hoechst 33258 Search Results


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  • 99
    Thermo Fisher hoechst 33258
    JC-10 assay for mitochondrial potential (MP). (A) iCell hepatocytes treated with indicated compounds for 60 min. Top : Cells stained with <t>Hoechst</t> 33258 (nuclei), and JC-10 (MP); bottom : overlay of Granularity analysis result masks. “Total” or “average number of granules per cell,” “total granule area,” or “total granule intensity” were used as output parameters. (B) Concentration-dependent responses of “average number of granules per cell” for several compounds and corresponding IC 50 values (in μM): antimycin A (blue circles), 0.005; CCCP (red squares), 0.047; valinomycin (green squares), 0.001; tolcapone (red triangles), 25.7; nefazodone (white circles), 33.6; leflunomide (purple diamonds), 34.3; ketoconazole (brown triangles), 71.4; isoniazid (grey squares), n/d; fusariotoxin (orange circles), 132; and flutamide (blue triangles), 22.2.
    Hoechst 33258, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 12042 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore bisbenzimide
    ECM rigidity regulates breast epithelial proliferation. T47D cells were cultured in floating and attached 3D collagen gels and allowed to grow for 7 d. The gels were then fixed and costained for Ki67, a marker of proliferation, and <t>bisbenzimide,</t> which will stain all nuclei regardless of cell cycle state. Cells in floating collagen gels have significantly (*P
    Bisbenzimide, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2729 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Beyotime hoechst 33258
    Knockdown of RTKN2 induces cell apoptosis ( A and B ) U2OS cells were stained with Annexin-V FITC/PI, and apoptotic cells were analyzed using flow cytometry. ( C ) Morphological changes associated with apoptosis were examined by <t>Hoechst</t> 33258 staining. ( D ) The apoptotic index was determined by (number of positively stained cells/total number of cells) × 100%. White arrowheads in the images indicate the nuclei of apoptotic cells (magnification, ×200). * P
    Hoechst 33258, supplied by Beyotime, used in various techniques. Bioz Stars score: 94/100, based on 1987 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore hoechst 33258 dye
    Nuclear localization of endogenous PC4, analyzed by immunofluorescence and confocal microscopy. (A) A total of 7 × 10 4 ), followed by incubation with goat anti-rabbit FITC-conjugated antibody. Nuclei were detected by <t>Hoechst</t> 33258 dye (corresponding photomicrographs on the right). Bar, 30 μm. (B) Percentage of cells with endogenous PC4 staining that was cytoplasmic only (black bars), nuclear and cytoplasmic (gray bars), or nuclear only (white bars). Values are calculated for each category as the percentages of the total number of cells scored at each time point (within three fields for each experiment). Means ± the SEM are from three independent experiments. The total number of cells counted for each time point is indicated at the top of the corresponding bar. (C) Confocal microscopy of endogenous PC4. A total of 7 × 10 4 C2C7 cells were seeded onto circular coverslips placed in 35-mm dishes. Cultures were treated, and endogenous PC4 was detected as in panel A. Nuclei are visualized in red by propidium iodide (Pr.i.). The panels on the left (merge) show the overlay of the green and red staining (in orange-green), which indicates the presence of endogenous PC4 protein in the nucleus. Bar, 10 μm.
    Hoechst 33258 Dye, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1312 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Beyotime hoechst 33258 staining
    Apoptosis induction studies of PPI. ( A ) AO\EB staining. ( B ) <t>Hoechst</t> 33258 staining. ( C ) TUNEL assay.
    Hoechst 33258 Staining, supplied by Beyotime, used in various techniques. Bioz Stars score: 92/100, based on 308 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Dojindo Labs hoechst 33258
    Long-term culture hiPS cells by automated system. ( A ) Fold change of cell number determined by split ratio. Black circles indicate each passaging point. ( B ) Fluorescent micrograph of hiPS cells automatically passaged 20 times. Cells were immunologically stained with antibodies against OCT 3/4, NANOG, SOX2, SSEA-4, TRA-1-60, and TRA-1-81. Cell nuclei were stained with Hoechst 33258. Scale bar: 200 μm. ( C ) qPCR analysis of gene expression of OCT 3/4 and NANOG. Expression levels were normalized to expression levels of unpassaged cells (mean ± sd., n = 3). P0, unpassaged cells; P20, cells automatically passaged for 20 times; EB, embryoid body. N.S., not significant. ( D ) Representative histograms of FACS analysis. ( E ) FACS quantification of cells throughout culture. ( F ) Global gene expression analysis. Cluster analysis was performed with all detected genes ( G ) Karyotype analysis of hiPS cells automatically passaged 20 times.
    Hoechst 33258, supplied by Dojindo Labs, used in various techniques. Bioz Stars score: 93/100, based on 468 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Polysciences inc hoechst 33258
    Time course of 8-nitroG formation in In 2 O 3 -treated cells. (A) Immunofluorescent images of 8-nitroG formation in In 2 O 3 -treated cells. RAW 264.7 cells were treated with 20 µg/m l  of In 2 O 3  for indicated durations at 37°C. 8-NitroG formation was detected by immunocytochemistry as described in Materials and Methods section. The nucleus was stained with Hoechst 33258. Magnification, ×200. (B) Quantitative image analysis of 8-nitroG formation in In 2 O 3 -treated cells. The staining intensity per cell was quantified with an ImageJ software. The vertical axis shows the fold increase in the staining intensity compared with that of the control at 2 h, which was set at 1. The data were expressed as means ± SD of 3-4 independent experiments. * p
    Hoechst 33258, supplied by Polysciences inc, used in various techniques. Bioz Stars score: 93/100, based on 295 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Beyotime hoechst 33258 staining solution
    <t>Hoechst</t> 33258 staining in KIAA0100-knockdown U937 cells. Cells were stained with Hoechst 33258 detected and the apoptotic cells were calculated by fluorescent photomicrographs at 200×. The results showed the apoptotic cells were the most in KIAA0100-knockdown U937 cells. The experiments was performed in triplicate and data were displayed as means±SE. **P
    Hoechst 33258 Staining Solution, supplied by Beyotime, used in various techniques. Bioz Stars score: 91/100, based on 361 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Beyotime hoechst 33258 staining kit
    BSO and NAC influence the α-hederin-induced apoptosis of SMMC-7721 cells. SMMC-7721 cells were incubated with α-hederin (10 μmol/L) with or without BSO (2 mmol/L) or NAC (5 mmol/L) pretreatment. A: Cell apoptosis was determined by <t>Hoechst</t> 33258 staining; B: ROS levels in SMMC-7721 cells; C and D: Effect of α-hederin on intracellular GSH and ATP levels. a P
    Hoechst 33258 Staining Kit, supplied by Beyotime, used in various techniques. Bioz Stars score: 89/100, based on 152 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson hoechst 33258
    A correlation between IGF-1R activation at focal adhesion-like structures and EMT transcriptomic signatures in erlotinib-refractory PC-9 cells. Left. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and vinculin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-vinculin antibodies, as specified, and <t>Hoechst</t> 33258 for nuclear counterstaining. Images show representative portions of erlotinib-sensitive PC-9 parental cells and erlotinib-refractory PC-9 POOLs captured in different channels for PP-IGF-1R Tyr1161 ( green ), vinculin ( red ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software. Right. EMT expression profiling of erlotinib-refractory PC-9 pooled cells versus erlotinib-sensitive PC-9 parental cells. Figure shows a representative scatter plot of the difference in the relative abundance of 19 mRNAs whose levels, in published results, are significantly altered during activation/deactivation of the EMT genetic program. Green and red symbols (≥ 2-fold) denote downregulation and upregulation versus basal expression in PC-9 parental cells, respectively; grey symbols denote fold-change results that need to be validated with a sufficient number of biological replicates (i.e., fold-change results may have greater variation of p value > 0.05 or the p value for the fold-change is either unavailable or relatively high [p > 0.05] or, alternatively, they are uninterpretable because the gene's average threshold cycles were either not determined or greater than the defined cut-off value [default 35] in both samples).
    Hoechst 33258, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 347 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Keygen Biotech hoechst 33258
    Icaritin induces K562 cells or primary cells apoptosis. A. Morphological features for apoptosis in untreated and Icaritin-treated K562 cells were revealed by <t>Hoechst</t> 33258 staining. Condensed chromatin and apoptotic body could be found in Icaritin-treated K562 cells by fluorescence microscope (Olympus, B×50); ×400. B. Percentages of apoptotic cells in various concentration of Icaritin-treated K562 cells (Hoechst 33258 staining). The results represent mean±SD of triplicate experiments. C. Flow-cytometry analysis showed externalized PS, revealed by Annexin staining, increased significantly in 32 µM Icaritin-treated K562 cells. D. Percentages of apoptotic cells in Icaritin-treated fresh primary cells from CML-BP patients bone marrow (n = 5) based on annexin V expression assays. Error bars represent SD of experiments. E. Cell cycle analysis showing sub-G1 content in Icaritin (32 µM)-treated cells (right panel) and control (left panel). F. Effects of Icaritin on caspase-9, caspase-3, Apaf-1, bax, bcl-2 and cyt-c protein expression (western blot results). β-actin is used as loading control. G. Effects of Icaritin on apoptosis in Imatinib-resistant cells, CD34+cells from one CML patient with Imatinib-resistance, and CD34+ cells from 3 cases with CML-BC (Annexin V analysis).
    Hoechst 33258, supplied by Keygen Biotech, used in various techniques. Bioz Stars score: 92/100, based on 94 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Nacalai hoechst 33258
    Immunohistochemical staining and electron micrographs of Sox17-transduced ball-like cell clusters. (A) Expression of CD31 and c-Kit was observed in Sox17-transduced ball-like cell clusters. After Cytospin centrifugation, ball-like cell clusters were stained with anti-CD31 (red) or anti-c-Kit (red) antibodies. Nuclei were stained with <t>Hoechst</t> 33258 (blue). Images on the right are higher magnifications of the boxed areas in the images on the left. Bar, 100 μm. (B) Electron micrographs of IRES-GFP (Mock)- and Sox17-IRES-GFP-transduced cells. Bar, 10 μm.
    Hoechst 33258, supplied by Nacalai, used in various techniques. Bioz Stars score: 94/100, based on 268 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    JC-10 assay for mitochondrial potential (MP). (A) iCell hepatocytes treated with indicated compounds for 60 min. Top : Cells stained with Hoechst 33258 (nuclei), and JC-10 (MP); bottom : overlay of Granularity analysis result masks. “Total” or “average number of granules per cell,” “total granule area,” or “total granule intensity” were used as output parameters. (B) Concentration-dependent responses of “average number of granules per cell” for several compounds and corresponding IC 50 values (in μM): antimycin A (blue circles), 0.005; CCCP (red squares), 0.047; valinomycin (green squares), 0.001; tolcapone (red triangles), 25.7; nefazodone (white circles), 33.6; leflunomide (purple diamonds), 34.3; ketoconazole (brown triangles), 71.4; isoniazid (grey squares), n/d; fusariotoxin (orange circles), 132; and flutamide (blue triangles), 22.2.

    Journal: Assay and Drug Development Technologies

    Article Title: High-Content Assays for Hepatotoxicity Using Induced Pluripotent Stem Cell–Derived Cells

    doi: 10.1089/adt.2013.520

    Figure Lengend Snippet: JC-10 assay for mitochondrial potential (MP). (A) iCell hepatocytes treated with indicated compounds for 60 min. Top : Cells stained with Hoechst 33258 (nuclei), and JC-10 (MP); bottom : overlay of Granularity analysis result masks. “Total” or “average number of granules per cell,” “total granule area,” or “total granule intensity” were used as output parameters. (B) Concentration-dependent responses of “average number of granules per cell” for several compounds and corresponding IC 50 values (in μM): antimycin A (blue circles), 0.005; CCCP (red squares), 0.047; valinomycin (green squares), 0.001; tolcapone (red triangles), 25.7; nefazodone (white circles), 33.6; leflunomide (purple diamonds), 34.3; ketoconazole (brown triangles), 71.4; isoniazid (grey squares), n/d; fusariotoxin (orange circles), 132; and flutamide (blue triangles), 22.2.

