fluorescence activated cell sorting  (Millipore)


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    Structured Review

    Millipore fluorescence activated cell sorting
    Fluorescence Activated Cell Sorting, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 28 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/fluorescence activated cell sorting/product/Millipore
    Average 99 stars, based on 28 article reviews
    Price from $9.99 to $1999.99
    fluorescence activated cell sorting - by Bioz Stars, 2020-05
    99/100 stars

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    Related Articles

    TUNEL Assay:

    Article Title: Autophagy mediates the beneficial effect of hypoxic preconditioning on bone marrow mesenchymal stem cells for the therapy of myocardial infarction
    Article Snippet: .. TUNEL staining was performed with fluorescein-dUTP (In Situ Cell Death Detection Kit; Roche Diagnostics) for apoptotic cell nuclei and DAPI (Sigma) to stain all cell nuclei. .. Additional staining for troponin I or GFP was performed for the identification of myocardium and transplanted BM-MSCs, respectively.

    In Situ:

    Article Title: Autophagy mediates the beneficial effect of hypoxic preconditioning on bone marrow mesenchymal stem cells for the therapy of myocardial infarction
    Article Snippet: .. TUNEL staining was performed with fluorescein-dUTP (In Situ Cell Death Detection Kit; Roche Diagnostics) for apoptotic cell nuclei and DAPI (Sigma) to stain all cell nuclei. .. Additional staining for troponin I or GFP was performed for the identification of myocardium and transplanted BM-MSCs, respectively.

    CCK-8 Assay:

    Article Title: Planar and Cell Aggregate-Like Assemblies Consisting of Microreactors and HepG2 Cells
    Article Snippet: .. Materials Sodium Alg, PLL (molecular weight of 40–60 kDa), sodium chloride (NaCl), calcium chloride (CaCl2 ), tris(hydroxymethyl)aminomethane (TRIS), HEPES, sodium bicarbonate (NaHCO3 ), Triton X-100, Span 80, TWEEN 80, 1 H,1 H,2 H,2 H-perfluorooctyltriethoxysilane (PFOTES), acetic acid, ethanol, chloroform (purity of ≥99.5%), phalloidin tetramethylrhodamine B isothiocyanate (phalloidin), DAPI, primary antibody monoclonal antivinculin, FDA and propidium iodide (PI), catalase from bovine liver (10 000 units mg–1 solid, 240 kDa), from horseradish peroxidase (HRP, 250–330 units mg–1 solid), cell counting kit-8 (CCK-8), and hydrogen peroxide (H2 O2 , 30 w/w %) were purchased from Sigma-Aldrich. .. 5-(4,6-Dichlorotriazinyl) aminofluorescein (5-DTAF), the secondary antibody Alexa Fluor 488 F(ab′)2 fragment of goat antimouse IgG, DyLight 633 maleimide, Quant-iT PicoGreen cell proliferation assay, and the Amplex Red catalase assay kit were purchased from Thermo Fisher Scientific.

    Incubation:

    Article Title: Dynamics of macrophage populations of the liver after subtotal hepatectomy in rats
    Article Snippet: .. After incubation with FITC-conjugated secondary antibodies (Abcam, Cambridge, UK) and the signal development, cell nuclei were counterstained with DAPI (Sigma-Aldrich, St. Louis, MO, USA). .. Macrophages derived from blood monocytes were selectively identified by anti-CX3CR1 immunostaining (Abcam, Cambridge, UK) and anti-CD11b immunostaining (Santa-Cruz, USA) in formalin-fixed paraffin-embedded hepatic tissue sections, in comparison with the splenic tissue sections for a positive control.

    Article Title: Biliary Polyunsaturated Fatty Acids and Telocytes in Gallstone Disease
    Article Snippet: .. After washing in PBS, the slides were incubated with (4,6-dichlorotriazinyl)aminofluorescein (DTAF)-conjugated streptavidin (016-010-084; 1:500 in PBS; Jackson ImmunoResearch) for 1 h. After a final rinse in PBS, the nuclei were counterstained with 4′,6-diamidino-2-phenylindole (DAPI) (D9542; 1:30,000; Sigma-Aldrich) for 30 s. The sections were mounted in Vectashield medium (H-1000; Vector Laboratories, Burlingame, CA, USA) to minimize fluorophore photobleaching. .. Slides were examined using an Olympus BX50 epifluorescence microscope (Olympus, Tokyo, Japan) equipped with an Olympus DP71 digital CCD camera.

    Staining:

    Article Title: Pyroptosis triggers pore-induced intracellular traps (PITs) that capture bacteria and lead to their clearance by efferocytosis
    Article Snippet: .. Fixed cells were permeabilized, blocked in 5% BSA/PBS, and stained with 0.1 µM Alexa Fluor 555 Phalloidin (Invitrogen) or antivimentin or antitubulin antibodies, 2 mM DAPI (Sigma-Aldrich), or anti–caspase-1 (Genentech). .. For live cell microscopy, cells were imaged in a humidified chamber at 5% CO2 37°C starting 15 min after infection and for 2 h in phenol-free imaging media.

    Article Title: Targeting ERK1/2-bim signaling cascades by BH3-mimetic ABT-737 as an alternative therapeutic strategy for oral cancer
    Article Snippet: .. 4′-6-diamidino-2-phenylindole (DAPI) staining To detect of nuclear morphological changes of apoptotic cells, cells were stained with DAPI solution (Sigma-Aldrich, Louis, MO, USA). .. Briefly, detached cells were fixed in 100% methanol at RT for 10 min, deposited on slides, and stained with DAPI solution (2 μg/ml).

