Structured Review

Biowest SAS fbs
NEFAs directly reduce Mg 2+ concentrations. Linear regression analyses of Mg 2+ and NEFA concentration in ( a ) BSA dissolved in 1 <t>mmol/l</t> MgCl 2 ( y = −0.12 x + 0.83, r 2 = 0.97, p ≤ 0.05), ( b ) <t>FBS</t> ( y = −0.10 x + 1.36, r 2 = 0.90, p ≤ 0.05) and ( c ) MgCl 2 solution ( y = −0.08 x + 1.00, r 2 = 0.99, p ≤ 0.05). Results from one representative experiment are shown. The experiment was repeated three additional times with similar results
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Images

1) Product Images from "Increased NEFA levels reduce blood Mg2+ in hypertriacylglycerolaemic states via direct binding of NEFA to Mg2+"

Article Title: Increased NEFA levels reduce blood Mg2+ in hypertriacylglycerolaemic states via direct binding of NEFA to Mg2+

Journal: Diabetologia

doi: 10.1007/s00125-018-4771-3

NEFAs directly reduce Mg 2+ concentrations. Linear regression analyses of Mg 2+ and NEFA concentration in ( a ) BSA dissolved in 1 mmol/l MgCl 2 ( y = −0.12 x + 0.83, r 2 = 0.97, p ≤ 0.05), ( b ) FBS ( y = −0.10 x + 1.36, r 2 = 0.90, p ≤ 0.05) and ( c ) MgCl 2 solution ( y = −0.08 x + 1.00, r 2 = 0.99, p ≤ 0.05). Results from one representative experiment are shown. The experiment was repeated three additional times with similar results
Figure Legend Snippet: NEFAs directly reduce Mg 2+ concentrations. Linear regression analyses of Mg 2+ and NEFA concentration in ( a ) BSA dissolved in 1 mmol/l MgCl 2 ( y = −0.12 x + 0.83, r 2 = 0.97, p ≤ 0.05), ( b ) FBS ( y = −0.10 x + 1.36, r 2 = 0.90, p ≤ 0.05) and ( c ) MgCl 2 solution ( y = −0.08 x + 1.00, r 2 = 0.99, p ≤ 0.05). Results from one representative experiment are shown. The experiment was repeated three additional times with similar results

Techniques Used: Concentration Assay

2) Product Images from "Hexacosenoyl-CoA is the most abundant very long-chain acyl-CoA in ATP binding cassette transporter D1-deficient cells [S]"

Article Title: Hexacosenoyl-CoA is the most abundant very long-chain acyl-CoA in ATP binding cassette transporter D1-deficient cells [S]

Journal: Journal of Lipid Research

doi: 10.1194/jlr.P119000325

Pulse-chase analysis of VLCFA-CoA species using FA D 2 -18:1. Metabolic profiles of 15 acyl-CoA species containing a deuterium-labeled (A) or nonlabeled (B) acyl moiety in ABCD1-KO (#1) and WT HeLa cells were examined. Cells were cultured in medium containing 10% FBS and treated with 30 μM of FA D 2 -18:1 and the MβCD complex for an hour, after which the medium was replaced with fresh medium containing 10% FBS. The cells were then harvested at 1, 2, 4, 6, and 24 h after treatment. Acyl-CoA species were extracted and analyzed by LC-MS/MS analysis. The MRM transitions (Q1/Q3) used are indicated in each panel. Statistical analyses were performed with Student’s t -test; * P
Figure Legend Snippet: Pulse-chase analysis of VLCFA-CoA species using FA D 2 -18:1. Metabolic profiles of 15 acyl-CoA species containing a deuterium-labeled (A) or nonlabeled (B) acyl moiety in ABCD1-KO (#1) and WT HeLa cells were examined. Cells were cultured in medium containing 10% FBS and treated with 30 μM of FA D 2 -18:1 and the MβCD complex for an hour, after which the medium was replaced with fresh medium containing 10% FBS. The cells were then harvested at 1, 2, 4, 6, and 24 h after treatment. Acyl-CoA species were extracted and analyzed by LC-MS/MS analysis. The MRM transitions (Q1/Q3) used are indicated in each panel. Statistical analyses were performed with Student’s t -test; * P

Techniques Used: Pulse Chase, Labeling, Cell Culture, Liquid Chromatography with Mass Spectroscopy