    Article Snippet: For live cell imaging assays, cell media was removed and cells incubated with a mixture of the following reagents in phosphate-buffered saline (PBS): 1 μM Calcein AM, 2 μg/mL Hoechst 33258, and 200 nM of MitoTracker Orange for 30 min at which point the staining solution was replaced with PBS containing 0.1% of fetal bovine serum (FBS; Gibco, Invitrogen) and images were acquired using the ImageXpress® Micro XL system (Molecular Devices, LLC, Sunnyvale, CA) with a 10× or 4× objective.

    Techniques: Staining, Concentration Assay

    Cytoskeleton integrity assessment by actin staining.  (A)  iCell hepatocytes treated with 30 mM of aflatoxin B1 for 72 h, then fixed and stained with AlexaFluor-488-Phalloidin and Hoechst 33258.  (B)  Concentration-dependent responses for several compounds using “number of actin-positive cells” as a readout. The IC 50 s for the tested compounds were aflatoxin G1 (red squares), 1.9 μM; staurosporine (orange diamonds), 2.0 μM; aflatoxin B1 (blue circles), 5.4 μM; idarubicin (grey squares), 7.4 μM; mitomycin C (green triangles), 19 μM; and retinoic acid (purple circles), 480 μM.

    Journal: Assay and Drug Development Technologies

    Article Title: High-Content Assays for Hepatotoxicity Using Induced Pluripotent Stem Cell–Derived Cells

    doi: 10.1089/adt.2013.520

    Figure Lengend Snippet: Cytoskeleton integrity assessment by actin staining. (A) iCell hepatocytes treated with 30 mM of aflatoxin B1 for 72 h, then fixed and stained with AlexaFluor-488-Phalloidin and Hoechst 33258. (B) Concentration-dependent responses for several compounds using “number of actin-positive cells” as a readout. The IC 50 s for the tested compounds were aflatoxin G1 (red squares), 1.9 μM; staurosporine (orange diamonds), 2.0 μM; aflatoxin B1 (blue circles), 5.4 μM; idarubicin (grey squares), 7.4 μM; mitomycin C (green triangles), 19 μM; and retinoic acid (purple circles), 480 μM.

    Article Snippet: For live cell imaging assays, cell media was removed and cells incubated with a mixture of the following reagents in phosphate-buffered saline (PBS): 1 μM Calcein AM, 2 μg/mL Hoechst 33258, and 200 nM of MitoTracker Orange for 30 min at which point the staining solution was replaced with PBS containing 0.1% of fetal bovine serum (FBS; Gibco, Invitrogen) and images were acquired using the ImageXpress® Micro XL system (Molecular Devices, LLC, Sunnyvale, CA) with a 10× or 4× objective.

    Techniques: Staining, Concentration Assay

    Live cell imaging toxicity assessment. (A) iCell hepatocytes treated with indicated concentrations of amitryptiline for 72 h, then stained with Calcein AM, Hoechst 33258, and MitoTracker Orange. Images were taken using a 10× objective and analyzed using Multi-Wavelength Cell Scoring module. (B) Image processing by Multi-Wavelength Cell Scoring application module. Masks shown define Calcein AM staining (green), MitoTracker staining (orange), and Hoechst 33258 staining (grey). (C) Comparison of IC 50 s obtained by using total cells (nuclear count, blue circles), Calcein AM–positive cells (green triangles), or MitoTracker Orange–positive cells (red squares). NT, no significant toxicity detected from the four-parameter curve fit to the data.

    Journal: Assay and Drug Development Technologies

    Article Title: High-Content Assays for Hepatotoxicity Using Induced Pluripotent Stem Cell–Derived Cells

    doi: 10.1089/adt.2013.520

    Figure Lengend Snippet: Live cell imaging toxicity assessment. (A) iCell hepatocytes treated with indicated concentrations of amitryptiline for 72 h, then stained with Calcein AM, Hoechst 33258, and MitoTracker Orange. Images were taken using a 10× objective and analyzed using Multi-Wavelength Cell Scoring module. (B) Image processing by Multi-Wavelength Cell Scoring application module. Masks shown define Calcein AM staining (green), MitoTracker staining (orange), and Hoechst 33258 staining (grey). (C) Comparison of IC 50 s obtained by using total cells (nuclear count, blue circles), Calcein AM–positive cells (green triangles), or MitoTracker Orange–positive cells (red squares). NT, no significant toxicity detected from the four-parameter curve fit to the data.

    Article Snippet: For live cell imaging assays, cell media was removed and cells incubated with a mixture of the following reagents in phosphate-buffered saline (PBS): 1 μM Calcein AM, 2 μg/mL Hoechst 33258, and 200 nM of MitoTracker Orange for 30 min at which point the staining solution was replaced with PBS containing 0.1% of fetal bovine serum (FBS; Gibco, Invitrogen) and images were acquired using the ImageXpress® Micro XL system (Molecular Devices, LLC, Sunnyvale, CA) with a 10× or 4× objective.

    Techniques: Live Cell Imaging, Staining

    Phenotypic characterization of hepatotoxicity by quantitation of the average and total stained cell areas. Images of iCell hepatocytes treated with 100 μM of indicated compounds for 72 h, then stained with Calcein AM and Hoechst 33258. Examples presented show impact of different compounds on total positive cell area and/or average positive cell area. Images were acquired using a 10× objective.

    Journal: Assay and Drug Development Technologies

    Article Title: High-Content Assays for Hepatotoxicity Using Induced Pluripotent Stem Cell–Derived Cells

    doi: 10.1089/adt.2013.520

    Figure Lengend Snippet: Phenotypic characterization of hepatotoxicity by quantitation of the average and total stained cell areas. Images of iCell hepatocytes treated with 100 μM of indicated compounds for 72 h, then stained with Calcein AM and Hoechst 33258. Examples presented show impact of different compounds on total positive cell area and/or average positive cell area. Images were acquired using a 10× objective.

    Article Snippet: For live cell imaging assays, cell media was removed and cells incubated with a mixture of the following reagents in phosphate-buffered saline (PBS): 1 μM Calcein AM, 2 μg/mL Hoechst 33258, and 200 nM of MitoTracker Orange for 30 min at which point the staining solution was replaced with PBS containing 0.1% of fetal bovine serum (FBS; Gibco, Invitrogen) and images were acquired using the ImageXpress® Micro XL system (Molecular Devices, LLC, Sunnyvale, CA) with a 10× or 4× objective.

    Techniques: Quantitation Assay, Staining

    Phenotypic characterization of toxicity by nuclear shape and intensity.  (A)  Images of iCell hepatocytes treated with 10 mM of staurosporine, stained with Hoechst 33258. Images were acquired using a 10× objective.  (B)  Concentration-dependent responses for several compounds using “average nuclear area” as a readout. The IC 50 s for the tested compounds were aflatoxin G1 (red squares), 1.9 μM; staurosporine (orange diamonds), 1.9 μM; aflatoxin B1 (blue circles), 5.5 μM; idarubicin (grey squares), 7.4 μM, mitomycin C (green triangles), 18 μM; and retinoic acid (purple circles), 482 μM.

    Journal: Assay and Drug Development Technologies

    Article Title: High-Content Assays for Hepatotoxicity Using Induced Pluripotent Stem Cell–Derived Cells

    doi: 10.1089/adt.2013.520

    Figure Lengend Snippet: Phenotypic characterization of toxicity by nuclear shape and intensity. (A) Images of iCell hepatocytes treated with 10 mM of staurosporine, stained with Hoechst 33258. Images were acquired using a 10× objective. (B) Concentration-dependent responses for several compounds using “average nuclear area” as a readout. The IC 50 s for the tested compounds were aflatoxin G1 (red squares), 1.9 μM; staurosporine (orange diamonds), 1.9 μM; aflatoxin B1 (blue circles), 5.5 μM; idarubicin (grey squares), 7.4 μM, mitomycin C (green triangles), 18 μM; and retinoic acid (purple circles), 482 μM.

    Article Snippet: For live cell imaging assays, cell media was removed and cells incubated with a mixture of the following reagents in phosphate-buffered saline (PBS): 1 μM Calcein AM, 2 μg/mL Hoechst 33258, and 200 nM of MitoTracker Orange for 30 min at which point the staining solution was replaced with PBS containing 0.1% of fetal bovine serum (FBS; Gibco, Invitrogen) and images were acquired using the ImageXpress® Micro XL system (Molecular Devices, LLC, Sunnyvale, CA) with a 10× or 4× objective.

    Techniques: Staining, Concentration Assay

    Confocal images of fibers and scaffold encapsulated with PEI–DNA nanoparticles. PEI–DNA nanoparticles encapsulated within the fibers were fluorescently stained with Hoechst 33258 dye. (A) Phase microscopy of the bead region of a single

    Journal:

    Article Title: Nonviral Gene Delivery from Nonwoven Fibrous Scaffolds Fabricated by Interfacial Complexation of Polyelectrolytes

    doi: 10.1016/j.ymthe.2005.12.016

    Figure Lengend Snippet: Confocal images of fibers and scaffold encapsulated with PEI–DNA nanoparticles. PEI–DNA nanoparticles encapsulated within the fibers were fluorescently stained with Hoechst 33258 dye. (A) Phase microscopy of the bead region of a single

    Article Snippet: To confirm the presence of nanoparticles, single fibers as well as scaffold samples were stained with Hoechst 33258 dye (Molecular Probes) and observed using a Nikon TE3000 Eclipse inverted microscope equipped with a UV filter (350/450 nm).

    Techniques: Staining, Microscopy

    Long-lasting beneficial effects of MT5-MMP deficiency in the brains of TgMT5 −/− mice. Representative confocal microphotographs of seven mice per group and the corresponding quantifications showing strong reductions in plaque load (6E10, red ) and microglial reactivity (Iba1, green ) in the neocortex ( a ) and hippocampus ( b ) of 16-month-old TgMT5 −/− mice compared to age-matched Tg mice. The nuclear marker Hoechst is in blue . Scale bar 200 µm. NCx neocortex, Sub subiculum, DG dentate gyrus. c Representative confocal microphotographs showing a better-preserved neuronal network (MAP-2, red ) and increased synaptophysin immunostaining (Syn, green ) in layers IV–V of the neocortex. Scale bar 20 µm. d Western blot of cortical soluble fractions showing increased levels of synaptophysin in TgMT5 −/− mice compared to Tg. Values are the mean ± SEM of seven actin-normalized optical densities (O.D.); ** p

    Journal: Cellular and Molecular Life Sciences

    Article Title: MT5-MMP is a new pro-amyloidogenic proteinase that promotes amyloid pathology and cognitive decline in a transgenic mouse model of Alzheimer’s disease

    doi: 10.1007/s00018-015-1992-1

    Figure Lengend Snippet: Long-lasting beneficial effects of MT5-MMP deficiency in the brains of TgMT5 −/− mice. Representative confocal microphotographs of seven mice per group and the corresponding quantifications showing strong reductions in plaque load (6E10, red ) and microglial reactivity (Iba1, green ) in the neocortex ( a ) and hippocampus ( b ) of 16-month-old TgMT5 −/− mice compared to age-matched Tg mice. The nuclear marker Hoechst is in blue . Scale bar 200 µm. NCx neocortex, Sub subiculum, DG dentate gyrus. c Representative confocal microphotographs showing a better-preserved neuronal network (MAP-2, red ) and increased synaptophysin immunostaining (Syn, green ) in layers IV–V of the neocortex. Scale bar 20 µm. d Western blot of cortical soluble fractions showing increased levels of synaptophysin in TgMT5 −/− mice compared to Tg. Values are the mean ± SEM of seven actin-normalized optical densities (O.D.); ** p

    Article Snippet: Nuclei were stained with Hoechst #33258 (0.5 µg/ml, Life Technologies).

