

    39) Product Images from "β-Carotene Inhibits Expression of Matrix Metalloproteinase-10 and Invasion in Helicobacter pylori-Infected Gastric Epithelial Cells"

    Article Title: β-Carotene Inhibits Expression of Matrix Metalloproteinase-10 and Invasion in Helicobacter pylori-Infected Gastric Epithelial Cells

    Journal: Molecules

    doi: 10.3390/molecules26061567

    β-Carotene inhibits cell invasion induced by H. pylori in AGS cells. AGS cells were pretreated with β-carotene (0.2 µM) for 2 h and then infected with H. pylori for 24 h. Invasive cells were measured by staining them on Matrigel-coated filters with 4′,6-diamidino-2-phenylindole (DAPI) and visualizing them under a confocal laser scanning microscope (left panel). The graph represents the relative percentage of invasive cells (right panel). All data are shown as the mean ± S.E. of three independent experiments. The percentage of invasive cells in none (the cells without any treatment or infection) was set as 100%. * p
    Figure Legend Snippet: β-Carotene inhibits cell invasion induced by H. pylori in AGS cells. AGS cells were pretreated with β-carotene (0.2 µM) for 2 h and then infected with H. pylori for 24 h. Invasive cells were measured by staining them on Matrigel-coated filters with 4′,6-diamidino-2-phenylindole (DAPI) and visualizing them under a confocal laser scanning microscope (left panel). The graph represents the relative percentage of invasive cells (right panel). All data are shown as the mean ± S.E. of three independent experiments. The percentage of invasive cells in none (the cells without any treatment or infection) was set as 100%. * p

    Techniques Used: Infection, Staining, Laser-Scanning Microscopy

    Related Articles

    Software:

    Article Title: Lysosomotropic drugs activate TFEB via lysosomal membrane fluidization and consequent inhibition of mTORC1 activity
    Article Snippet: Fluorescence images were taken along with the corresponding bright field, diffusion interference contrast image. .. For FRAP measurements, the integrated settings in the Zeiss confocal software were utilized. ..

    Article Title: A TFEB nuclear export signal integrates amino acid supply and glucose availability
    Article Snippet: Coverslips were then mounted on glass microscope slides (VWR) with Vectorshield H-1000 mounting medium (Vector laboratories). .. A Zeiss LSM710 and accompanying Zen software were used to acquire confocal images (Zeiss, Oberkochen, Germany). ..

    Article Title: The Role of the Microglial Cx3cr1 Pathway in the Postnatal Maturation of Retinal Photoreceptors
    Article Snippet: For photoreceptor cilium staining, retinal samples were used unfixed, snap frozen in Tissue-Tek O.C.T. compound (Sakura Finetek USA), and then processed as described above. .. All retinal sections and wholemounts were imaged using a confocal microscope (Meta, Pascal LSM-5 and LSM800, Zen Software, Zeiss) with either a 40× or 60× objective. ..

    Article Title: Reduction in expression of the astrocyte glutamate transporter, GLT1, worsens functional and histological outcomes following traumatic spinal cord injury
    Article Snippet: Images were acquired on either Zeiss Imager Z1 microscope (Roper Scientific; Trenton, NJ) or on a Zeiss laser confocal microscope. .. Images were analyzed using either Metamorph or Zeiss confocal software. .. Adobe Photoshop CS (Adobe, San Jose, CA) was used to prepare figures.

    Article Title: Histone variant H3.3 orchestrates neural stem cell differentiation in the developing brain
    Article Snippet: .. Brain sections (at least nine sections) were analyzed by confocal microscopy, and the GFP-positive cell number or multi-marker co-labeled cells in different regions were counted by the confocal microscopy software (ZEISS Zen 2010, Zeiss, Oberkochen, Germany) and Photoshop CS4 software (Adobe Systems Inc., New York, NY, USA). .. For morphological and quantitative analysis of cultured neurons and IUE brain slides, the number of branches and branch length of the neurons (at least eleven cells) were calculated using confocal microscopy software (ZEISS Zen 2010).