Metabolic analysis of VLCFA-CoA species using FA D 2 -18:1. Metabolic profiles of each deuterium-labeled 18:1- and 26:1-CoA species (A) and nonlabeled 18:1- and 26:1-CoA species (B) were examined in ABCD1-KO (#1) and WT HeLa cells. The metabolic profiles of the other labeled and nonlabeled acyl-CoA species are depicted in supplemental Fig. S3. Cells were cultured in medium containing 10% FBS and treated with 30 μM of FA D 2 -18:1 and the MβCD complex. The cells were harvested at 1, 3, 5, 8, and 24 h after treatment. The MRM transitions (Q1/Q3) used are indicated in each panel. Statistical analyses were performed with the Student’s t -test; * P
Figure Legend Snippet: Metabolic analysis of VLCFA-CoA species using FA D 2 -18:1. Metabolic profiles of each deuterium-labeled 18:1- and 26:1-CoA species (A) and nonlabeled 18:1- and 26:1-CoA species (B) were examined in ABCD1-KO (#1) and WT HeLa cells. The metabolic profiles of the other labeled and nonlabeled acyl-CoA species are depicted in supplemental Fig. S3. Cells were cultured in medium containing 10% FBS and treated with 30 μM of FA D 2 -18:1 and the MβCD complex. The cells were harvested at 1, 3, 5, 8, and 24 h after treatment. The MRM transitions (Q1/Q3) used are indicated in each panel. Statistical analyses were performed with the Student’s t -test; * P

Techniques Used: Labeling, Cell Culture

Metabolic analysis of VLCFA-CoA species using FA D 4 -26:0. Metabolic profiles of 26:0- and 26:1-CoA species containing a deuterium-labeled (A) or nonlabeled (B) acyl moiety in ABCD1-KO (#1) and WT HeLa cells (WT). Cells were cultured in medium containing 10% FBS and were treated with 30 μM of FA D 4 -26:0 and the MβCD complex. Cells were harvested at 1, 3, 5, 8, and 24 h after treatment. We confirmed that the retention times of each deuterium-labeled acyl-CoA species were almost identical with those of the corresponding nonlabeled VLCFA-CoA species. The MRM transitions (Q1/Q3) used are indicated in each panel. Data represent the mean ± SD, n = 3. Statistical analyses were performed with the Student’s t -test; * P
Figure Legend Snippet: Metabolic analysis of VLCFA-CoA species using FA D 4 -26:0. Metabolic profiles of 26:0- and 26:1-CoA species containing a deuterium-labeled (A) or nonlabeled (B) acyl moiety in ABCD1-KO (#1) and WT HeLa cells (WT). Cells were cultured in medium containing 10% FBS and were treated with 30 μM of FA D 4 -26:0 and the MβCD complex. Cells were harvested at 1, 3, 5, 8, and 24 h after treatment. We confirmed that the retention times of each deuterium-labeled acyl-CoA species were almost identical with those of the corresponding nonlabeled VLCFA-CoA species. The MRM transitions (Q1/Q3) used are indicated in each panel. Data represent the mean ± SD, n = 3. Statistical analyses were performed with the Student’s t -test; * P

Techniques Used: Labeling, Cell Culture

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Article Snippet: After this was done, the Biowest FBS and ATL BIOL FBS were selected for full growth curve comparisons (both 10%, just following the cells more closely).

Article Title: Human Cells Grown With or Without Substitutes for Fetal Bovine Serum
Article Snippet: For these studies we selected only the best supplements: ATCC FBS, Gibco FBS, Biowest FBS, ATL BIOL FBS, Seradigm FBS, and the Fetalgro® alternative.

Article Title: Human Cells Grown With or Without Substitutes for Fetal Bovine Serum
Article Snippet: They were ATCC® FBS, Gibco FBS, Biowest FBS, ATL BIOL FBS, Seradigm FBS and RMBI FetalGro® .

Transfection:

Article Title: Ligand chain length drives activation of lipid G protein-coupled receptors
Article Snippet: Cells were passaged, after one wash with Ca2+ and Mg2+ -free Dubelcco’s Phosphate Saline Buffer (PBS), using Trypsin-EDTA mixture at 170 U/mL–0.2 mg/mL (BioWhittaker, Lonza). .. Transient transfection for S1P1 expression AequoScreen® CHO-K1 parental cell line stably transfected with pCAEQG plasmid leading to the co-expression of apoaequorin along with the promiscuous G protein subtype Gα16 were grown as described in section B, except from the replacement of regular FBS by Ultra-low Endotoxin FBS (BioWest). .. CHO-K1 parental cells were seeded in 6-well pates (Greiner) in medium without antibiotics 24 h before transfection, to reach a 50–70% confluency.