    Article Title: Genetic Validation of Leishmania donovani Lysyl-tRNA Synthetase Shows that It Is Indispensable for Parasite Growth and Infectivity
    Article Snippet: .. The nuclear and the kinetoplastid DNA were then stained with 1 µg/ml of DAPI (Sigma) for 15 min. .. The fluorescence of the stained parasites was visualized by a confocal laser scanning microscope (Olympus FluoView FV1000 with PLAPON 60× O objective lenses; numerical aperture [NA], 1.42).

    Article Title: Autophagy mediates the beneficial effect of hypoxic preconditioning on bone marrow mesenchymal stem cells for the therapy of myocardial infarction
    Article Snippet: .. TUNEL staining was performed with fluorescein-dUTP (In Situ Cell Death Detection Kit; Roche Diagnostics) for apoptotic cell nuclei and DAPI (Sigma) to stain all cell nuclei. .. Additional staining for troponin I or GFP was performed for the identification of myocardium and transplanted BM-MSCs, respectively.

    Molecular Weight:

    Article Title: Planar and Cell Aggregate-Like Assemblies Consisting of Microreactors and HepG2 Cells
    Article Snippet: .. Materials Sodium Alg, PLL (molecular weight of 40–60 kDa), sodium chloride (NaCl), calcium chloride (CaCl2 ), tris(hydroxymethyl)aminomethane (TRIS), HEPES, sodium bicarbonate (NaHCO3 ), Triton X-100, Span 80, TWEEN 80, 1 H,1 H,2 H,2 H-perfluorooctyltriethoxysilane (PFOTES), acetic acid, ethanol, chloroform (purity of ≥99.5%), phalloidin tetramethylrhodamine B isothiocyanate (phalloidin), DAPI, primary antibody monoclonal antivinculin, FDA and propidium iodide (PI), catalase from bovine liver (10 000 units mg–1 solid, 240 kDa), from horseradish peroxidase (HRP, 250–330 units mg–1 solid), cell counting kit-8 (CCK-8), and hydrogen peroxide (H2 O2 , 30 w/w %) were purchased from Sigma-Aldrich. .. 5-(4,6-Dichlorotriazinyl) aminofluorescein (5-DTAF), the secondary antibody Alexa Fluor 488 F(ab′)2 fragment of goat antimouse IgG, DyLight 633 maleimide, Quant-iT PicoGreen cell proliferation assay, and the Amplex Red catalase assay kit were purchased from Thermo Fisher Scientific.