    Article Title: Postnatal Refinement of Auditory Hair Cell Planar Polarity Deficits Occurs in the Absence of Vangl2
    Article Snippet: Tissue was subsequently washed with PBS-T, mounted using Fluoro-Gel (Electron Microscopy Sciences), and imaged using a Carl Zeiss LSM710 confocal microscope. .. Three-dimensional (3D) reconstructions of individual Deiters' cells were generated using Zen software (Carl Zeiss) from confocal stacks. .. The following commercial antibodies were used in this study: Actin (mAB1501; Millipore), β1β2-tubulin (T8535; Sigma), Myosin VIIa (25-6790; Proteus Biosciences), Oncomodulin (SC7446; Santa Cruz Biotechnology), Pericentrin (PRB432C; Covance), Vangl1 (HPA025235; Sigma), Vangl2 (SC46561; Santa Cruz Biotechnology), and Zeb1 (A301–921A; Bethyl).

    Article Title: The secreted cell signal Folded Gastrulation regulates glial morphogenesis and axon guidance in Drosophila
    Article Snippet: For the side view of the CNS in , dissected CNS of stained embryos were mounted on their side and images were taken as a z-series. .. A projected image from the z-series was obtained using the Zeiss confocal software. ..

    other:

    Article Title: Protection from Disulfide Stress by Inhibition of Pap1 Nuclear Export in Schizosaccharomyces pombe
    Article Snippet: 4.8 software) or confocal microscopy (Zeiss LSM 800 confocal microscope with Airyscan using Zen imaging software).

    Microscopy:

    Article Title: The Role of the Microglial Cx3cr1 Pathway in the Postnatal Maturation of Retinal Photoreceptors
    Article Snippet: For photoreceptor cilium staining, retinal samples were used unfixed, snap frozen in Tissue-Tek O.C.T. compound (Sakura Finetek USA), and then processed as described above. .. All retinal sections and wholemounts were imaged using a confocal microscope (Meta, Pascal LSM-5 and LSM800, Zen Software, Zeiss) with either a 40× or 60× objective. ..

    Confocal Microscopy:

    Article Title: Histone variant H3.3 orchestrates neural stem cell differentiation in the developing brain
    Article Snippet: .. Brain sections (at least nine sections) were analyzed by confocal microscopy, and the GFP-positive cell number or multi-marker co-labeled cells in different regions were counted by the confocal microscopy software (ZEISS Zen 2010, Zeiss, Oberkochen, Germany) and Photoshop CS4 software (Adobe Systems Inc., New York, NY, USA). .. For morphological and quantitative analysis of cultured neurons and IUE brain slides, the number of branches and branch length of the neurons (at least eleven cells) were calculated using confocal microscopy software (ZEISS Zen 2010).

    Generated:

    Article Title: Postnatal Refinement of Auditory Hair Cell Planar Polarity Deficits Occurs in the Absence of Vangl2
    Article Snippet: Tissue was subsequently washed with PBS-T, mounted using Fluoro-Gel (Electron Microscopy Sciences), and imaged using a Carl Zeiss LSM710 confocal microscope. .. Three-dimensional (3D) reconstructions of individual Deiters' cells were generated using Zen software (Carl Zeiss) from confocal stacks. .. The following commercial antibodies were used in this study: Actin (mAB1501; Millipore), β1β2-tubulin (T8535; Sigma), Myosin VIIa (25-6790; Proteus Biosciences), Oncomodulin (SC7446; Santa Cruz Biotechnology), Pericentrin (PRB432C; Covance), Vangl1 (HPA025235; Sigma), Vangl2 (SC46561; Santa Cruz Biotechnology), and Zeb1 (A301–921A; Bethyl).