    Techniques: Mouse Assay, Marker, Immunostaining, Western Blot

    Decreased glial reactivity and IL-1β levels in the brains of TgMT5 −/− mice. a Representative epifluorescence microphotographs showing changes in astroglia (GFAP) and microglia (Iba1) reactivity in the hippocampus (Hc) and neocortex (NCx) of Tg and TgMT5 −/− mice, counterstained with nuclear marker Hoechst ( blue ). Scale bar 1 mm. b Quantification of changes in GFAP and Iba1 immunostaining measured as the percentage of the immunostained area/total area of the hippocampus or neocortex. Values are the mean ± SEM of seven brains per group; ** p

    Journal: Cellular and Molecular Life Sciences

    Article Title: MT5-MMP is a new pro-amyloidogenic proteinase that promotes amyloid pathology and cognitive decline in a transgenic mouse model of Alzheimer’s disease

    doi: 10.1007/s00018-015-1992-1

    Figure Lengend Snippet: Decreased glial reactivity and IL-1β levels in the brains of TgMT5 −/− mice. a Representative epifluorescence microphotographs showing changes in astroglia (GFAP) and microglia (Iba1) reactivity in the hippocampus (Hc) and neocortex (NCx) of Tg and TgMT5 −/− mice, counterstained with nuclear marker Hoechst ( blue ). Scale bar 1 mm. b Quantification of changes in GFAP and Iba1 immunostaining measured as the percentage of the immunostained area/total area of the hippocampus or neocortex. Values are the mean ± SEM of seven brains per group; ** p

    Article Snippet: Nuclei were stained with Hoechst #33258 (0.5 µg/ml, Life Technologies).

    Techniques: Mouse Assay, Marker, Immunostaining

    Decreased Hsp70-2 protein levels in Bat3 −/− male GCs. (A–J) P42 Bat3 +/+ and Bat3 −/− testis sections were stained with Hsp70-2 (A–D) and Bat3 (E and F) antibodies. Hoechst33258 for DNA staining (I and J). Merged images (G and H). (K–V) Defective accumulation of Hsp70-2 at an early stage of spermatogenesis. P7 (K–P) and P14 (Q–V) Bat3 +/+ and Bat3 −/− testis sections were stained using Hsp70-2 (K, L, Q, and R) and Bat3 (M, N, S, and T) antibodies. DNA was stained by Hoechst 33258 (O, P, U, and V). (W and X) Bat3 and Hsp70-2 mRNA (W) and protein (X) levels in Bat3 +/+ and Bat3 −/− male GCs were examined at the indicated developmental stages. Bars: (B) 100 μm; (D, L, and R) 20 μm.

    Journal: The Journal of Cell Biology

    Article Title: Bat3 deficiency accelerates the degradation of Hsp70-2/HspA2 during spermatogenesis

    doi: 10.1083/jcb.200802113

    Figure Lengend Snippet: Decreased Hsp70-2 protein levels in Bat3 −/− male GCs. (A–J) P42 Bat3 +/+ and Bat3 −/− testis sections were stained with Hsp70-2 (A–D) and Bat3 (E and F) antibodies. Hoechst33258 for DNA staining (I and J). Merged images (G and H). (K–V) Defective accumulation of Hsp70-2 at an early stage of spermatogenesis. P7 (K–P) and P14 (Q–V) Bat3 +/+ and Bat3 −/− testis sections were stained using Hsp70-2 (K, L, Q, and R) and Bat3 (M, N, S, and T) antibodies. DNA was stained by Hoechst 33258 (O, P, U, and V). (W and X) Bat3 and Hsp70-2 mRNA (W) and protein (X) levels in Bat3 +/+ and Bat3 −/− male GCs were examined at the indicated developmental stages. Bars: (B) 100 μm; (D, L, and R) 20 μm.

    Article Snippet: Hoechst33258 (Invitrogen) was used to visualize nuclei.

    Techniques: Staining

    ECM rigidity regulates breast epithelial proliferation. T47D cells were cultured in floating and attached 3D collagen gels and allowed to grow for 7 d. The gels were then fixed and costained for Ki67, a marker of proliferation, and bisbenzimide, which will stain all nuclei regardless of cell cycle state. Cells in floating collagen gels have significantly (*P

    Journal: The Journal of Cell Biology

    Article Title: ROCK-generated contractility regulates breast epithelial cell differentiation in response to the physical properties of a three-dimensional collagen matrix

    doi: 10.1083/jcb.200305010

    Figure Lengend Snippet: ECM rigidity regulates breast epithelial proliferation. T47D cells were cultured in floating and attached 3D collagen gels and allowed to grow for 7 d. The gels were then fixed and costained for Ki67, a marker of proliferation, and bisbenzimide, which will stain all nuclei regardless of cell cycle state. Cells in floating collagen gels have significantly (*P

    Article Snippet: After extensive washing in PBS, 100 μl anti–rabbit TRITC (1:100; Jackson ImmunoResearch Laboratories) or anti–mouse TRITC (1:100; Jackson ImmunoResearch Laboratories) with phalloidin-FITC (1:1000; Jackson ImmunoResearch Laboratories), Bisbenzimide (Sigma-Aldrich), and 1% donkey serum in PBS was added for 30 min at room temperature.

    Techniques: Cell Culture, Marker, Staining

    Active intestinal epithelial stem cells [aISC, sex determining region Y-box 9 (SOX9 + )] are less resistant to ischemic injury than reserve intestinal epithelial stem cells [rISC, homeodomain only protein X (HOPX) + ] and undergo apoptosis following prolonged ischemic injury. Fluorescent multiplex immunohistochemistry with tyramide signal amplification (TSA) was used to image each ISC population and colocalize aISC and rISC with cleaved caspase 3 (CC3) in tissues subjected to 1–4 h of ischemia ( n  = 4).  A : SOX9 +  cells (red) within the crypt base of control 2- and 4-h ischemic tissue.  B : HOPX +  cells (white) within the crypt base of control 2- and 4-h ischemic tissue.  C  and  E : a progressive increase of SOX9 + CC3 +  colocalization with increasing duration of ischemic injury that reached statistical significance by 4 h ischemia was observed (dotted line outlines colocalized SOX9 + CC3 +  cell).  D  and  E : consistent with the hypothesis that rISC are resistant to injury, we noted a complete lack of HOPX + CC3 +  coexpression in short-duration ischemia (1 or 2 h) and no significant coexpression in the prolonged ischemic injured intestine (dotted line outlines colocalized HOPX + CC3 +  cell). Nuclei were stained with bisbenzimide H 33258 (blue). Values are means ± SD  a Statistical significant difference from control,  P

    Journal: American Journal of Physiology - Gastrointestinal and Liver Physiology

    Article Title: The Engineered Gut: Use of Stem Cells and Tissue Engineering to Study Physiological Mechanisms and Disease Processes: Preservation of reserve intestinal epithelial stem cells following severe ischemic injury

    doi: 10.1152/ajpgi.00262.2018

    Figure Lengend Snippet: Active intestinal epithelial stem cells [aISC, sex determining region Y-box 9 (SOX9 + )] are less resistant to ischemic injury than reserve intestinal epithelial stem cells [rISC, homeodomain only protein X (HOPX) + ] and undergo apoptosis following prolonged ischemic injury. Fluorescent multiplex immunohistochemistry with tyramide signal amplification (TSA) was used to image each ISC population and colocalize aISC and rISC with cleaved caspase 3 (CC3) in tissues subjected to 1–4 h of ischemia ( n = 4). A : SOX9 + cells (red) within the crypt base of control 2- and 4-h ischemic tissue. B : HOPX + cells (white) within the crypt base of control 2- and 4-h ischemic tissue. C and E : a progressive increase of SOX9 + CC3 + colocalization with increasing duration of ischemic injury that reached statistical significance by 4 h ischemia was observed (dotted line outlines colocalized SOX9 + CC3 + cell). D and E : consistent with the hypothesis that rISC are resistant to injury, we noted a complete lack of HOPX + CC3 + coexpression in short-duration ischemia (1 or 2 h) and no significant coexpression in the prolonged ischemic injured intestine (dotted line outlines colocalized HOPX + CC3 + cell). Nuclei were stained with bisbenzimide H 33258 (blue). Values are means ± SD a Statistical significant difference from control, P

    Article Snippet: Following washing, nuclear counterstaining was performed with bisbenzimide H 33258 (1:500; Sigma).

    Techniques: Fluorescent Multiplex Immunohistochemistry, Amplification, Staining

    Increasing durations of ischemia differentially impact intestinal epithelial stem cell (ISC) populations. Immunohistochemical analysis of tissues obtained from in vivo mesenteric vascular occlusion for 1–4 h and evaluated for expression of proliferating cell nuclear antigen (PCNA, green), Ki-67 (green), sex determining region Y-box 9 (SOX9, red), homeodomain only protein X (HOPX, green), and cleaved caspase 3 (CC3, red) ( n  = 4–7).  A  and  B : following prolonged ischemic injury (3 and 4 h), crypt damage with concomitant epithelial cell loss was apparent with decreased numbers of cells expressing PCNA and Ki-67.  C – E : consistent with the hypothesis that reserve intestinal epithelial stem cell (rISC) are more resistant to injury, the number of SOX9 +  cells and SOX9 + Ki-67 +  were decreased following 3 h and 4 h ischemia, whereas the number of cells expressing the rISC biomarker HOPX remained unchanged despite the duration of ischemic injury.  F : a significantly increased number of crypt cells expressed CC3, but only in 3 and 4 h of ischemia, indicating that the extensive cell loss observed with prolonged ischemia was in large part attributable to apoptosis. Nuclei were stained with bisbenzimide H 33258 (blue). Dotted line outlines crypts extending down to the muscularis mucosa. Values are means ± SD.  a Statistical significant difference from control,  P

    Journal: American Journal of Physiology - Gastrointestinal and Liver Physiology

    Article Title: The Engineered Gut: Use of Stem Cells and Tissue Engineering to Study Physiological Mechanisms and Disease Processes: Preservation of reserve intestinal epithelial stem cells following severe ischemic injury

    doi: 10.1152/ajpgi.00262.2018

    Figure Lengend Snippet: Increasing durations of ischemia differentially impact intestinal epithelial stem cell (ISC) populations. Immunohistochemical analysis of tissues obtained from in vivo mesenteric vascular occlusion for 1–4 h and evaluated for expression of proliferating cell nuclear antigen (PCNA, green), Ki-67 (green), sex determining region Y-box 9 (SOX9, red), homeodomain only protein X (HOPX, green), and cleaved caspase 3 (CC3, red) ( n = 4–7). A and B : following prolonged ischemic injury (3 and 4 h), crypt damage with concomitant epithelial cell loss was apparent with decreased numbers of cells expressing PCNA and Ki-67. C – E : consistent with the hypothesis that reserve intestinal epithelial stem cell (rISC) are more resistant to injury, the number of SOX9 + cells and SOX9 + Ki-67 + were decreased following 3 h and 4 h ischemia, whereas the number of cells expressing the rISC biomarker HOPX remained unchanged despite the duration of ischemic injury. F : a significantly increased number of crypt cells expressed CC3, but only in 3 and 4 h of ischemia, indicating that the extensive cell loss observed with prolonged ischemia was in large part attributable to apoptosis. Nuclei were stained with bisbenzimide H 33258 (blue). Dotted line outlines crypts extending down to the muscularis mucosa. Values are means ± SD. a Statistical significant difference from control, P

    Article Snippet: Following washing, nuclear counterstaining was performed with bisbenzimide H 33258 (1:500; Sigma).

    Techniques: Immunohistochemistry, In Vivo, Expressing, Biomarker Assay, Staining

    Subregion-specific patterns of gene expression within the hippocampus I. A – E, U – Y , Genes enriched in the dentate gyrus ( A–C ), CA1–CA3 ( D, E ), CA3 or CA2 plus CA3 (CA2/3; U–W ), and the dentate gyrus plus CA1 ( X, Y ). A – J, U – DD, In situ hybridization for Dsp ( A, F ), Mcmd6 ( B, G ), calretinin ( C,H ), Lpl ( D, I ), Mrg1b ( E, J ), Bok ( U,Z ) Socs2 ( V,AA ), Cadps ( W,BB ), calbindin ( X, CC ), and Ngef ( Y,DD ) on coronal sections through the hippocampus at low magnification ( A–E, U–Y ) and high magnification ( F–J, Z–DD ). Bisbenzimide staining of the same sections is shown in K–O and EE–II , and gene chip data for each gene are shown in P–T and JJ–NN . Arrowheads in F–O and Z–II delimit the boundaries of CA1 and also the boundary between CA2 and CA3 in Z and EE . The arrow in H denotes the hilus of the dentate gyrus. Scale bars, 1 mm.