Expressing:

Article Title: Ligand chain length drives activation of lipid G protein-coupled receptors
Article Snippet: Cells were passaged, after one wash with Ca2+ and Mg2+ -free Dubelcco’s Phosphate Saline Buffer (PBS), using Trypsin-EDTA mixture at 170 U/mL–0.2 mg/mL (BioWhittaker, Lonza). .. Transient transfection for S1P1 expression AequoScreen® CHO-K1 parental cell line stably transfected with pCAEQG plasmid leading to the co-expression of apoaequorin along with the promiscuous G protein subtype Gα16 were grown as described in section B, except from the replacement of regular FBS by Ultra-low Endotoxin FBS (BioWest). .. CHO-K1 parental cells were seeded in 6-well pates (Greiner) in medium without antibiotics 24 h before transfection, to reach a 50–70% confluency.

Stable Transfection:

Article Title: Ligand chain length drives activation of lipid G protein-coupled receptors
Article Snippet: Cells were passaged, after one wash with Ca2+ and Mg2+ -free Dubelcco’s Phosphate Saline Buffer (PBS), using Trypsin-EDTA mixture at 170 U/mL–0.2 mg/mL (BioWhittaker, Lonza). .. Transient transfection for S1P1 expression AequoScreen® CHO-K1 parental cell line stably transfected with pCAEQG plasmid leading to the co-expression of apoaequorin along with the promiscuous G protein subtype Gα16 were grown as described in section B, except from the replacement of regular FBS by Ultra-low Endotoxin FBS (BioWest). .. CHO-K1 parental cells were seeded in 6-well pates (Greiner) in medium without antibiotics 24 h before transfection, to reach a 50–70% confluency.

Plasmid Preparation:

Article Title: Ligand chain length drives activation of lipid G protein-coupled receptors
Article Snippet: Cells were passaged, after one wash with Ca2+ and Mg2+ -free Dubelcco’s Phosphate Saline Buffer (PBS), using Trypsin-EDTA mixture at 170 U/mL–0.2 mg/mL (BioWhittaker, Lonza). .. Transient transfection for S1P1 expression AequoScreen® CHO-K1 parental cell line stably transfected with pCAEQG plasmid leading to the co-expression of apoaequorin along with the promiscuous G protein subtype Gα16 were grown as described in section B, except from the replacement of regular FBS by Ultra-low Endotoxin FBS (BioWest). .. CHO-K1 parental cells were seeded in 6-well pates (Greiner) in medium without antibiotics 24 h before transfection, to reach a 50–70% confluency.

Cell Culture:

Article Title: Thiazoles with cyclopropyl fragment as antifungal, anticonvulsant, and anti-Toxoplasma gondii agents: synthesis, toxicity evaluation, and molecular docking study
Article Snippet: Antiparasitic activity .. Cell and parasite culture L929 cell line (ATCC® CCL-1™) was cultured in IMDM (IMDM/10%FBS/P/S) (Iscove’s Modified Dulbecco’s Medium—Biowest) culture medium with the addition of 10% FBS (Fetal Bovine Serum—Biowest), 100 μg/ml streptomycin (Sigma), 100 U/ml penicillin (Sigma). .. Vero cell line (ATCC® CCL-81™ ) were maintained in EMEM (EMEM/10%FBS/P/S) (Eagle’s Minimum Essential Medium ATCC® 30-2003™) culture medium supplemented with 10% FBS, 100 μg/ml streptomycin, 100 U/ml penicillin.

Modification:

Article Title: Thiazoles with cyclopropyl fragment as antifungal, anticonvulsant, and anti-Toxoplasma gondii agents: synthesis, toxicity evaluation, and molecular docking study
Article Snippet: Antiparasitic activity .. Cell and parasite culture L929 cell line (ATCC® CCL-1™) was cultured in IMDM (IMDM/10%FBS/P/S) (Iscove’s Modified Dulbecco’s Medium—Biowest) culture medium with the addition of 10% FBS (Fetal Bovine Serum—Biowest), 100 μg/ml streptomycin (Sigma), 100 U/ml penicillin (Sigma). .. Vero cell line (ATCC® CCL-81™ ) were maintained in EMEM (EMEM/10%FBS/P/S) (Eagle’s Minimum Essential Medium ATCC® 30-2003™) culture medium supplemented with 10% FBS, 100 μg/ml streptomycin, 100 U/ml penicillin.