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Carl Zeiss 510 meta zeiss confocal laser scanning microscope
    Molecular analysis of Met kinetic signature- mRNA and protein levels of selected genes in high and low Met expressing cells. (A) Total cellular RNA, was isolated from low (MCF7) and high Met (MDA231) cell cultures and mRNA expression of Met, Survivin, Pbk, Cyclin E1 and Ki67 was evaluated by quantitative real time PCR and compared mRNA levels of the housekeeping GAPDH gene. The primers used for the quantification of gene expression are listed in Table S2 . A gray box denotes MCF7 cell line samples and a black box denotes MDA231 cell line samples (B) Samples from low (MCF7) and high Met (MDA231) cells were subjected to western blot (WB) analysis, before and 15 min and 60 min after treatment with HGF/SF, using antibodies against Met and activated Met (p-Met) and (C) antibodies against ERK K-23, p-ERK E-4, E-Cadherin, Survivin and Actin C4. (D, E) Subcellular localization of survivin in fluorescence (IF) analysis of Low (MCF7) and high Met (MDA231) cells after treatment with HGF/SF at 0 min, 10 min, 30 min and 24 h. The cells were Immunostained using anti-Survivin antibody. Immunofluorescence was examined using a <t>510</t> Meta <t>Zeiss</t> confocal laser scanning microscope (CLSM). Survivin quantification was performed on at least five confocal images per slide. Cell outline was defined based on Nomarski images; nuclei were defined based on the DAPI staining. Average pixel intensity was calculated separately for the nucleus and cytoplasm areas. (F) IF analysis of temporal kinetics of Survivin protein expression following treatment with HGF/SF .
    510 Meta Zeiss Confocal Laser Scanning Microscope, supplied by Carl Zeiss, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/510 meta zeiss confocal laser scanning microscope/product/Carl Zeiss
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    510 meta zeiss confocal laser scanning microscope - by Bioz Stars, 2021-06
    99/100 stars
      Buy from Supplier

    99
    Carl Zeiss lsm 510 meta laser scanning confocal microscope system
    Coro1B Localizes to the Periphery of VSMCs (A) Confocal images were acquired after transfecting VSMCs with siNegCtrl or siCoro1B using the Amaxa electroporation system. Cells were serum starved for 48 hrs and then fixed and permeabilized. Immunofluorescence of Coro1B (red), DAPI (blue) and phalloidin (green) was then detected using the Zeiss <t>LSM</t> 510 <t>META</t> Laser Scanning Confocal Microscope System. (B ) VSMCs were transfected with empty vector or Myc- Coro1B with the Amaxa electroporation system, serum starved for 48 hrs and then fixed and permeabilized. Immunofluorescent images using DAPI (blue), phalloidin (green), Coro1B (red) and Myc (magenta) antibodies were then acquired.
    Lsm 510 Meta Laser Scanning Confocal Microscope System, supplied by Carl Zeiss, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/lsm 510 meta laser scanning confocal microscope system/product/Carl Zeiss
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    lsm 510 meta laser scanning confocal microscope system - by Bioz Stars, 2021-06
    99/100 stars
      Buy from Supplier

    Image Search Results


    Molecular analysis of Met kinetic signature- mRNA and protein levels of selected genes in high and low Met expressing cells. (A) Total cellular RNA, was isolated from low (MCF7) and high Met (MDA231) cell cultures and mRNA expression of Met, Survivin, Pbk, Cyclin E1 and Ki67 was evaluated by quantitative real time PCR and compared mRNA levels of the housekeeping GAPDH gene. The primers used for the quantification of gene expression are listed in Table S2 . A gray box denotes MCF7 cell line samples and a black box denotes MDA231 cell line samples (B) Samples from low (MCF7) and high Met (MDA231) cells were subjected to western blot (WB) analysis, before and 15 min and 60 min after treatment with HGF/SF, using antibodies against Met and activated Met (p-Met) and (C) antibodies against ERK K-23, p-ERK E-4, E-Cadherin, Survivin and Actin C4. (D, E) Subcellular localization of survivin in fluorescence (IF) analysis of Low (MCF7) and high Met (MDA231) cells after treatment with HGF/SF at 0 min, 10 min, 30 min and 24 h. The cells were Immunostained using anti-Survivin antibody. Immunofluorescence was examined using a 510 Meta Zeiss confocal laser scanning microscope (CLSM). Survivin quantification was performed on at least five confocal images per slide. Cell outline was defined based on Nomarski images; nuclei were defined based on the DAPI staining. Average pixel intensity was calculated separately for the nucleus and cytoplasm areas. (F) IF analysis of temporal kinetics of Survivin protein expression following treatment with HGF/SF .