    Journal: The Journal of Neuroscience

    Article Title: Defining a Molecular Atlas of the Hippocampus Using DNA Microarrays and High-Throughput In Situ Hybridization

    doi: 10.1523/JNEUROSCI.4710-03.2004

    Figure Lengend Snippet: Subregion-specific patterns of gene expression within the hippocampus I. A – E, U – Y , Genes enriched in the dentate gyrus ( A–C ), CA1–CA3 ( D, E ), CA3 or CA2 plus CA3 (CA2/3; U–W ), and the dentate gyrus plus CA1 ( X, Y ). A – J, U – DD, In situ hybridization for Dsp ( A, F ), Mcmd6 ( B, G ), calretinin ( C,H ), Lpl ( D, I ), Mrg1b ( E, J ), Bok ( U,Z ) Socs2 ( V,AA ), Cadps ( W,BB ), calbindin ( X, CC ), and Ngef ( Y,DD ) on coronal sections through the hippocampus at low magnification ( A–E, U–Y ) and high magnification ( F–J, Z–DD ). Bisbenzimide staining of the same sections is shown in K–O and EE–II , and gene chip data for each gene are shown in P–T and JJ–NN . Arrowheads in F–O and Z–II delimit the boundaries of CA1 and also the boundary between CA2 and CA3 in Z and EE . The arrow in H denotes the hilus of the dentate gyrus. Scale bars, 1 mm.

    Article Snippet: Slides were then developed and stained with the fluorescent dye bisbenzimide (Sigma) to allow localization of silver grains to individual cells.

    Techniques: Expressing, In Situ Hybridization, Staining, Chromatin Immunoprecipitation

    Subregion-specific patterns of gene expression within the hippocampus II. A – D, Q – T , Genes enriched in CA1 ( A, B ), the dentate gyrus plus CA2/3 ( C,D ), the dentate gyrus plus CA2 ( Q,R ), and CA1 plus CA3 ( S,T ). A – H, Q – X, In situ hybridization for Nov ( A, E ), Etv1 ( B, F ), Tgfb2 ( C,G ), Frzb ( D, H ), Pcp4 ( Q,U ), Tiam1 ( R, V ), Tyro3 ( S,W ), and Prss19 ( T,X ) on coronal sections through the hippocampus at low magnification ( A–D, Q–T ) and high magnification ( E–H, U–X ). Bisbenzimide staining of the same sections is shown in I–L and Y–BB , and gene chip data for each gene are shown in M–P and CC–FF . Arrowheads in E–L and U–BB delimit the boundaries of CA1 and also the boundary between CA2 and CA3 in U–BB . Scale bars, 1 mm.

    Journal: The Journal of Neuroscience

    Article Title: Defining a Molecular Atlas of the Hippocampus Using DNA Microarrays and High-Throughput In Situ Hybridization

    doi: 10.1523/JNEUROSCI.4710-03.2004

    Figure Lengend Snippet: Subregion-specific patterns of gene expression within the hippocampus II. A – D, Q – T , Genes enriched in CA1 ( A, B ), the dentate gyrus plus CA2/3 ( C,D ), the dentate gyrus plus CA2 ( Q,R ), and CA1 plus CA3 ( S,T ). A – H, Q – X, In situ hybridization for Nov ( A, E ), Etv1 ( B, F ), Tgfb2 ( C,G ), Frzb ( D, H ), Pcp4 ( Q,U ), Tiam1 ( R, V ), Tyro3 ( S,W ), and Prss19 ( T,X ) on coronal sections through the hippocampus at low magnification ( A–D, Q–T ) and high magnification ( E–H, U–X ). Bisbenzimide staining of the same sections is shown in I–L and Y–BB , and gene chip data for each gene are shown in M–P and CC–FF . Arrowheads in E–L and U–BB delimit the boundaries of CA1 and also the boundary between CA2 and CA3 in U–BB . Scale bars, 1 mm.

    Article Snippet: Slides were then developed and stained with the fluorescent dye bisbenzimide (Sigma) to allow localization of silver grains to individual cells.

    Techniques: Expressing, In Situ Hybridization, Staining, Chromatin Immunoprecipitation

    Knockdown of RTKN2 induces cell apoptosis ( A and B ) U2OS cells were stained with Annexin-V FITC/PI, and apoptotic cells were analyzed using flow cytometry. ( C ) Morphological changes associated with apoptosis were examined by Hoechst 33258 staining. ( D ) The apoptotic index was determined by (number of positively stained cells/total number of cells) × 100%. White arrowheads in the images indicate the nuclei of apoptotic cells (magnification, ×200). * P

    Journal: Bioscience Reports

    Article Title: Rhotekin 2 silencing inhibits proliferation and induces apoptosis in human osteosarcoma cells

    doi: 10.1042/BSR20181384

    Figure Lengend Snippet: Knockdown of RTKN2 induces cell apoptosis ( A and B ) U2OS cells were stained with Annexin-V FITC/PI, and apoptotic cells were analyzed using flow cytometry. ( C ) Morphological changes associated with apoptosis were examined by Hoechst 33258 staining. ( D ) The apoptotic index was determined by (number of positively stained cells/total number of cells) × 100%. White arrowheads in the images indicate the nuclei of apoptotic cells (magnification, ×200). * P

    Article Snippet: Then cells were stained with 10 mg/l Hoechst 33258 for 10 min at room temperature in the dark place.

    Techniques: Staining, Flow Cytometry, Cytometry

    Nuclear localization of endogenous PC4, analyzed by immunofluorescence and confocal microscopy. (A) A total of 7 × 10 4 ), followed by incubation with goat anti-rabbit FITC-conjugated antibody. Nuclei were detected by Hoechst 33258 dye (corresponding photomicrographs on the right). Bar, 30 μm. (B) Percentage of cells with endogenous PC4 staining that was cytoplasmic only (black bars), nuclear and cytoplasmic (gray bars), or nuclear only (white bars). Values are calculated for each category as the percentages of the total number of cells scored at each time point (within three fields for each experiment). Means ± the SEM are from three independent experiments. The total number of cells counted for each time point is indicated at the top of the corresponding bar. (C) Confocal microscopy of endogenous PC4. A total of 7 × 10 4 C2C7 cells were seeded onto circular coverslips placed in 35-mm dishes. Cultures were treated, and endogenous PC4 was detected as in panel A. Nuclei are visualized in red by propidium iodide (Pr.i.). The panels on the left (merge) show the overlay of the green and red staining (in orange-green), which indicates the presence of endogenous PC4 protein in the nucleus. Bar, 10 μm.

    Journal: Molecular and Cellular Biology

    Article Title: PC4 Coactivates MyoD by Relieving the Histone Deacetylase 4-Mediated Inhibition of Myocyte Enhancer Factor 2C

    doi: 10.1128/MCB.25.6.2242-2259.2005

    Figure Lengend Snippet: Nuclear localization of endogenous PC4, analyzed by immunofluorescence and confocal microscopy. (A) A total of 7 × 10 4 ), followed by incubation with goat anti-rabbit FITC-conjugated antibody. Nuclei were detected by Hoechst 33258 dye (corresponding photomicrographs on the right). Bar, 30 μm. (B) Percentage of cells with endogenous PC4 staining that was cytoplasmic only (black bars), nuclear and cytoplasmic (gray bars), or nuclear only (white bars). Values are calculated for each category as the percentages of the total number of cells scored at each time point (within three fields for each experiment). Means ± the SEM are from three independent experiments. The total number of cells counted for each time point is indicated at the top of the corresponding bar. (C) Confocal microscopy of endogenous PC4. A total of 7 × 10 4 C2C7 cells were seeded onto circular coverslips placed in 35-mm dishes. Cultures were treated, and endogenous PC4 was detected as in panel A. Nuclei are visualized in red by propidium iodide (Pr.i.). The panels on the left (merge) show the overlay of the green and red staining (in orange-green), which indicates the presence of endogenous PC4 protein in the nucleus. Bar, 10 μm.

    Article Snippet: To detect nuclei, cells were incubated at the end of the immunofluorescence staining procedure for 2 min in Hoechst 33258 dye diluted in PBS at 1 μg/ml (Sigma), washed twice in PBS, and mounted as described above.

    Techniques: Immunofluorescence, Confocal Microscopy, Incubation, Staining

    Characterization of cell death using fluorescent microscopy and Western blotting. ( A) Hoechst staining of 5o and 5m treated Mahlavu and Huh7 cells with apoptotic nuclei at 24 h and 72 h. ( B) PARP in Mahlavu and Huh7 cells treated with 5o and 5m for 72 h. Actin was used for equal loading. ( C) The bar graphs representing relative band intensities of PARP and cleaved-PARP, which were normalized with their actin loading controls.

    Journal: Journal of Enzyme Inhibition and Medicinal Chemistry

    Article Title: Synthesis and cellular bioactivities of novel isoxazole derivatives incorporating an arylpiperazine moiety as anticancer agents

    doi: 10.1080/14756366.2018.1504041

    Figure Lengend Snippet: Characterization of cell death using fluorescent microscopy and Western blotting. ( A) Hoechst staining of 5o and 5m treated Mahlavu and Huh7 cells with apoptotic nuclei at 24 h and 72 h. ( B) PARP in Mahlavu and Huh7 cells treated with 5o and 5m for 72 h. Actin was used for equal loading. ( C) The bar graphs representing relative band intensities of PARP and cleaved-PARP, which were normalized with their actin loading controls.

    Article Snippet: Then, the cells were stained with 1 µg/ml Hoechst (#33258, Sigma).

    Techniques: Microscopy, Western Blot, Staining

    Hoechst33258 fluorescent staining after SR140333 treatment (A, SR140333 treated cells; B, control) . T47D cells were cultured in DMEM contained 10%FBS and SR140333 was added at logarithmic growth phase (on day 3, at about 30% cell confluences). We carried out Hoechst33258 staining on specimens obtained by the cover slip culture method. After treated with SR140333 for 24 h, T47D cells showed slower proliferation profile and visible apoptosis was detected by Hoechst33258.

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    Article Title: SR140333 counteracts NK-1 mediated cell proliferation in human breast cancer cell line T47D

    doi: 10.1186/1756-9966-29-55

    Figure Lengend Snippet: Hoechst33258 fluorescent staining after SR140333 treatment (A, SR140333 treated cells; B, control) . T47D cells were cultured in DMEM contained 10%FBS and SR140333 was added at logarithmic growth phase (on day 3, at about 30% cell confluences). We carried out Hoechst33258 staining on specimens obtained by the cover slip culture method. After treated with SR140333 for 24 h, T47D cells showed slower proliferation profile and visible apoptosis was detected by Hoechst33258.

    Article Snippet: MTT, DMSO and Hoechst33258 were purchased from Sigma (Saint Louis, USA).

    Techniques: Staining, Cell Culture

    HSP70 colocalized with viral dsRNA in infected MACR-145 cells. MARC-145 cells were mock infected or infected with PRRSV at an MOI of 0.1. Cells were fixed at 24 h.p.i and subjected to IFA to detect dsRNA (red) and HSP70 (green) by using mouse anti-dsRNA (J2) MAb and rabbit anti-HSP70 polyclonal antibody, respectively. Nuclei were stained with Hoechst dye 33258 (blue). The images of cells were acquired with Leica TCS SP5 confocal microscope.

    Journal: BMC Microbiology

    Article Title: Inhibition of HSP70 reduces porcine reproductive and respiratory syndrome virus replication in vitro

    doi: 10.1186/1471-2180-14-64

    Figure Lengend Snippet: HSP70 colocalized with viral dsRNA in infected MACR-145 cells. MARC-145 cells were mock infected or infected with PRRSV at an MOI of 0.1. Cells were fixed at 24 h.p.i and subjected to IFA to detect dsRNA (red) and HSP70 (green) by using mouse anti-dsRNA (J2) MAb and rabbit anti-HSP70 polyclonal antibody, respectively. Nuclei were stained with Hoechst dye 33258 (blue). The images of cells were acquired with Leica TCS SP5 confocal microscope.