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  • 92
    Millipore 3 o trans p coumaroyltormentic acid
    The effect of 3 - O - p <t>-coumaroyltormentic</t> acid on the viability and mammosphere-forming ability of MCF-7 and MDA-MB-231 cells. ( A ) MCF-7 and MDA-MB-231 cells were treated with increasing concentrations of 3 - O - trans - p -coumaroyltormentic acid for 48 h. The antiproliferative effect of 3 - O - trans - p -coumaroyltormentic acid was measured by the MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2 H -tetrazolium) assay. ( B ) Effect of 3 - O - trans - p -coumaroyltormentic acid on the formation of mammospheres derived from MCF-7 and MDA-MB-231 cells. The mammospheres were incubated with either 3 - O - trans - p -coumaroyltormentic acid (10 and 20 μM) or DMSO for seven days (scale bar = 100 μm). ( C ) MCF-7 and MDA-MB-231 cells were treated with increasing concentrations of 3 - O - cis - p -coumaroyltormentic acid for 48 h. The antiproliferative effect of 3 - O - cis - p -coumaroyltormentic acid was measured by the MTS assay. ( D ) Effect of 3 - O - cis - p -coumaroyltormentic acid on the formation of mammospheres derived from MCF-7 and MDA-MB-231 cells. The mammospheres were incubated with either 3 - O - cis - p -coumaroyltormentic acid (20 and 40 μM) or DMSO. MCF-7 and MDA-MB-231 cells were treated with 3 - O - cis - p -coumaroyltormentic acid or DMSO in CSC culture media for seven days. Images were obtained by microscopy at 10× magnification and were representative mammospheres (scale bar = 100 μm). ( E ) Effect of 3 - O - trans - p -coumaroyltormentic acid on migratory potential of human breast cancer cells. The wound healing of MDA-MB-231 cells with or without 3 - O - trans - p -coumaroyltormentic acid photographed at 0 and 18 h (scale bar = 100 μm). ( F ) Effect of 3 - O - trans - p -coumaroyltormentic acid on colony formation on human breast cancer cells. The dissociated 1000 MDA-231-MB cells were seeded in six-well plates and treated with an indicated concentration of 3 - O - trans - p -coumaroyltormentic acid and DMSO for seven days. Representative images of colonies were recorded. The data shown represent the mean ± SD of three independent experiments. * p
    3 O Trans P Coumaroyltormentic Acid, supplied by Millipore, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/3 o trans p coumaroyltormentic acid/product/Millipore
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    89
    Millipore anti dectin 1
    Gene expression and translation of <t>dectin-1</t> signaling proteins. a Illustration of selected signaling molecules downstream of dectin-1; b Polysome profiles and c quantification of signalosome genes (qPCR) in monosome, disome, and light and heavy polysome fractions (monocytes). Data are from 4 subjects per age group (boxes and whiskers; RQ = relative quantification); d Quantification of signalosome genes (qPCR) in total RNA fractions (4 to 5 subjects/age group; mean ± SD); e Surface expression of dectin-1 (flow cytometry, mononuclear cells, gated on CD14-expressing cells; data pooled from 10 to 23 subjects per age group; boxes and whiskers); f Representative (cropped) Western blot of MALT1 and Bcl10 protein expression in monocytes after 0 to 60 min LPS stimulation. Representative blot is from a 29 weeks gestation sample. Images cropped from same blot probes with each antibody; cumulative quantification of 4 independent Western blot experiments for g MALT1 and h Bcl10 (mean ± SD)
    Anti Dectin 1, supplied by Millipore, used in various techniques. Bioz Stars score: 89/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti dectin 1/product/Millipore
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    93
    Millipore mhc class ii molecule
    Collagen-induced arthritis fibroblast-like synoviocytes (CIA-FLS) express major histocompatibility complex <t>(MHC)</t> class II molecules that are regulated through KCa1.1, as well as antigen-presenting proteins . a CIA-FLS plasma membrane expression of MHC class II protein following stimulation for 72 h with IFN-γ ( gray circles ), paxilline (Pax; black squares ), or interferon (IFN)-γ and paxilline ( open squares ). Data are presented as mean ± SEM ( n = 3–8 CIA-FLS donors). b Flow cytometric histogram of CIA-FLS stained for the plasma membrane expression of MHC class II protein in cells treated with paxilline for 72 h ( dashed line ), stimulated for 72 h with IFN-γ ( solid line ), or stimulated for 72 h with IFN-γ and paxilline ( dotted line ). The shaded histogram represents background staining. c MHC class II molecule expression of CIA-FLS that were permeabilized with saponin prior to staining for MHC class II in cells treated for 72 h with IFN-γ ( gray circles ), paxilline ( black squares ), or IFN-γ and paxilline ( open squares ). Data are presented as mean ± SEM ( n = 3–6 CIA-FLS donors). d Flow cytometric histogram of CIA-FLS stained for MHC class II molecules following permeabilization with saponin in cells treated for 72 h with IFN-γ ( solid line ), paxilline ( dashed line ), or IFN-γ and paxilline ( dotted line ). The shaded histogram represents background staining. e Expression of intercellular adhesion molecule (ICAM)-1, <t>B7-H3,</t> and CD40 by CIA-FLS following treatment for 72 h with IFN-γ, paxilline, or IFN-γ and paxilline. Data are presented as mean ± SEM ( n = 6 CIA-FLS donors). * p
    Mhc Class Ii Molecule, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Millipore membrane bound cg activity measurements sputum
    Introducing small-molecule FRET flow cytometry. (a–d) Representative flow cytometry neutrophil-specific ratiometric measurement of <t>surface-bound</t> <t>CG</t> <t>activity</t> at different time points after mSAM addition. (e) Average D/A ratio measured on <t>sputum</t> neutrophils derived from 11 human subjects (6 healthy donors and 5 CF) after 10 min and (f) their correlation with the respective microscopy measurement ( n = 50–70 for microscopy, n ≥ 1000 for flow cytometry). Data are represented as mean ± SD. Each human sample was measured in duplicate. Statistics were calculated by Wilcoxon rank sum and Spearman rank correlation tests.
    Membrane Bound Cg Activity Measurements Sputum, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/membrane bound cg activity measurements sputum/product/Millipore
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    Image Search Results


    The effect of 3 - O - p -coumaroyltormentic acid on the viability and mammosphere-forming ability of MCF-7 and MDA-MB-231 cells. ( A ) MCF-7 and MDA-MB-231 cells were treated with increasing concentrations of 3 - O - trans - p -coumaroyltormentic acid for 48 h. The antiproliferative effect of 3 - O - trans - p -coumaroyltormentic acid was measured by the MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2 H -tetrazolium) assay. ( B ) Effect of 3 - O - trans - p -coumaroyltormentic acid on the formation of mammospheres derived from MCF-7 and MDA-MB-231 cells. The mammospheres were incubated with either 3 - O - trans - p -coumaroyltormentic acid (10 and 20 μM) or DMSO for seven days (scale bar = 100 μm). ( C ) MCF-7 and MDA-MB-231 cells were treated with increasing concentrations of 3 - O - cis - p -coumaroyltormentic acid for 48 h. The antiproliferative effect of 3 - O - cis - p -coumaroyltormentic acid was measured by the MTS assay. ( D ) Effect of 3 - O - cis - p -coumaroyltormentic acid on the formation of mammospheres derived from MCF-7 and MDA-MB-231 cells. The mammospheres were incubated with either 3 - O - cis - p -coumaroyltormentic acid (20 and 40 μM) or DMSO. MCF-7 and MDA-MB-231 cells were treated with 3 - O - cis - p -coumaroyltormentic acid or DMSO in CSC culture media for seven days. Images were obtained by microscopy at 10× magnification and were representative mammospheres (scale bar = 100 μm). ( E ) Effect of 3 - O - trans - p -coumaroyltormentic acid on migratory potential of human breast cancer cells. The wound healing of MDA-MB-231 cells with or without 3 - O - trans - p -coumaroyltormentic acid photographed at 0 and 18 h (scale bar = 100 μm). ( F ) Effect of 3 - O - trans - p -coumaroyltormentic acid on colony formation on human breast cancer cells. The dissociated 1000 MDA-231-MB cells were seeded in six-well plates and treated with an indicated concentration of 3 - O - trans - p -coumaroyltormentic acid and DMSO for seven days. Representative images of colonies were recorded. The data shown represent the mean ± SD of three independent experiments. * p