    Journal: PLoS ONE

    Article Title: Met Kinetic Signature Derived from the Response to HGF/SF in a Cellular Model Predicts Breast Cancer Patient Survival

    doi: 10.1371/journal.pone.0045969

    Figure Lengend Snippet: Molecular analysis of Met kinetic signature- mRNA and protein levels of selected genes in high and low Met expressing cells. (A) Total cellular RNA, was isolated from low (MCF7) and high Met (MDA231) cell cultures and mRNA expression of Met, Survivin, Pbk, Cyclin E1 and Ki67 was evaluated by quantitative real time PCR and compared mRNA levels of the housekeeping GAPDH gene. The primers used for the quantification of gene expression are listed in Table S2 . A gray box denotes MCF7 cell line samples and a black box denotes MDA231 cell line samples (B) Samples from low (MCF7) and high Met (MDA231) cells were subjected to western blot (WB) analysis, before and 15 min and 60 min after treatment with HGF/SF, using antibodies against Met and activated Met (p-Met) and (C) antibodies against ERK K-23, p-ERK E-4, E-Cadherin, Survivin and Actin C4. (D, E) Subcellular localization of survivin in fluorescence (IF) analysis of Low (MCF7) and high Met (MDA231) cells after treatment with HGF/SF at 0 min, 10 min, 30 min and 24 h. The cells were Immunostained using anti-Survivin antibody. Immunofluorescence was examined using a 510 Meta Zeiss confocal laser scanning microscope (CLSM). Survivin quantification was performed on at least five confocal images per slide. Cell outline was defined based on Nomarski images; nuclei were defined based on the DAPI staining. Average pixel intensity was calculated separately for the nucleus and cytoplasm areas. (F) IF analysis of temporal kinetics of Survivin protein expression following treatment with HGF/SF .

    Article Snippet: Slides were analyzed using a 510 Meta Zeiss confocal laser scanning microscope (CLSM).

    Techniques: Expressing, Isolation, Real-time Polymerase Chain Reaction, Western Blot, Fluorescence, Immunofluorescence, Laser-Scanning Microscopy, Confocal Laser Scanning Microscopy, Staining

    Characterization of induced ectopic xylem vessel elements. A to H, The excised cotyledons of 6-d-old wild-type seedlings were treated without (A, C, E, and G) or with (B, D, F, and H) KDB for 5 d. Cotyledons were observed using a confocal scanning laser microscope. The cotyledon cells were stained with safranin-O (A and B), probed with the LM10 xylan antibody (C and D), stained with pontamine (E and F), or stained with DAPI (G and H). Yellow arrowheads indicate the nucleus. Bars = 100 µm. I and J, Six-day-old XCP1pro : GUS seedlings were treated without (I) or with (J) KDB for 5 d. Bars = 500 µm.

    Journal: Plant Physiology

    Article Title: Transcription Factors VND1-VND3 Contribute to Cotyledon Xylem Vessel Formation 1Transcription Factors VND1-VND3 Contribute to Cotyledon Xylem Vessel Formation 1 [OPEN]

    doi: 10.1104/pp.17.00461

    Figure Lengend Snippet: Characterization of induced ectopic xylem vessel elements. A to H, The excised cotyledons of 6-d-old wild-type seedlings were treated without (A, C, E, and G) or with (B, D, F, and H) KDB for 5 d. Cotyledons were observed using a confocal scanning laser microscope. The cotyledon cells were stained with safranin-O (A and B), probed with the LM10 xylan antibody (C and D), stained with pontamine (E and F), or stained with DAPI (G and H). Yellow arrowheads indicate the nucleus. Bars = 100 µm. I and J, Six-day-old XCP1pro : GUS seedlings were treated without (I) or with (J) KDB for 5 d. Bars = 500 µm.

    Article Snippet: Xylan and cellulose were observed using Alexa Fluor 488 (wavelength, 493–556 nm) and Alexa Fluor 594 (wavelength, 585–734 nm), respectively, using a confocal laser scanning microscope system (Zeiss LSM 710).