    Article Snippet: After three times washes, the Hoechst dye 33258 (Sigma) was added to stain the nuclei.

    Techniques: Infection, Immunofluorescence, Staining, Microscopy

    Quercetin blocked the viral protein expression. (A) MARC-145 cells were heated (HS) or not, and 8 hours later cells were inoculated with PRRSV at an MOI of 0.1 for 1 hour. Cells without heat shock treatment were subsequently cultured with medium containing DMSO, Qct (100 μM), or no chemical (Ctrl). Cells were harvested at different times as indicated for Western blotting analysis. The levels of HSP70 and viral N protein were quantified by measuring band intensities and normalized with respect to the amount of β-actin. Data are mean ± SD, n = 3. (B) PRRSV-infected MARC-145 cells were treated with DMSO or Qct at the concentration as indicated. Mock-infected cells were untreated. IFA was performed at 24 h.p.i with anti-N antibody and Alexa Fluor 555-conjugated (red) anti-mouse secondary antibody. Nuclei were stained with Hoechst dye 33258 (blue). Bar, 200 μm.

    Journal: BMC Microbiology

    Article Title: Inhibition of HSP70 reduces porcine reproductive and respiratory syndrome virus replication in vitro

    doi: 10.1186/1471-2180-14-64

    Figure Lengend Snippet: Quercetin blocked the viral protein expression. (A) MARC-145 cells were heated (HS) or not, and 8 hours later cells were inoculated with PRRSV at an MOI of 0.1 for 1 hour. Cells without heat shock treatment were subsequently cultured with medium containing DMSO, Qct (100 μM), or no chemical (Ctrl). Cells were harvested at different times as indicated for Western blotting analysis. The levels of HSP70 and viral N protein were quantified by measuring band intensities and normalized with respect to the amount of β-actin. Data are mean ± SD, n = 3. (B) PRRSV-infected MARC-145 cells were treated with DMSO or Qct at the concentration as indicated. Mock-infected cells were untreated. IFA was performed at 24 h.p.i with anti-N antibody and Alexa Fluor 555-conjugated (red) anti-mouse secondary antibody. Nuclei were stained with Hoechst dye 33258 (blue). Bar, 200 μm.

    Article Snippet: After three times washes, the Hoechst dye 33258 (Sigma) was added to stain the nuclei.

    Techniques: Expressing, Cell Culture, Western Blot, Infection, Concentration Assay, Immunofluorescence, Staining

    The knockdown of HSP70 reduced the level of viral dsRNA RI. MARC-145 cells were transfected with no siRNA (Ctrl), scramble siRNA (Scr), or different concentrations of siRNAs targetting HSP70 (siHSP70). After 24 hours, cells were mock infected or infected with PRRSV at an MOI of 0.1. Cells were fixed at 24 h.p.i and IFA was performed to detect viral dsRNA (red) with anti-dsRNA (J2) antibody. Nuclei were stained with Hoechst dye 33258 (blue). Bar, 200 μm.

    Journal: BMC Microbiology

    Article Title: Inhibition of HSP70 reduces porcine reproductive and respiratory syndrome virus replication in vitro

    doi: 10.1186/1471-2180-14-64

    Figure Lengend Snippet: The knockdown of HSP70 reduced the level of viral dsRNA RI. MARC-145 cells were transfected with no siRNA (Ctrl), scramble siRNA (Scr), or different concentrations of siRNAs targetting HSP70 (siHSP70). After 24 hours, cells were mock infected or infected with PRRSV at an MOI of 0.1. Cells were fixed at 24 h.p.i and IFA was performed to detect viral dsRNA (red) with anti-dsRNA (J2) antibody. Nuclei were stained with Hoechst dye 33258 (blue). Bar, 200 μm.

    Article Snippet: After three times washes, the Hoechst dye 33258 (Sigma) was added to stain the nuclei.

    Techniques: Transfection, Infection, Immunofluorescence, Staining

    EVO induced apoptosis in Caki-1 cells. ( a ) Apoptosis-related morphological changes were detected by staining cells with Hoechst 33258. Apoptotic cells were defined as those with blue-stained nuclei that exhibited a fragmented/condensed appearance (red arrow). Original magnification, × 200. ( b ) Elevated plasma membrane permeability was detected by staining cells in the AO/EB double-staining assay (white arrow). Original magnification, × 200. ( c ) Ultrastructural changes in Caki-1 cells were detected via TEM. a, apoptotic body; c, condensed chromatin; mv, microvillus; N, nucleus; v, vacuole. ( d ) The 3ʹ OH ends of fragmented DNA of apoptotic cells were labelled in the TUNEL assay and viewed under a fluorescence microscope. Original magnification, × 400. ( e ) Bar graph showing the quantification of TUNEL-positive Caki-1 cells. ( f ) Annexin V-FITC and PI reactivity was used to detect and quantify apoptosis induced by EVO. The percentage of cells undergoing apoptosis was determined via flow cytometry. Mean values are shown with the SD and significant differences between the EVO-treated and untreated groups are indicated as  * (P 

    Journal: Scientific Reports

    Article Title: Cytological Assessments and Transcriptome Profiling Demonstrate that Evodiamine Inhibits Growth and Induces Apoptosis in a Renal Carcinoma Cell Line

    doi: 10.1038/s41598-017-12918-y

    Figure Lengend Snippet: EVO induced apoptosis in Caki-1 cells. ( a ) Apoptosis-related morphological changes were detected by staining cells with Hoechst 33258. Apoptotic cells were defined as those with blue-stained nuclei that exhibited a fragmented/condensed appearance (red arrow). Original magnification, × 200. ( b ) Elevated plasma membrane permeability was detected by staining cells in the AO/EB double-staining assay (white arrow). Original magnification, × 200. ( c ) Ultrastructural changes in Caki-1 cells were detected via TEM. a, apoptotic body; c, condensed chromatin; mv, microvillus; N, nucleus; v, vacuole. ( d ) The 3ʹ OH ends of fragmented DNA of apoptotic cells were labelled in the TUNEL assay and viewed under a fluorescence microscope. Original magnification, × 400. ( e ) Bar graph showing the quantification of TUNEL-positive Caki-1 cells. ( f ) Annexin V-FITC and PI reactivity was used to detect and quantify apoptosis induced by EVO. The percentage of cells undergoing apoptosis was determined via flow cytometry. Mean values are shown with the SD and significant differences between the EVO-treated and untreated groups are indicated as * (P 

    Article Snippet: Subsequently, the cells were stained with 4 ng/ml 4′,6-diamidino-2-phenylindole (DAPI) or 1 μg/ml Hoechst 33258 solution (Sigma–Aldrich) at 4 °C for 5–10 min. Stained cells in each group were observed using a confocal laser scanning microscope (Leica Microsystems, Hessen Wetzlar, Germany).

    Techniques: Staining, Permeability, Double Staining, Transmission Electron Microscopy, TUNEL Assay, Fluorescence, Microscopy, Flow Cytometry, Cytometry

    Hoechst 33,258 staining in U87MG cells. ( A ) Negative control; ( B ) Cells treated with compound  10  for 48 h at IC 50  concentration (arrow shows nuclear fragmentation, magnification ×400).

    Journal: Molecules : A Journal of Synthetic Chemistry and Natural Product Chemistry

    Article Title: Piplartine Analogues and Cytotoxic Evaluation against Glioblastoma

    doi: 10.3390/molecules23061382

    Figure Lengend Snippet: Hoechst 33,258 staining in U87MG cells. ( A ) Negative control; ( B ) Cells treated with compound 10 for 48 h at IC 50 concentration (arrow shows nuclear fragmentation, magnification ×400).

    Article Snippet: The cells were then stained with 1 μL Hoechst 33,258 (5 μg/mL; Sigma-Aldrich, Saint Louis, MO, USA) and incubated at room temperature in the dark for 5 min. Changes in nuclear morphology were visualized under a fluorescent microscope (excitation, 350 nm; emission, 460 nm; Leica® DM IL LED, Wetzlar, Germany).

    Techniques: Staining, Negative Control, Concentration Assay

    Cellular apoptosis in MKN-28 cells was analyzed by Hoechst 33258 staining and flow cytometry. (A) Apoptotic features were identified by observing chromatin condensation and fragmentation after Hoechst 33258 staining, as indicated by white arrows. (B) Detection of apoptosis via annexin V/PI staining (X-axis, annexin V; Y-axis, PI). The early apoptotic cells were defined as the sum of cells in the lower right quadrant of panel B. In A and B: (a) control; (b) 40μM SIN; (c) 100 mg/l 5-FU; and (d) 20 μM SIN + 50 mg/l 5-FU. (C) The apoptotic rate, as assessed via Hoechst 33258 staining. Bars indicate the mean ± SD (n=3). SIN, sinomenine; 5-FU, 5-fluorouracil; PI, propidium iodide.

    Journal: Oncology Letters

    Article Title: Sinomenine sensitizes gastric cancer cells to 5-fluorouracil in vitro and in vivo

    doi: 10.3892/ol.2013.1592

    Figure Lengend Snippet: Cellular apoptosis in MKN-28 cells was analyzed by Hoechst 33258 staining and flow cytometry. (A) Apoptotic features were identified by observing chromatin condensation and fragmentation after Hoechst 33258 staining, as indicated by white arrows. (B) Detection of apoptosis via annexin V/PI staining (X-axis, annexin V; Y-axis, PI). The early apoptotic cells were defined as the sum of cells in the lower right quadrant of panel B. In A and B: (a) control; (b) 40μM SIN; (c) 100 mg/l 5-FU; and (d) 20 μM SIN + 50 mg/l 5-FU. (C) The apoptotic rate, as assessed via Hoechst 33258 staining. Bars indicate the mean ± SD (n=3). SIN, sinomenine; 5-FU, 5-fluorouracil; PI, propidium iodide.

    Article Snippet: Detection of apoptotic cells by Hoechst 33258 staining and flow cytometry The morphological features of apoptotic cells (chromatin condensation and fragmentation) were detected by Hoechst 33258 staining as follows: MKN-28 cells were treated with 100 mg/l 5-FU, 40 μM SIN or 50mg/l 5-FU + 20 μM SIN for 24 h, followed by incubation with 20 μM Hoechst 33258 (Sigma-Aldrich) for 10 min at room temperature.

    Techniques: Staining, Flow Cytometry, Cytometry

    Apoptosis induction studies of PPI. ( A ) AO\EB staining. ( B ) Hoechst 33258 staining. ( C ) TUNEL assay.

    Journal: Chemistry Central Journal

    Article Title: Discovery and antitumor activities of constituents from Cyrtomium fortumei (J.) Smith rhizomes

    doi: 10.1186/1752-153X-7-24

    Figure Lengend Snippet: Apoptosis induction studies of PPI. ( A ) AO\EB staining. ( B ) Hoechst 33258 staining. ( C ) TUNEL assay.

    Article Snippet: The cells were washed twice with PBS, and were consequently stained with 0.5 mL of Hoechst 33258 staining (Beyotime Co., Jiangsu, China) for 5 min.

    Techniques: Staining, TUNEL Assay

    Long-term culture hiPS cells by automated system. ( A ) Fold change of cell number determined by split ratio. Black circles indicate each passaging point. ( B ) Fluorescent micrograph of hiPS cells automatically passaged 20 times. Cells were immunologically stained with antibodies against OCT 3/4, NANOG, SOX2, SSEA-4, TRA-1-60, and TRA-1-81. Cell nuclei were stained with Hoechst 33258. Scale bar: 200 μm. ( C ) qPCR analysis of gene expression of OCT 3/4 and NANOG. Expression levels were normalized to expression levels of unpassaged cells (mean ± sd., n = 3). P0, unpassaged cells; P20, cells automatically passaged for 20 times; EB, embryoid body. N.S., not significant. ( D ) Representative histograms of FACS analysis. ( E ) FACS quantification of cells throughout culture. ( F ) Global gene expression analysis. Cluster analysis was performed with all detected genes ( G ) Karyotype analysis of hiPS cells automatically passaged 20 times.