    Journal: International Journal of Molecular Sciences

    Article Title: Triterpene Acid (3-O-p-Coumaroyltormentic Acid) Isolated From Aronia Extracts Inhibits Breast Cancer Stem Cell Formation through Downregulation of c-Myc Protein

    doi: 10.3390/ijms19092528

    Figure Lengend Snippet: The effect of 3 - O - p -coumaroyltormentic acid on the viability and mammosphere-forming ability of MCF-7 and MDA-MB-231 cells. ( A ) MCF-7 and MDA-MB-231 cells were treated with increasing concentrations of 3 - O - trans - p -coumaroyltormentic acid for 48 h. The antiproliferative effect of 3 - O - trans - p -coumaroyltormentic acid was measured by the MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2 H -tetrazolium) assay. ( B ) Effect of 3 - O - trans - p -coumaroyltormentic acid on the formation of mammospheres derived from MCF-7 and MDA-MB-231 cells. The mammospheres were incubated with either 3 - O - trans - p -coumaroyltormentic acid (10 and 20 μM) or DMSO for seven days (scale bar = 100 μm). ( C ) MCF-7 and MDA-MB-231 cells were treated with increasing concentrations of 3 - O - cis - p -coumaroyltormentic acid for 48 h. The antiproliferative effect of 3 - O - cis - p -coumaroyltormentic acid was measured by the MTS assay. ( D ) Effect of 3 - O - cis - p -coumaroyltormentic acid on the formation of mammospheres derived from MCF-7 and MDA-MB-231 cells. The mammospheres were incubated with either 3 - O - cis - p -coumaroyltormentic acid (20 and 40 μM) or DMSO. MCF-7 and MDA-MB-231 cells were treated with 3 - O - cis - p -coumaroyltormentic acid or DMSO in CSC culture media for seven days. Images were obtained by microscopy at 10× magnification and were representative mammospheres (scale bar = 100 μm). ( E ) Effect of 3 - O - trans - p -coumaroyltormentic acid on migratory potential of human breast cancer cells. The wound healing of MDA-MB-231 cells with or without 3 - O - trans - p -coumaroyltormentic acid photographed at 0 and 18 h (scale bar = 100 μm). ( F ) Effect of 3 - O - trans - p -coumaroyltormentic acid on colony formation on human breast cancer cells. The dissociated 1000 MDA-231-MB cells were seeded in six-well plates and treated with an indicated concentration of 3 - O - trans - p -coumaroyltormentic acid and DMSO for seven days. Representative images of colonies were recorded. The data shown represent the mean ± SD of three independent experiments. * p

    Article Snippet: Western Blotting Proteins isolated from MCF-7 and MDA-MB-231 mammospheres treated with 3-O -trans -p -coumaroyltormentic acid were separated on a 10% gel with SDS-PAGE and transferred to a polyvinylidene difluoride membrane (Millipore, Billerica, MA, USA).

    Techniques: Multiple Displacement Amplification, Derivative Assay, Incubation, MTS Assay, Microscopy, Concentration Assay

    Effect of 3 - O - trans - p -coumaroyltormentic acid on the expression of CSC markers in breast cancer cell line. ( A ) The CD44 high /CD24 low cell population was analyzed by flow cytometry in MDA-MB-231 cells treated with either 3 - O - trans - p -coumaroyltormentic acid (20 μM) or DMSO for two days. For FACS (Flow Cytometer) analysis, 50,000 cells were acquired. Gating was based on binding of the control antibody (red cross). ( B ) The effect of 3 - O - trans - p -coumaroyltormentic acid on the ALDH-positive cell population. MDA-MB-231 cells were treated with either 3 - O - trans - p -coumaroyltormentic acid (20 μM) or DMSO for two days and subjected to the ALDEFLUOR assay and FACS analysis. A set of representative flow cytometer dot plots is shown. The lower panel shows ALDH-positive cells in the presence of the ALDH inhibitor diethylaminobenzaldehyde (DEAB) as a negative control, and the upper panel represents ALDH-positive cells in the absence of DEAB. The ALDH-positive population was gated in the box (red dot line box).

    Journal: International Journal of Molecular Sciences

    Article Title: Triterpene Acid (3-O-p-Coumaroyltormentic Acid) Isolated From Aronia Extracts Inhibits Breast Cancer Stem Cell Formation through Downregulation of c-Myc Protein

    doi: 10.3390/ijms19092528

    Figure Lengend Snippet: Effect of 3 - O - trans - p -coumaroyltormentic acid on the expression of CSC markers in breast cancer cell line. ( A ) The CD44 high /CD24 low cell population was analyzed by flow cytometry in MDA-MB-231 cells treated with either 3 - O - trans - p -coumaroyltormentic acid (20 μM) or DMSO for two days. For FACS (Flow Cytometer) analysis, 50,000 cells were acquired. Gating was based on binding of the control antibody (red cross). ( B ) The effect of 3 - O - trans - p -coumaroyltormentic acid on the ALDH-positive cell population. MDA-MB-231 cells were treated with either 3 - O - trans - p -coumaroyltormentic acid (20 μM) or DMSO for two days and subjected to the ALDEFLUOR assay and FACS analysis. A set of representative flow cytometer dot plots is shown. The lower panel shows ALDH-positive cells in the presence of the ALDH inhibitor diethylaminobenzaldehyde (DEAB) as a negative control, and the upper panel represents ALDH-positive cells in the absence of DEAB. The ALDH-positive population was gated in the box (red dot line box).