    Techniques: Microscopy, Staining

    The distribution pattern of lamprey Vav3 in VLRB + lymphocytes after stimulation with LPS and PHA. Lymphocytes were isolated from animals before and after treated with LPS or PHA. The cells were first incubated with the rabbit anti-recombinant lamprey Vav3 poly-clonal antibody and the mouse anti-VLRB mono-clonal antibody and then were incubated with Alexa Fluor 555-conjugated goat anti-mouse IgG (red) and Alexa Fluor 488-conjugated goat anti-rabbit IgG (green), as described in Materials and Methods Section. The nuclei were stained with Hoechst 33258 dye (blue). Cells were examined by a Zeiss LSM710 confocal laser scanning microscope. The bar represents 10 μm. The big and round cells (VLRB and lamprey Vav3 double positive) marked with arrows are plasmacytes.

    Journal: International Journal of Molecular Sciences

    Article Title: A Novel Vav3 Homolog Identified in Lamprey, Lampetra japonica, with Roles in Lipopolysaccharide-Mediated Immune Response

    doi: 10.3390/ijms18102035

    Figure Lengend Snippet: The distribution pattern of lamprey Vav3 in VLRB + lymphocytes after stimulation with LPS and PHA. Lymphocytes were isolated from animals before and after treated with LPS or PHA. The cells were first incubated with the rabbit anti-recombinant lamprey Vav3 poly-clonal antibody and the mouse anti-VLRB mono-clonal antibody and then were incubated with Alexa Fluor 555-conjugated goat anti-mouse IgG (red) and Alexa Fluor 488-conjugated goat anti-rabbit IgG (green), as described in Materials and Methods Section. The nuclei were stained with Hoechst 33258 dye (blue). Cells were examined by a Zeiss LSM710 confocal laser scanning microscope. The bar represents 10 μm. The big and round cells (VLRB and lamprey Vav3 double positive) marked with arrows are plasmacytes.

    Article Snippet: Cells were observed by a Zeiss LSM710 Confocal Laser Scanning Microscope (Oberkochen, Germany) and each type of cells (including big and round VLRB+ plasmacytes and small VLRB+ lymphocytes) were counted and analyzed in 5 microscope fields by using Zeiss ZEN LE software.

    Techniques: Isolation, Incubation, Recombinant, Staining, Laser-Scanning Microscopy

    Coro1B Localizes to the Periphery of VSMCs (A) Confocal images were acquired after transfecting VSMCs with siNegCtrl or siCoro1B using the Amaxa electroporation system. Cells were serum starved for 48 hrs and then fixed and permeabilized. Immunofluorescence of Coro1B (red), DAPI (blue) and phalloidin (green) was then detected using the Zeiss LSM 510 META Laser Scanning Confocal Microscope System. (B ) VSMCs were transfected with empty vector or Myc- Coro1B with the Amaxa electroporation system, serum starved for 48 hrs and then fixed and permeabilized. Immunofluorescent images using DAPI (blue), phalloidin (green), Coro1B (red) and Myc (magenta) antibodies were then acquired.

    Journal: Circulation research

    Article Title: Role of Coronin 1B in PDGF-induced Migration of Vascular Smooth Muscle Cells

    doi: 10.1161/CIRCRESAHA.111.255745

    Figure Lengend Snippet: Coro1B Localizes to the Periphery of VSMCs (A) Confocal images were acquired after transfecting VSMCs with siNegCtrl or siCoro1B using the Amaxa electroporation system. Cells were serum starved for 48 hrs and then fixed and permeabilized. Immunofluorescence of Coro1B (red), DAPI (blue) and phalloidin (green) was then detected using the Zeiss LSM 510 META Laser Scanning Confocal Microscope System. (B ) VSMCs were transfected with empty vector or Myc- Coro1B with the Amaxa electroporation system, serum starved for 48 hrs and then fixed and permeabilized. Immunofluorescent images using DAPI (blue), phalloidin (green), Coro1B (red) and Myc (magenta) antibodies were then acquired.

    Article Snippet: Images were acquired with a Zeiss LSM 510 META Laser Scanning Confocal Microscope System using a 63x oil objective lens (numerical aperture: 1.40) and Zeiss ZEN acquisition software.

    Techniques: Electroporation, Immunofluorescence, Microscopy, Transfection, Plasmid Preparation