    Journal: Scientific Reports

    Article Title: Long-term maintenance of human induced pluripotent stem cells by automated cell culture system

    doi: 10.1038/srep16647

    Figure Lengend Snippet: Long-term culture hiPS cells by automated system. ( A ) Fold change of cell number determined by split ratio. Black circles indicate each passaging point. ( B ) Fluorescent micrograph of hiPS cells automatically passaged 20 times. Cells were immunologically stained with antibodies against OCT 3/4, NANOG, SOX2, SSEA-4, TRA-1-60, and TRA-1-81. Cell nuclei were stained with Hoechst 33258. Scale bar: 200 μm. ( C ) qPCR analysis of gene expression of OCT 3/4 and NANOG. Expression levels were normalized to expression levels of unpassaged cells (mean ± sd., n = 3). P0, unpassaged cells; P20, cells automatically passaged for 20 times; EB, embryoid body. N.S., not significant. ( D ) Representative histograms of FACS analysis. ( E ) FACS quantification of cells throughout culture. ( F ) Global gene expression analysis. Cluster analysis was performed with all detected genes ( G ) Karyotype analysis of hiPS cells automatically passaged 20 times.

    Article Snippet: The cell nuclei were counterstained with 1 μg/mL Hoechst 33258 (Dojindo Laboratories, Kumamoto, Japan).

    Techniques: Passaging, Staining, Real-time Polymerase Chain Reaction, Expressing, FACS

    Time course of 8-nitroG formation in In 2 O 3 -treated cells. (A) Immunofluorescent images of 8-nitroG formation in In 2 O 3 -treated cells. RAW 264.7 cells were treated with 20 µg/m l  of In 2 O 3  for indicated durations at 37°C. 8-NitroG formation was detected by immunocytochemistry as described in Materials and Methods section. The nucleus was stained with Hoechst 33258. Magnification, ×200. (B) Quantitative image analysis of 8-nitroG formation in In 2 O 3 -treated cells. The staining intensity per cell was quantified with an ImageJ software. The vertical axis shows the fold increase in the staining intensity compared with that of the control at 2 h, which was set at 1. The data were expressed as means ± SD of 3-4 independent experiments. * p

    Journal: Journal of Occupational Health

    Article Title: Nitrative DNA damage in cultured macrophages exposed to indium oxide

    doi: 10.1539/joh.17-0146-OA

    Figure Lengend Snippet: Time course of 8-nitroG formation in In 2 O 3 -treated cells. (A) Immunofluorescent images of 8-nitroG formation in In 2 O 3 -treated cells. RAW 264.7 cells were treated with 20 µg/m l of In 2 O 3 for indicated durations at 37°C. 8-NitroG formation was detected by immunocytochemistry as described in Materials and Methods section. The nucleus was stained with Hoechst 33258. Magnification, ×200. (B) Quantitative image analysis of 8-nitroG formation in In 2 O 3 -treated cells. The staining intensity per cell was quantified with an ImageJ software. The vertical axis shows the fold increase in the staining intensity compared with that of the control at 2 h, which was set at 1. The data were expressed as means ± SD of 3-4 independent experiments. * p

    Article Snippet: Then the cells were incubated with fluorescent secondary antibody [Alexa 594-labeled goat antibody against rabbit IgG (1:400, molecular probes, Eugene, OR, USA) ] for 3 h. The nuclei were stained with 5 μM Hoechst 33258 (Polysciences Inc., Warrington, PA, USA).

    Techniques: Immunocytochemistry, Staining, Software

    Effects of iNOS and endocytosis inhibitors on In 2 O 3 -induced DNA damage and their cytotoxicity. (A) Effects of iNOS and endocytosis inhibitors on In 2 O 3 -induced 8-nitroG formation. RAW 264.7 cells were treated with 20 µg/m l  of In 2 O 3  for 4 h at 37°C. The cells were co-treated with 1 µM 1400W, 0.5 mM MBCD or 50 µM MDC. 8-NitroG formation was detected by immunocytochemistry as described in Materials and Methods section. The nucleus was stained with Hoechst 33258. Magnification, ×200. (B) Cytotoxicity of In 2 O 3  plus inhibitors for iNOS and endocytosis. RAW 264.7 cells were treated with 20 µg/m l  In 2 O 3  plus 1 µM 1400W, 0.5 mM MBCD or 50 µM MDC for 4 h at 37°C. Cell viability was evaluated by trypan blue exclusion assay. The data were expressed as means ± SD of 4 independent experiments.

    Journal: Journal of Occupational Health

    Article Title: Nitrative DNA damage in cultured macrophages exposed to indium oxide

    doi: 10.1539/joh.17-0146-OA

    Figure Lengend Snippet: Effects of iNOS and endocytosis inhibitors on In 2 O 3 -induced DNA damage and their cytotoxicity. (A) Effects of iNOS and endocytosis inhibitors on In 2 O 3 -induced 8-nitroG formation. RAW 264.7 cells were treated with 20 µg/m l of In 2 O 3 for 4 h at 37°C. The cells were co-treated with 1 µM 1400W, 0.5 mM MBCD or 50 µM MDC. 8-NitroG formation was detected by immunocytochemistry as described in Materials and Methods section. The nucleus was stained with Hoechst 33258. Magnification, ×200. (B) Cytotoxicity of In 2 O 3 plus inhibitors for iNOS and endocytosis. RAW 264.7 cells were treated with 20 µg/m l In 2 O 3 plus 1 µM 1400W, 0.5 mM MBCD or 50 µM MDC for 4 h at 37°C. Cell viability was evaluated by trypan blue exclusion assay. The data were expressed as means ± SD of 4 independent experiments.

    Article Snippet: Then the cells were incubated with fluorescent secondary antibody [Alexa 594-labeled goat antibody against rabbit IgG (1:400, molecular probes, Eugene, OR, USA) ] for 3 h. The nuclei were stained with 5 μM Hoechst 33258 (Polysciences Inc., Warrington, PA, USA).

    Techniques: Immunocytochemistry, Staining, Trypan Blue Exclusion Assay

    8-NitroG formation in In 2 O 3 -treated cells. (A) Immunofluorescent images of 8-nitroG formation in In 2 O 3 -treated cells. RAW 264.7 cells were treated with indicated concentrations of In 2 O 3  for 4 h at 37°C. 8-NitroG formation was detected by immunocytochemistry as described in Materials and Methods section. The nucleus was stained with Hoechst 33258. Magnification, ×200. (B) Quantitative image analysis of 8-nitroG formation in In 2 O 3 -treated cells. The staining intensity per cell was quantified with an ImageJ software. The vertical axis shows the fold increase in the staining intensity compared with that of the control, which was set at 1. The data were expressed as means ± SD of 3-4 independent experiments. * p

    Journal: Journal of Occupational Health

    Article Title: Nitrative DNA damage in cultured macrophages exposed to indium oxide

    doi: 10.1539/joh.17-0146-OA

    Figure Lengend Snippet: 8-NitroG formation in In 2 O 3 -treated cells. (A) Immunofluorescent images of 8-nitroG formation in In 2 O 3 -treated cells. RAW 264.7 cells were treated with indicated concentrations of In 2 O 3 for 4 h at 37°C. 8-NitroG formation was detected by immunocytochemistry as described in Materials and Methods section. The nucleus was stained with Hoechst 33258. Magnification, ×200. (B) Quantitative image analysis of 8-nitroG formation in In 2 O 3 -treated cells. The staining intensity per cell was quantified with an ImageJ software. The vertical axis shows the fold increase in the staining intensity compared with that of the control, which was set at 1. The data were expressed as means ± SD of 3-4 independent experiments. * p

    Article Snippet: Then the cells were incubated with fluorescent secondary antibody [Alexa 594-labeled goat antibody against rabbit IgG (1:400, molecular probes, Eugene, OR, USA) ] for 3 h. The nuclei were stained with 5 μM Hoechst 33258 (Polysciences Inc., Warrington, PA, USA).

    Techniques: Immunocytochemistry, Staining, Software

    Hoechst 33258 staining in KIAA0100-knockdown U937 cells. Cells were stained with Hoechst 33258 detected and the apoptotic cells were calculated by fluorescent photomicrographs at 200×. The results showed the apoptotic cells were the most in KIAA0100-knockdown U937 cells. The experiments was performed in triplicate and data were displayed as means±SE. **P

    Journal: Journal of Pharmaceutical Analysis

    Article Title: Bioinformatic prediction and functional characterization of human KIAA0100 gene

    doi: 10.1016/j.jpha.2016.09.003

    Figure Lengend Snippet: Hoechst 33258 staining in KIAA0100-knockdown U937 cells. Cells were stained with Hoechst 33258 detected and the apoptotic cells were calculated by fluorescent photomicrographs at 200×. The results showed the apoptotic cells were the most in KIAA0100-knockdown U937 cells. The experiments was performed in triplicate and data were displayed as means±SE. **P

    Article Snippet: Morphological study The log-phase Cas9-U937 cells were infected by the most effective lentivirus-sgRNA-EGFPMCS , at 120 h after infection, cells were collected and washed twice in 4 °C PBS, and stained with Hoechst 33258 staining solution according to the manufacturer's instructions (Beyotime, China).

    Techniques: Staining

    BSO and NAC influence the α-hederin-induced apoptosis of SMMC-7721 cells. SMMC-7721 cells were incubated with α-hederin (10 μmol/L) with or without BSO (2 mmol/L) or NAC (5 mmol/L) pretreatment. A: Cell apoptosis was determined by Hoechst 33258 staining; B: ROS levels in SMMC-7721 cells; C and D: Effect of α-hederin on intracellular GSH and ATP levels. a P

    Journal: World Journal of Gastroenterology

    Article Title: Mitochondrial pathway mediated by reactive oxygen species involvement in α-hederin-induced apoptosis in hepatocellular carcinoma cells

    doi: 10.3748/wjg.v24.i17.1901

    Figure Lengend Snippet: BSO and NAC influence the α-hederin-induced apoptosis of SMMC-7721 cells. SMMC-7721 cells were incubated with α-hederin (10 μmol/L) with or without BSO (2 mmol/L) or NAC (5 mmol/L) pretreatment. A: Cell apoptosis was determined by Hoechst 33258 staining; B: ROS levels in SMMC-7721 cells; C and D: Effect of α-hederin on intracellular GSH and ATP levels. a P

    Article Snippet: Cell apoptosis assays Apoptotic cells were examined using the Hoechst 33258 staining kit (Beyotime).

    Techniques: Incubation, Staining

    α-hederin reduces hepatocellular carcinoma cell viability and induces the apoptosis of hepatocellular carcinoma cells through GSH depletion and reactive oxygen species accumulation. A: Cell Counting Kit-8 assays showed that α-hederin inhibits the viability of hepatocellular carcinoma cells (SMMC-7721, HepG-2, and Huh-7) in a dose- and time-dependent manner; B: SMMC-7721 cells were incubated with α-hederin (0, 5 μmol/L, 10 μmol/L, or 20 μmol/L) and stained with Hoechst 33258. Apoptotic cells were identified by fragmented and condensed nuclei under a fluorescence microscope. The percentage of apoptotic cells was calculated,  P  for trend

    Journal: World Journal of Gastroenterology

    Article Title: Mitochondrial pathway mediated by reactive oxygen species involvement in α-hederin-induced apoptosis in hepatocellular carcinoma cells

    doi: 10.3748/wjg.v24.i17.1901

    Figure Lengend Snippet: α-hederin reduces hepatocellular carcinoma cell viability and induces the apoptosis of hepatocellular carcinoma cells through GSH depletion and reactive oxygen species accumulation. A: Cell Counting Kit-8 assays showed that α-hederin inhibits the viability of hepatocellular carcinoma cells (SMMC-7721, HepG-2, and Huh-7) in a dose- and time-dependent manner; B: SMMC-7721 cells were incubated with α-hederin (0, 5 μmol/L, 10 μmol/L, or 20 μmol/L) and stained with Hoechst 33258. Apoptotic cells were identified by fragmented and condensed nuclei under a fluorescence microscope. The percentage of apoptotic cells was calculated, P for trend

    Article Snippet: Cell apoptosis assays Apoptotic cells were examined using the Hoechst 33258 staining kit (Beyotime).