    Article Snippet: Western Blotting Proteins isolated from MCF-7 and MDA-MB-231 mammospheres treated with 3-O -trans -p -coumaroyltormentic acid were separated on a 10% gel with SDS-PAGE and transferred to a polyvinylidene difluoride membrane (Millipore, Billerica, MA, USA).

    Techniques: Expressing, Flow Cytometry, Cytometry, Multiple Displacement Amplification, FACS, Binding Assay, Negative Control

    Molecular structure of the CSC inhibitor isolated from aronia. ( A , B ) Molecular mass of the CSC inhibitor derived from aronia based on electrospray ionization (ESI) mass spectrometry. ( C , D ) Molecular structure of 3 - O - trans - p -coumaroyltormentic acid and 3 - O - cis - p -coumaroyltormentic acid.

    Journal: International Journal of Molecular Sciences

    Article Title: Triterpene Acid (3-O-p-Coumaroyltormentic Acid) Isolated From Aronia Extracts Inhibits Breast Cancer Stem Cell Formation through Downregulation of c-Myc Protein

    doi: 10.3390/ijms19092528

    Figure Lengend Snippet: Molecular structure of the CSC inhibitor isolated from aronia. ( A , B ) Molecular mass of the CSC inhibitor derived from aronia based on electrospray ionization (ESI) mass spectrometry. ( C , D ) Molecular structure of 3 - O - trans - p -coumaroyltormentic acid and 3 - O - cis - p -coumaroyltormentic acid.

    Article Snippet: Western Blotting Proteins isolated from MCF-7 and MDA-MB-231 mammospheres treated with 3-O -trans -p -coumaroyltormentic acid were separated on a 10% gel with SDS-PAGE and transferred to a polyvinylidene difluoride membrane (Millipore, Billerica, MA, USA).

    Techniques: Isolation, Derivative Assay, Mass Spectrometry

    Effect of 3 - O - trans - p -coumaroyltormentic acid on the gene expression of CSC markers and mammosphere growth. ( A ) Transcriptional expression of the CSC markers Sox2, CD44, and Oct4 genes was determined in 3 - O - trans - p -coumaroyltormentic acid-treated and DMSO-treated mammospheres using specific primers for these genes and real-time RT-PCR. β-Actin served as an internal control. ( B ) Effect of 3 - O - trans - p -coumaroyltormentic acid on mammosphere growth. 3 - O - trans - p -coumaroyltormentic acid prevents mammosphere growth. After 3 - O - trans - p -coumaroyltormentic acid and DMSO treatment for two days, the mammospheres were dissociated into a single-cell suspension and plated in a six-cm dish with an equal number of cells. At 24 h after plating, the cells were counted. At two days and three days after replating, cells were counted in triplicate and plotted as the mean value. The data shown represent the mean ± SD of three independent experiments. * p

    Journal: International Journal of Molecular Sciences

    Article Title: Triterpene Acid (3-O-p-Coumaroyltormentic Acid) Isolated From Aronia Extracts Inhibits Breast Cancer Stem Cell Formation through Downregulation of c-Myc Protein

    doi: 10.3390/ijms19092528

    Figure Lengend Snippet: Effect of 3 - O - trans - p -coumaroyltormentic acid on the gene expression of CSC markers and mammosphere growth. ( A ) Transcriptional expression of the CSC markers Sox2, CD44, and Oct4 genes was determined in 3 - O - trans - p -coumaroyltormentic acid-treated and DMSO-treated mammospheres using specific primers for these genes and real-time RT-PCR. β-Actin served as an internal control. ( B ) Effect of 3 - O - trans - p -coumaroyltormentic acid on mammosphere growth. 3 - O - trans - p -coumaroyltormentic acid prevents mammosphere growth. After 3 - O - trans - p -coumaroyltormentic acid and DMSO treatment for two days, the mammospheres were dissociated into a single-cell suspension and plated in a six-cm dish with an equal number of cells. At 24 h after plating, the cells were counted. At two days and three days after replating, cells were counted in triplicate and plotted as the mean value. The data shown represent the mean ± SD of three independent experiments. * p

    Article Snippet: Western Blotting Proteins isolated from MCF-7 and MDA-MB-231 mammospheres treated with 3-O -trans -p -coumaroyltormentic acid were separated on a 10% gel with SDS-PAGE and transferred to a polyvinylidene difluoride membrane (Millipore, Billerica, MA, USA).

    Techniques: Expressing, Quantitative RT-PCR

    Gene expression and translation of dectin-1 signaling proteins. a Illustration of selected signaling molecules downstream of dectin-1; b Polysome profiles and c quantification of signalosome genes (qPCR) in monosome, disome, and light and heavy polysome fractions (monocytes). Data are from 4 subjects per age group (boxes and whiskers; RQ = relative quantification); d Quantification of signalosome genes (qPCR) in total RNA fractions (4 to 5 subjects/age group; mean ± SD); e Surface expression of dectin-1 (flow cytometry, mononuclear cells, gated on CD14-expressing cells; data pooled from 10 to 23 subjects per age group; boxes and whiskers); f Representative (cropped) Western blot of MALT1 and Bcl10 protein expression in monocytes after 0 to 60 min LPS stimulation. Representative blot is from a 29 weeks gestation sample. Images cropped from same blot probes with each antibody; cumulative quantification of 4 independent Western blot experiments for g MALT1 and h Bcl10 (mean ± SD)