    Techniques: Cell Counting, Incubation, Staining, Fluorescence, Microscopy

    A correlation between IGF-1R activation at focal adhesion-like structures and EMT transcriptomic signatures in erlotinib-refractory PC-9 cells. Left. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and vinculin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-vinculin antibodies, as specified, and Hoechst 33258 for nuclear counterstaining. Images show representative portions of erlotinib-sensitive PC-9 parental cells and erlotinib-refractory PC-9 POOLs captured in different channels for PP-IGF-1R Tyr1161 ( green ), vinculin ( red ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software. Right. EMT expression profiling of erlotinib-refractory PC-9 pooled cells versus erlotinib-sensitive PC-9 parental cells. Figure shows a representative scatter plot of the difference in the relative abundance of 19 mRNAs whose levels, in published results, are significantly altered during activation/deactivation of the EMT genetic program. Green and red symbols (≥ 2-fold) denote downregulation and upregulation versus basal expression in PC-9 parental cells, respectively; grey symbols denote fold-change results that need to be validated with a sufficient number of biological replicates (i.e., fold-change results may have greater variation of p value > 0.05 or the p value for the fold-change is either unavailable or relatively high [p > 0.05] or, alternatively, they are uninterpretable because the gene's average threshold cycles were either not determined or greater than the defined cut-off value [default 35] in both samples).

    Journal: Scientific Reports

    Article Title: IGF-1R/epithelial-to-mesenchymal transition (EMT) crosstalk suppresses the erlotinib-sensitizing effect of EGFR exon 19 deletion mutations

    doi: 10.1038/srep02560

    Figure Lengend Snippet: A correlation between IGF-1R activation at focal adhesion-like structures and EMT transcriptomic signatures in erlotinib-refractory PC-9 cells. Left. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and vinculin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-vinculin antibodies, as specified, and Hoechst 33258 for nuclear counterstaining. Images show representative portions of erlotinib-sensitive PC-9 parental cells and erlotinib-refractory PC-9 POOLs captured in different channels for PP-IGF-1R Tyr1161 ( green ), vinculin ( red ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software. Right. EMT expression profiling of erlotinib-refractory PC-9 pooled cells versus erlotinib-sensitive PC-9 parental cells. Figure shows a representative scatter plot of the difference in the relative abundance of 19 mRNAs whose levels, in published results, are significantly altered during activation/deactivation of the EMT genetic program. Green and red symbols (≥ 2-fold) denote downregulation and upregulation versus basal expression in PC-9 parental cells, respectively; grey symbols denote fold-change results that need to be validated with a sufficient number of biological replicates (i.e., fold-change results may have greater variation of p value > 0.05 or the p value for the fold-change is either unavailable or relatively high [p > 0.05] or, alternatively, they are uninterpretable because the gene's average threshold cycles were either not determined or greater than the defined cut-off value [default 35] in both samples).

    Article Snippet: The images were captured in different channels for Alexa Fluor® 488 (pseudocolored green), Alexa Fluor® 594 (pseudocolored red), and Hoechst 33258 (pseudocolored blue) using a BD Pathway™ 855 Bioimager System (Becton Dickinson Biosciences, San Jose, California) with 20 × or 40 × objectives (NA 075 Olympus).

    Techniques: Activation Assay, Staining, Software, Expressing

    Effects of erlotinib treatment on the epithelial/mesenchymal phenotype of NSCLC cells. A431 (positive control), PC-9, H460, H1975, and H1993 cells were grown until 75%–80% confluence, serum-starved for 24 h, and then treated for 72 h with graded concentrations (1 and 10 μmol/L) of erlotinib. Left panels. After fixation and permeabilization, sub-cellular distribution of the epithelial marker E-cadherin was assessed following staining with an anti-E-cadherin antibody and Hoechst 33258 for nuclear counterstaining. Middle panels . After fixation and permeabilization, sub-cellular distribution of F-actin was assessed following staining with an anti-F-actin antibody and Hoechst 33258 for nuclear counterstaining. Right panels. After fixation and permeabilization, sub-cellular distribution of the mesenchymal-specific marker vimentin was assessed following staining with an anti-vimentin antibody and Hoechst 33258 for nuclear counterstaining. Images show representative 4 × 4 montages of untreated- or erlotinib-treated cell cultures captured in different channels for E-cadherin ( green ), F-actin ( red ), vimentin ( green ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Journal: Scientific Reports

    Article Title: IGF-1R/epithelial-to-mesenchymal transition (EMT) crosstalk suppresses the erlotinib-sensitizing effect of EGFR exon 19 deletion mutations

    doi: 10.1038/srep02560

    Figure Lengend Snippet: Effects of erlotinib treatment on the epithelial/mesenchymal phenotype of NSCLC cells. A431 (positive control), PC-9, H460, H1975, and H1993 cells were grown until 75%–80% confluence, serum-starved for 24 h, and then treated for 72 h with graded concentrations (1 and 10 μmol/L) of erlotinib. Left panels. After fixation and permeabilization, sub-cellular distribution of the epithelial marker E-cadherin was assessed following staining with an anti-E-cadherin antibody and Hoechst 33258 for nuclear counterstaining. Middle panels . After fixation and permeabilization, sub-cellular distribution of F-actin was assessed following staining with an anti-F-actin antibody and Hoechst 33258 for nuclear counterstaining. Right panels. After fixation and permeabilization, sub-cellular distribution of the mesenchymal-specific marker vimentin was assessed following staining with an anti-vimentin antibody and Hoechst 33258 for nuclear counterstaining. Images show representative 4 × 4 montages of untreated- or erlotinib-treated cell cultures captured in different channels for E-cadherin ( green ), F-actin ( red ), vimentin ( green ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Article Snippet: The images were captured in different channels for Alexa Fluor® 488 (pseudocolored green), Alexa Fluor® 594 (pseudocolored red), and Hoechst 33258 (pseudocolored blue) using a BD Pathway™ 855 Bioimager System (Becton Dickinson Biosciences, San Jose, California) with 20 × or 40 × objectives (NA 075 Olympus).

    Techniques: Positive Control, Marker, Staining, Software

    Cytoskeleton organization and distribution of the active form of IGF-1R in erlotinib-refractory PC-9 cells. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and F-actin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-F-actin antibodies, as specified, and Hoechst 33258 for nuclear counterstaining. Images show representative portions of erlotinib-sensitive PC-9 parental cells and erlotinib-refractory PC-9 POOLs captured in different channels for PP-IGF-1R Tyr1161 ( green ), F-actin ( red ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Journal: Scientific Reports

    Article Title: IGF-1R/epithelial-to-mesenchymal transition (EMT) crosstalk suppresses the erlotinib-sensitizing effect of EGFR exon 19 deletion mutations

    doi: 10.1038/srep02560

    Figure Lengend Snippet: Cytoskeleton organization and distribution of the active form of IGF-1R in erlotinib-refractory PC-9 cells. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and F-actin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-F-actin antibodies, as specified, and Hoechst 33258 for nuclear counterstaining. Images show representative portions of erlotinib-sensitive PC-9 parental cells and erlotinib-refractory PC-9 POOLs captured in different channels for PP-IGF-1R Tyr1161 ( green ), F-actin ( red ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Article Snippet: The images were captured in different channels for Alexa Fluor® 488 (pseudocolored green), Alexa Fluor® 594 (pseudocolored red), and Hoechst 33258 (pseudocolored blue) using a BD Pathway™ 855 Bioimager System (Becton Dickinson Biosciences, San Jose, California) with 20 × or 40 × objectives (NA 075 Olympus).

    Techniques: Staining, Software

    TGFβ1-induced EMT is sufficient to promote refractoriness to erlotinib in PC-9 cells. Left panels. The effects of sequential ( top ) or concurrent ( bottom ) treatment with the EMT inducer TGFβ1 on cell viability of PC-9 cells exposed to erlotinib were measured by MTT uptake assays. Figure shows dose-response graphs as% of untreated cell populations (untreated control cells = 100% cell viability). Results are means ( columns ) and 95% confidence intervals ( bars ) of three independent experiments made in triplicate. Statistically significant differences (one-factor ANOVA analysis) between MTT uptakes in treated and untreated control cells are shown. Statistical tests were two-sided. Right panels. Images show representative portions (n = 3) of control- or TGFβ1-treated PC-9 cell cultures at the start of erlotinib treatment ( top ) or once the treatment with erlotinib is finished ( bottom ) and captured in different channels for F-actin ( red ), vimentin (green), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Journal: Scientific Reports

    Article Title: IGF-1R/epithelial-to-mesenchymal transition (EMT) crosstalk suppresses the erlotinib-sensitizing effect of EGFR exon 19 deletion mutations

    doi: 10.1038/srep02560

    Figure Lengend Snippet: TGFβ1-induced EMT is sufficient to promote refractoriness to erlotinib in PC-9 cells. Left panels. The effects of sequential ( top ) or concurrent ( bottom ) treatment with the EMT inducer TGFβ1 on cell viability of PC-9 cells exposed to erlotinib were measured by MTT uptake assays. Figure shows dose-response graphs as% of untreated cell populations (untreated control cells = 100% cell viability). Results are means ( columns ) and 95% confidence intervals ( bars ) of three independent experiments made in triplicate. Statistically significant differences (one-factor ANOVA analysis) between MTT uptakes in treated and untreated control cells are shown. Statistical tests were two-sided. Right panels. Images show representative portions (n = 3) of control- or TGFβ1-treated PC-9 cell cultures at the start of erlotinib treatment ( top ) or once the treatment with erlotinib is finished ( bottom ) and captured in different channels for F-actin ( red ), vimentin (green), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Article Snippet: The images were captured in different channels for Alexa Fluor® 488 (pseudocolored green), Alexa Fluor® 594 (pseudocolored red), and Hoechst 33258 (pseudocolored blue) using a BD Pathway™ 855 Bioimager System (Becton Dickinson Biosciences, San Jose, California) with 20 × or 40 × objectives (NA 075 Olympus).

    Techniques: MTT Assay, Software

    TGFβ1 induces a bona fide EMT in erlotinib-sensitive PC-9 cells. (A). PC-9 cells were grown until 75%–80% confluence, serum-starved for 24 h, and then treated for 3 days with 10 ng/mL TGFβ1. Phase-contrast ( left ) and immunofluorescence ( right ) images were obtained and merged on BD Pathway 855 Bioimager System according the recommended assay procedure (BD Attovision software). Images show representative portions (n = 3) of untreated- or TGFβ1-treated PC-9 cells captured in different channels for F-actin ( red ), vimentin (green), and Hoechst 33258 ( blue ) with a 20 × objective. (B). Left. EMT expression profiling of PC-9/TGFβ1 cells versus untreated PC-9 cells. Figure shows a representative scatter plot of the difference in the relative abundance of 19 mRNAs whose levels, in published results, are significantly altered during activation/deactivation of the EMT genetic program (see legend in Fig. 3 ). Right. After fixation and permeabilization, sub-cellular distribution of the epithelial marker E-cadherin was assessed following staining with an anti-E-cadherin antibody and Hoechst 33258 for nuclear counterstaining. Images show representative portions of untreated- and TGFβ1-treated PC-9 cells captured in different channels for E-cadherin ( green ) and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Journal: Scientific Reports

    Article Title: IGF-1R/epithelial-to-mesenchymal transition (EMT) crosstalk suppresses the erlotinib-sensitizing effect of EGFR exon 19 deletion mutations

    doi: 10.1038/srep02560

    Figure Lengend Snippet: TGFβ1 induces a bona fide EMT in erlotinib-sensitive PC-9 cells. (A). PC-9 cells were grown until 75%–80% confluence, serum-starved for 24 h, and then treated for 3 days with 10 ng/mL TGFβ1. Phase-contrast ( left ) and immunofluorescence ( right ) images were obtained and merged on BD Pathway 855 Bioimager System according the recommended assay procedure (BD Attovision software). Images show representative portions (n = 3) of untreated- or TGFβ1-treated PC-9 cells captured in different channels for F-actin ( red ), vimentin (green), and Hoechst 33258 ( blue ) with a 20 × objective. (B). Left. EMT expression profiling of PC-9/TGFβ1 cells versus untreated PC-9 cells. Figure shows a representative scatter plot of the difference in the relative abundance of 19 mRNAs whose levels, in published results, are significantly altered during activation/deactivation of the EMT genetic program (see legend in Fig. 3 ). Right. After fixation and permeabilization, sub-cellular distribution of the epithelial marker E-cadherin was assessed following staining with an anti-E-cadherin antibody and Hoechst 33258 for nuclear counterstaining. Images show representative portions of untreated- and TGFβ1-treated PC-9 cells captured in different channels for E-cadherin ( green ) and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Article Snippet: The images were captured in different channels for Alexa Fluor® 488 (pseudocolored green), Alexa Fluor® 594 (pseudocolored red), and Hoechst 33258 (pseudocolored blue) using a BD Pathway™ 855 Bioimager System (Becton Dickinson Biosciences, San Jose, California) with 20 × or 40 × objectives (NA 075 Olympus).

    Techniques: Immunofluorescence, Software, Expressing, Activation Assay, Marker, Staining

    Pharmacological inhibition of IGF-1R activity impedes TGFβ1-induced EMT in PC-9 cells. PC-9 cells were grown until 75%–80% confluence, serum-starved for 24 h, and then treated for 3 days with 10 ng/mL TGFβ1 in the absence of presence of 1 μmol/L AG1024. Top panels. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and F-actin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-F-actin antibodies, as specified, and Hoechst 33258 for nuclear counterstaining. Middle panels . After fixation and permeabilization, sub-cellular distribution of the epithelial marker E-cadherin was assessed following staining with an anti-E-cadherin antibody and Hoechst 33258 for nuclear counterstaining. Bottom panels. After fixation and permeabilization, sub-cellular distribution of the mesenchymal-specific marker vimentin was assessed following staining with an anti-vimentin antibody and Hoechst 33258 for nuclear counterstaining. Images show representative portions of untreated-, TGFβ1-, AG1024-, or TGFβ1 + AG1024-treated PC-9 cells captured in different channels for PP-IGF-1R Tyr1161 ( green ), F-actin ( red ), E-cadherin ( green ), vimentin ( green ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Journal: Scientific Reports

    Article Title: IGF-1R/epithelial-to-mesenchymal transition (EMT) crosstalk suppresses the erlotinib-sensitizing effect of EGFR exon 19 deletion mutations

    doi: 10.1038/srep02560

    Figure Lengend Snippet: Pharmacological inhibition of IGF-1R activity impedes TGFβ1-induced EMT in PC-9 cells. PC-9 cells were grown until 75%–80% confluence, serum-starved for 24 h, and then treated for 3 days with 10 ng/mL TGFβ1 in the absence of presence of 1 μmol/L AG1024. Top panels. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and F-actin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-F-actin antibodies, as specified, and Hoechst 33258 for nuclear counterstaining. Middle panels . After fixation and permeabilization, sub-cellular distribution of the epithelial marker E-cadherin was assessed following staining with an anti-E-cadherin antibody and Hoechst 33258 for nuclear counterstaining. Bottom panels. After fixation and permeabilization, sub-cellular distribution of the mesenchymal-specific marker vimentin was assessed following staining with an anti-vimentin antibody and Hoechst 33258 for nuclear counterstaining. Images show representative portions of untreated-, TGFβ1-, AG1024-, or TGFβ1 + AG1024-treated PC-9 cells captured in different channels for PP-IGF-1R Tyr1161 ( green ), F-actin ( red ), E-cadherin ( green ), vimentin ( green ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Article Snippet: The images were captured in different channels for Alexa Fluor® 488 (pseudocolored green), Alexa Fluor® 594 (pseudocolored red), and Hoechst 33258 (pseudocolored blue) using a BD Pathway™ 855 Bioimager System (Becton Dickinson Biosciences, San Jose, California) with 20 × or 40 × objectives (NA 075 Olympus).

    Techniques: Inhibition, Activity Assay, Staining, Marker, Software

    Short-term exposure to erlotinib activates EMT-like features in PC-9 cells. PC-9 cells were grown until 75%–80% confluence, serum-starved for 24 h, and then treated for 72 h with 1 μmol/L erlotinib. Top panels. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and F-actin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-F-actin antibodies, as specified, and Hoechst 33258 for nuclear counterstaining. Middle panels . After fixation and permeabilization, sub-cellular distribution of the epithelial marker E-cadherin was assessed following staining with an anti-E-cadherin antibody and Hoechst 33258 for nuclear counterstaining. Bottom panels. After fixation and permeabilization, sub-cellular distribution of the mesenchymal-specific marker vimentin was assessed following staining with an anti-vimentin antibody and Hoechst 33258 for nuclear counterstaining. Images show representative portions (left panels) or 4 × 4 montages (right panels) of untreated- or erlotinib-treated PC-9 cell cultures captured in different channels for PP-IGF-1R Tyr1161 ( green ), F-actin ( red ), E-cadherin ( green ), vimentin ( green ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Journal: Scientific Reports

    Article Title: IGF-1R/epithelial-to-mesenchymal transition (EMT) crosstalk suppresses the erlotinib-sensitizing effect of EGFR exon 19 deletion mutations

    doi: 10.1038/srep02560

    Figure Lengend Snippet: Short-term exposure to erlotinib activates EMT-like features in PC-9 cells. PC-9 cells were grown until 75%–80% confluence, serum-starved for 24 h, and then treated for 72 h with 1 μmol/L erlotinib. Top panels. After fixation and permeabilization, cellular co-distribution of PP-IGF-1R Tyr1161 and F-actin was assessed following staining with anti-PP-IGF-1R Tyr1161 and anti-F-actin antibodies, as specified, and Hoechst 33258 for nuclear counterstaining. Middle panels . After fixation and permeabilization, sub-cellular distribution of the epithelial marker E-cadherin was assessed following staining with an anti-E-cadherin antibody and Hoechst 33258 for nuclear counterstaining. Bottom panels. After fixation and permeabilization, sub-cellular distribution of the mesenchymal-specific marker vimentin was assessed following staining with an anti-vimentin antibody and Hoechst 33258 for nuclear counterstaining. Images show representative portions (left panels) or 4 × 4 montages (right panels) of untreated- or erlotinib-treated PC-9 cell cultures captured in different channels for PP-IGF-1R Tyr1161 ( green ), F-actin ( red ), E-cadherin ( green ), vimentin ( green ), and Hoechst 33258 ( blue ) with a 20 × objective, and merged on BD Pathway 855 Bioimager System using BD Attovision software.

    Article Snippet: The images were captured in different channels for Alexa Fluor® 488 (pseudocolored green), Alexa Fluor® 594 (pseudocolored red), and Hoechst 33258 (pseudocolored blue) using a BD Pathway™ 855 Bioimager System (Becton Dickinson Biosciences, San Jose, California) with 20 × or 40 × objectives (NA 075 Olympus).

    Techniques: Staining, Marker, Software

    Icaritin induces K562 cells or primary cells apoptosis. A. Morphological features for apoptosis in untreated and Icaritin-treated K562 cells were revealed by Hoechst 33258 staining. Condensed chromatin and apoptotic body could be found in Icaritin-treated K562 cells by fluorescence microscope (Olympus, B×50); ×400. B. Percentages of apoptotic cells in various concentration of Icaritin-treated K562 cells (Hoechst 33258 staining). The results represent mean±SD of triplicate experiments. C. Flow-cytometry analysis showed externalized PS, revealed by Annexin staining, increased significantly in 32 µM Icaritin-treated K562 cells. D. Percentages of apoptotic cells in Icaritin-treated fresh primary cells from CML-BP patients bone marrow (n = 5) based on annexin V expression assays. Error bars represent SD of experiments. E. Cell cycle analysis showing sub-G1 content in Icaritin (32 µM)-treated cells (right panel) and control (left panel). F. Effects of Icaritin on caspase-9, caspase-3, Apaf-1, bax, bcl-2 and cyt-c protein expression (western blot results). β-actin is used as loading control. G. Effects of Icaritin on apoptosis in Imatinib-resistant cells, CD34+cells from one CML patient with Imatinib-resistance, and CD34+ cells from 3 cases with CML-BC (Annexin V analysis).

    Journal: PLoS ONE

    Article Title: Icaritin Shows Potent Anti-Leukemia Activity on Chronic Myeloid Leukemia In Vitro and In Vivo by Regulating MAPK/ERK/JNK and JAK2/STAT3 /AKT Signalings

    doi: 10.1371/journal.pone.0023720

    Figure Lengend Snippet: Icaritin induces K562 cells or primary cells apoptosis. A. Morphological features for apoptosis in untreated and Icaritin-treated K562 cells were revealed by Hoechst 33258 staining. Condensed chromatin and apoptotic body could be found in Icaritin-treated K562 cells by fluorescence microscope (Olympus, B×50); ×400. B. Percentages of apoptotic cells in various concentration of Icaritin-treated K562 cells (Hoechst 33258 staining). The results represent mean±SD of triplicate experiments. C. Flow-cytometry analysis showed externalized PS, revealed by Annexin staining, increased significantly in 32 µM Icaritin-treated K562 cells. D. Percentages of apoptotic cells in Icaritin-treated fresh primary cells from CML-BP patients bone marrow (n = 5) based on annexin V expression assays. Error bars represent SD of experiments. E. Cell cycle analysis showing sub-G1 content in Icaritin (32 µM)-treated cells (right panel) and control (left panel). F. Effects of Icaritin on caspase-9, caspase-3, Apaf-1, bax, bcl-2 and cyt-c protein expression (western blot results). β-actin is used as loading control. G. Effects of Icaritin on apoptosis in Imatinib-resistant cells, CD34+cells from one CML patient with Imatinib-resistance, and CD34+ cells from 3 cases with CML-BC (Annexin V analysis).

    Article Snippet: The apoptotic morphology of K562 was evaluated by Hoechst 33258 (KeyGen Biotech Co, Ltd) following manufacturer's protocol.

    Techniques: Staining, Fluorescence, Microscopy, Concentration Assay, Flow Cytometry, Cytometry, Expressing, Cell Cycle Assay, Western Blot

    Immunohistochemical staining and electron micrographs of Sox17-transduced ball-like cell clusters. (A) Expression of CD31 and c-Kit was observed in Sox17-transduced ball-like cell clusters. After Cytospin centrifugation, ball-like cell clusters were stained with anti-CD31 (red) or anti-c-Kit (red) antibodies. Nuclei were stained with Hoechst 33258 (blue). Images on the right are higher magnifications of the boxed areas in the images on the left. Bar, 100 μm. (B) Electron micrographs of IRES-GFP (Mock)- and Sox17-IRES-GFP-transduced cells. Bar, 10 μm.

    Journal: Molecular and Cellular Biology

    Article Title: Sox17-Mediated Maintenance of Fetal Intra-Aortic Hematopoietic Cell Clusters

    doi: 10.1128/MCB.01485-13

    Figure Lengend Snippet: Immunohistochemical staining and electron micrographs of Sox17-transduced ball-like cell clusters. (A) Expression of CD31 and c-Kit was observed in Sox17-transduced ball-like cell clusters. After Cytospin centrifugation, ball-like cell clusters were stained with anti-CD31 (red) or anti-c-Kit (red) antibodies. Nuclei were stained with Hoechst 33258 (blue). Images on the right are higher magnifications of the boxed areas in the images on the left. Bar, 100 μm. (B) Electron micrographs of IRES-GFP (Mock)- and Sox17-IRES-GFP-transduced cells. Bar, 10 μm.

    Article Snippet: After washing, nuclei were counterstained with Hoechst 33258 (Nacalai Tesque, Kyoto, Japan).

    Techniques: Immunohistochemistry, Staining, Expressing, Centrifugation