    Journal: Nature Communications

    Article Title: Cellular metabolism constrains innate immune responses in early human ontogeny

    doi: 10.1038/s41467-018-07215-9

    Figure Lengend Snippet: Gene expression and translation of dectin-1 signaling proteins. a Illustration of selected signaling molecules downstream of dectin-1; b Polysome profiles and c quantification of signalosome genes (qPCR) in monosome, disome, and light and heavy polysome fractions (monocytes). Data are from 4 subjects per age group (boxes and whiskers; RQ = relative quantification); d Quantification of signalosome genes (qPCR) in total RNA fractions (4 to 5 subjects/age group; mean ± SD); e Surface expression of dectin-1 (flow cytometry, mononuclear cells, gated on CD14-expressing cells; data pooled from 10 to 23 subjects per age group; boxes and whiskers); f Representative (cropped) Western blot of MALT1 and Bcl10 protein expression in monocytes after 0 to 60 min LPS stimulation. Representative blot is from a 29 weeks gestation sample. Images cropped from same blot probes with each antibody; cumulative quantification of 4 independent Western blot experiments for g MALT1 and h Bcl10 (mean ± SD)

    Article Snippet: For blocking experiments, MNCs were pre-incubated for 1 h at 37 °C with neutralizing anti-dectin-1 or anti-dectin-2 antibodies, mepazine hydrochloride (10 µM, EMD Millipore #5005000001), cycloheximide (100 μg ml-1 , Sigma-Aldrich #C4859), ST 2825 (a MyD88 inhibitor, 10 µM, AdooQ Bioscience, #A15248-1), GW5074 (a Raf-1 inhibitor, 1 µM, Cayman Chemical #10010368-1), Piceatannol (a Syk inhibitor, 20 µM, Cayman Chemical, #10009355-5).

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Flow Cytometry, Cytometry, Western Blot

    MALT1 is essential for Candida recognition and curdlan signaling in human monocytes. IL-1β production after stimulation (24 h) with a curdlan or b LPS, or with c C. albicans or d C. parapsilosis , and upon inhibition (i) of MALT1 (using mepazine hydrochloride), MyD88 (ST2825), Raf (GW5074) or Syk (Piceatannol), or blocking of dectin-1 or dectin-2 using antibodies (data pooled from 6 experiments (6 subjects); mean ± SD); e Effect of MALT1 inhibition, dectin-1-blocking antibody or actin polymerization inhibition (using cytochalasin- d ) on phagocytosis of Candida by monocytes (boxes and whiskers; data also from 6 experiments). Statistical significance was calculated using 2-sided paired t -tests

    Journal: Nature Communications

    Article Title: Cellular metabolism constrains innate immune responses in early human ontogeny

    doi: 10.1038/s41467-018-07215-9

    Figure Lengend Snippet: MALT1 is essential for Candida recognition and curdlan signaling in human monocytes. IL-1β production after stimulation (24 h) with a curdlan or b LPS, or with c C. albicans or d C. parapsilosis , and upon inhibition (i) of MALT1 (using mepazine hydrochloride), MyD88 (ST2825), Raf (GW5074) or Syk (Piceatannol), or blocking of dectin-1 or dectin-2 using antibodies (data pooled from 6 experiments (6 subjects); mean ± SD); e Effect of MALT1 inhibition, dectin-1-blocking antibody or actin polymerization inhibition (using cytochalasin- d ) on phagocytosis of Candida by monocytes (boxes and whiskers; data also from 6 experiments). Statistical significance was calculated using 2-sided paired t -tests

    Article Snippet: For blocking experiments, MNCs were pre-incubated for 1 h at 37 °C with neutralizing anti-dectin-1 or anti-dectin-2 antibodies, mepazine hydrochloride (10 µM, EMD Millipore #5005000001), cycloheximide (100 μg ml-1 , Sigma-Aldrich #C4859), ST 2825 (a MyD88 inhibitor, 10 µM, AdooQ Bioscience, #A15248-1), GW5074 (a Raf-1 inhibitor, 1 µM, Cayman Chemical #10010368-1), Piceatannol (a Syk inhibitor, 20 µM, Cayman Chemical, #10009355-5).

    Techniques: Inhibition, Blocking Assay

    Impaired glycolysis and mTOR activity in preterm neonatal monocytes. a Effect of blocking glycolysis on cytokine responses to LPS stimulation (24 h) in mononuclear cells (2-sided paired t -tests; boxes and whiskers; 4 to 9 samples per condition; 2-DG = 2-deoxy-d-glucose); b Extracellular acidification rates (ECAR, normalized to total protein content), under baseline glucose-free conditions, after addition of glucose, oligomycin, and 2-DG (representative experiment from a 29 week gestation preterm sample); c Glycolytic capacity (cumulative data pooled from 4 independent experiments; 3–9 subjects per age group; mean ± SD; p value by Mann–Whitney U test); d Lactate production in mononuclear cells after LPS stimulation (24 h); p value = effect of age by 2-way ANOVA. Data from 9 to 10 subjects per age group; boxes and whiskers; e Effect of blocking glycolysis or translation (using cycloheximide, CHX) on the phagocytosis of C. albicans (C a ); boxes and whiskers; 3 subjects; Cyto-D = cytochalasin-D; f Depiction of signaling events between Toll-like receptor and Raf-1-mediated dectin-1 activation, mTOR phosphorylation, and increased glycolysis and protein synthesis; g Western blot image (cropped) of mTOR/4EBP1 expression and mTOR phosphorylation (monocytes; preterm sample from 29 weeks gestation; cropped images from same blot probed for mTOR, β-actin, and 4EBP1, and stripped/re-probed for phospho-mTOR); h Expression of mTOR-related genes (monocytes; 6–12 per age group; boxes and whiskers; 2-way ANOVA across age groups, only significant p values are shown)

    Journal: Nature Communications

    Article Title: Cellular metabolism constrains innate immune responses in early human ontogeny

    doi: 10.1038/s41467-018-07215-9

    Figure Lengend Snippet: Impaired glycolysis and mTOR activity in preterm neonatal monocytes. a Effect of blocking glycolysis on cytokine responses to LPS stimulation (24 h) in mononuclear cells (2-sided paired t -tests; boxes and whiskers; 4 to 9 samples per condition; 2-DG = 2-deoxy-d-glucose); b Extracellular acidification rates (ECAR, normalized to total protein content), under baseline glucose-free conditions, after addition of glucose, oligomycin, and 2-DG (representative experiment from a 29 week gestation preterm sample); c Glycolytic capacity (cumulative data pooled from 4 independent experiments; 3–9 subjects per age group; mean ± SD; p value by Mann–Whitney U test); d Lactate production in mononuclear cells after LPS stimulation (24 h); p value = effect of age by 2-way ANOVA. Data from 9 to 10 subjects per age group; boxes and whiskers; e Effect of blocking glycolysis or translation (using cycloheximide, CHX) on the phagocytosis of C. albicans (C a ); boxes and whiskers; 3 subjects; Cyto-D = cytochalasin-D; f Depiction of signaling events between Toll-like receptor and Raf-1-mediated dectin-1 activation, mTOR phosphorylation, and increased glycolysis and protein synthesis; g Western blot image (cropped) of mTOR/4EBP1 expression and mTOR phosphorylation (monocytes; preterm sample from 29 weeks gestation; cropped images from same blot probed for mTOR, β-actin, and 4EBP1, and stripped/re-probed for phospho-mTOR); h Expression of mTOR-related genes (monocytes; 6–12 per age group; boxes and whiskers; 2-way ANOVA across age groups, only significant p values are shown)

    Article Snippet: For blocking experiments, MNCs were pre-incubated for 1 h at 37 °C with neutralizing anti-dectin-1 or anti-dectin-2 antibodies, mepazine hydrochloride (10 µM, EMD Millipore #5005000001), cycloheximide (100 μg ml-1 , Sigma-Aldrich #C4859), ST 2825 (a MyD88 inhibitor, 10 µM, AdooQ Bioscience, #A15248-1), GW5074 (a Raf-1 inhibitor, 1 µM, Cayman Chemical #10010368-1), Piceatannol (a Syk inhibitor, 20 µM, Cayman Chemical, #10009355-5).

    Techniques: Activity Assay, Blocking Assay, MANN-WHITNEY, Activation Assay, Western Blot, Expressing

    Importance of dectin-1 in response to Candida in neonatal monocytes. a Antibody blocking of dectin-1 reduces cytokine production to C. albicans (C a ) in adult mononuclear cells (one-sided t -test with Welch’s correction for unequal variance; 6 to 12 subjects per condition; boxes and whiskers); b Phagocytosis of C. albicans upon blocking with anti-dectin-1 receptor antibody, same-isotype control or cytochalasin-D (Cyto-D); (11 to 18 subjects per age group; boxes and whiskers); Blocking of c C. albicans (C a ) or d C. parapsilosis (C p ) phagocytosis using laminarin (blocking dectin-1), mannan (blocking dectin-2 and mannose receptor), anti-DC-SIGN, and anti-CD206 antibodies, or a combination of all three antibodies (bar graph with mean ± standard deviation (SD); 2 to 3 subjects per age group)

    Journal: Nature Communications

    Article Title: Cellular metabolism constrains innate immune responses in early human ontogeny

    doi: 10.1038/s41467-018-07215-9

    Figure Lengend Snippet: Importance of dectin-1 in response to Candida in neonatal monocytes. a Antibody blocking of dectin-1 reduces cytokine production to C. albicans (C a ) in adult mononuclear cells (one-sided t -test with Welch’s correction for unequal variance; 6 to 12 subjects per condition; boxes and whiskers); b Phagocytosis of C. albicans upon blocking with anti-dectin-1 receptor antibody, same-isotype control or cytochalasin-D (Cyto-D); (11 to 18 subjects per age group; boxes and whiskers); Blocking of c C. albicans (C a ) or d C. parapsilosis (C p ) phagocytosis using laminarin (blocking dectin-1), mannan (blocking dectin-2 and mannose receptor), anti-DC-SIGN, and anti-CD206 antibodies, or a combination of all three antibodies (bar graph with mean ± standard deviation (SD); 2 to 3 subjects per age group)

    Article Snippet: For blocking experiments, MNCs were pre-incubated for 1 h at 37 °C with neutralizing anti-dectin-1 or anti-dectin-2 antibodies, mepazine hydrochloride (10 µM, EMD Millipore #5005000001), cycloheximide (100 μg ml-1 , Sigma-Aldrich #C4859), ST 2825 (a MyD88 inhibitor, 10 µM, AdooQ Bioscience, #A15248-1), GW5074 (a Raf-1 inhibitor, 1 µM, Cayman Chemical #10010368-1), Piceatannol (a Syk inhibitor, 20 µM, Cayman Chemical, #10009355-5).

    Techniques: Blocking Assay, Standard Deviation

    Collagen-induced arthritis fibroblast-like synoviocytes (CIA-FLS) express major histocompatibility complex (MHC) class II molecules that are regulated through KCa1.1, as well as antigen-presenting proteins . a CIA-FLS plasma membrane expression of MHC class II protein following stimulation for 72 h with IFN-γ ( gray circles ), paxilline (Pax; black squares ), or interferon (IFN)-γ and paxilline ( open squares ). Data are presented as mean ± SEM ( n = 3–8 CIA-FLS donors). b Flow cytometric histogram of CIA-FLS stained for the plasma membrane expression of MHC class II protein in cells treated with paxilline for 72 h ( dashed line ), stimulated for 72 h with IFN-γ ( solid line ), or stimulated for 72 h with IFN-γ and paxilline ( dotted line ). The shaded histogram represents background staining. c MHC class II molecule expression of CIA-FLS that were permeabilized with saponin prior to staining for MHC class II in cells treated for 72 h with IFN-γ ( gray circles ), paxilline ( black squares ), or IFN-γ and paxilline ( open squares ). Data are presented as mean ± SEM ( n = 3–6 CIA-FLS donors). d Flow cytometric histogram of CIA-FLS stained for MHC class II molecules following permeabilization with saponin in cells treated for 72 h with IFN-γ ( solid line ), paxilline ( dashed line ), or IFN-γ and paxilline ( dotted line ). The shaded histogram represents background staining. e Expression of intercellular adhesion molecule (ICAM)-1, B7-H3, and CD40 by CIA-FLS following treatment for 72 h with IFN-γ, paxilline, or IFN-γ and paxilline. Data are presented as mean ± SEM ( n = 6 CIA-FLS donors). * p

    Journal: Arthritis Research & Therapy

    Article Title: KCa1.1 and Kv1.3 channels regulate the interactions between fibroblast-like synoviocytes and T lymphocytes during rheumatoid arthritis

    doi: 10.1186/s13075-018-1783-9

    Figure Lengend Snippet: Collagen-induced arthritis fibroblast-like synoviocytes (CIA-FLS) express major histocompatibility complex (MHC) class II molecules that are regulated through KCa1.1, as well as antigen-presenting proteins . a CIA-FLS plasma membrane expression of MHC class II protein following stimulation for 72 h with IFN-γ ( gray circles ), paxilline (Pax; black squares ), or interferon (IFN)-γ and paxilline ( open squares ). Data are presented as mean ± SEM ( n = 3–8 CIA-FLS donors). b Flow cytometric histogram of CIA-FLS stained for the plasma membrane expression of MHC class II protein in cells treated with paxilline for 72 h ( dashed line ), stimulated for 72 h with IFN-γ ( solid line ), or stimulated for 72 h with IFN-γ and paxilline ( dotted line ). The shaded histogram represents background staining. c MHC class II molecule expression of CIA-FLS that were permeabilized with saponin prior to staining for MHC class II in cells treated for 72 h with IFN-γ ( gray circles ), paxilline ( black squares ), or IFN-γ and paxilline ( open squares ). Data are presented as mean ± SEM ( n = 3–6 CIA-FLS donors). d Flow cytometric histogram of CIA-FLS stained for MHC class II molecules following permeabilization with saponin in cells treated for 72 h with IFN-γ ( solid line ), paxilline ( dashed line ), or IFN-γ and paxilline ( dotted line ). The shaded histogram represents background staining. e Expression of intercellular adhesion molecule (ICAM)-1, B7-H3, and CD40 by CIA-FLS following treatment for 72 h with IFN-γ, paxilline, or IFN-γ and paxilline. Data are presented as mean ± SEM ( n = 6 CIA-FLS donors). * p

    Article Snippet: Measuring MHC class II molecule, B7-H3, ICAM-1, and CD40 expression levels in CIA-FLS CIA-FLS were treated with 100 ng/ml recombinant IFN-γ (MilliporeSigma, Burlington, MA, USA) for 72 h in the presence or absence of 20 μM paxilline.

    Techniques: Expressing, Flow Cytometry, Staining

    Introducing small-molecule FRET flow cytometry. (a–d) Representative flow cytometry neutrophil-specific ratiometric measurement of surface-bound CG activity at different time points after mSAM addition. (e) Average D/A ratio measured on sputum neutrophils derived from 11 human subjects (6 healthy donors and 5 CF) after 10 min and (f) their correlation with the respective microscopy measurement ( n = 50–70 for microscopy, n ≥ 1000 for flow cytometry). Data are represented as mean ± SD. Each human sample was measured in duplicate. Statistics were calculated by Wilcoxon rank sum and Spearman rank correlation tests.

    Journal: ACS Central Science

    Article Title: Cathepsin G Activity as a New Marker for Detecting Airway Inflammation by Microscopy and Flow Cytometry

    doi: 10.1021/acscentsci.8b00933

    Figure Lengend Snippet: Introducing small-molecule FRET flow cytometry. (a–d) Representative flow cytometry neutrophil-specific ratiometric measurement of surface-bound CG activity at different time points after mSAM addition. (e) Average D/A ratio measured on sputum neutrophils derived from 11 human subjects (6 healthy donors and 5 CF) after 10 min and (f) their correlation with the respective microscopy measurement ( n = 50–70 for microscopy, n ≥ 1000 for flow cytometry). Data are represented as mean ± SD. Each human sample was measured in duplicate. Statistics were calculated by Wilcoxon rank sum and Spearman rank correlation tests.

    Article Snippet: Sputum Processing and Membrane-Bound CG Activity Measurements Sputum was separated from saliva by adding 4 volumes of 10% Sputolysin solution (Calbiochem, Darmstadt, Germany) followed by 15 min of mild shaking at room temperature.

    Techniques: Flow Cytometry, Cytometry, Activity Assay, Derivative Assay, Microscopy