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    Biowest SAS fbs
    (A,B) Time of addition assay for analysis of mechanism of CAM and SCM against influenza A/shanghai/37T (2009H1N1) infection. MDCK cells cultured in <t>DMEM</t> supplemented with 10% <t>FBS</t> (Biowest, France), penicillin (100 U/ml), and streptomycin (10 μg/ml) at 37°C/5% CO 2 were infected with influenza A/shanghai/37T (2009H1N1) at 0.1 MOI in DMEM containing 2 μg/mL TPCK-Trypsin. CAM and SCM diluted in DMEM to give a final concentration of 20 μM were added to mixture of virus and cells at 0, 0.5, 1, 2, 4, and 12 h post infection. Inhibition activity of CAM and SCM was measured by CPE reduction assay using CCK-8, according to the manufacturer’s instruction. The data were presented as means ± SE of triplicate assays and the experiment was repeated at least twice.
    Fbs, supplied by Biowest SAS, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Biowest SAS dextran charcoal treated fbs
    Release of NO from JS-K . (A) Generation of NO from JS-K in <t>RPMI-1640</t> medium containing 10% fetal bovine serum was determined indirectly by photorimetric detection of nitrite according to Green et al. [ 31 ]. (B) Detection of nitrotyrosine by fluorescence microscopy in 22Rv1 cells: Generation of intracellular NO was detected indirectly by detection of nitrotyrosine (green). Cell nuclei were stained with DAPI. [A] untreated cells, [B] 2 μM JS-K neg , [C] 2 μM JS-K, [D] 200 μM DETA/NO.
    Dextran Charcoal Treated Fbs, supplied by Biowest SAS, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Biowest SAS fetal bovine serum fbs
    Crizotinib-induced apoptosis in Ba/F3 cells transformed by NPM-ALK. Ba/F3 cells were infected with an empty virus (-) and retrovirus expressing NPM-ALK or TEL-JAK2. (A) Whole cell lysates were immunoblotted with an anti-ALK antibody, anti-Flag antibody, anti-JAK2 antibody, anti-phospho-STAT3 antibody, anti-STAT3 antibody, anti-phospho-STAT5 antibody, anti-STAT5 antibody or anti-β-actin antibody. (B) Transduced Ba/F3 cells were incubated with <t>RPMI</t> containing 10% <t>FBS</t> in the absence of IL-3 for 2 days. Viable cell numbers at 24 hr and 48 hr were counted by the Trypan blue exclusion method. Values are the mean ± SD of three independent experiments. (C-F) Transduced Ba/F3 cells were incubated with RPMI containing 10% FBS in the presence of crizotinib (0.25, 0.5, 1 μM) and MMC (0.1, 1, 10 μg/mL) for 24 hr. (C) Cell viability was measured by the WST assay. Values are the mean ± S.D. of four independent experiments. (D) Cells were fixed, treated with propidium iodide, and subjected to a flow cytometric analysis. The percentages of cells in the sub-G1 phase were graphed. Values are the mean ± S.D. of four independent experiments. (E) To evaluate the apoptotic cell death, the chromatin DNA was isolated from cells and subjected to agarose gel electrophoresis. (F) Whole cell lysates were prepared and the phosphorylation of STAT3 and STAT5 was analyzed by immunoblotting. The relative phosphorylation level of STAT3 and STAT5 is shown in the graphs. Values are given as the mean ± SD of three independent experiments. *** P
    Fetal Bovine Serum Fbs, supplied by Biowest SAS, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    (A,B) Time of addition assay for analysis of mechanism of CAM and SCM against influenza A/shanghai/37T (2009H1N1) infection. MDCK cells cultured in DMEM supplemented with 10% FBS (Biowest, France), penicillin (100 U/ml), and streptomycin (10 μg/ml) at 37°C/5% CO 2 were infected with influenza A/shanghai/37T (2009H1N1) at 0.1 MOI in DMEM containing 2 μg/mL TPCK-Trypsin. CAM and SCM diluted in DMEM to give a final concentration of 20 μM were added to mixture of virus and cells at 0, 0.5, 1, 2, 4, and 12 h post infection. Inhibition activity of CAM and SCM was measured by CPE reduction assay using CCK-8, according to the manufacturer’s instruction. The data were presented as means ± SE of triplicate assays and the experiment was repeated at least twice.

    Journal: Frontiers in Microbiology

    Article Title: The Antihistamine Drugs Carbinoxamine Maleate and Chlorpheniramine Maleate Exhibit Potent Antiviral Activity Against a Broad Spectrum of Influenza Viruses

    doi: 10.3389/fmicb.2018.02643

    Figure Lengend Snippet: (A,B) Time of addition assay for analysis of mechanism of CAM and SCM against influenza A/shanghai/37T (2009H1N1) infection. MDCK cells cultured in DMEM supplemented with 10% FBS (Biowest, France), penicillin (100 U/ml), and streptomycin (10 μg/ml) at 37°C/5% CO 2 were infected with influenza A/shanghai/37T (2009H1N1) at 0.1 MOI in DMEM containing 2 μg/mL TPCK-Trypsin. CAM and SCM diluted in DMEM to give a final concentration of 20 μM were added to mixture of virus and cells at 0, 0.5, 1, 2, 4, and 12 h post infection. Inhibition activity of CAM and SCM was measured by CPE reduction assay using CCK-8, according to the manufacturer’s instruction. The data were presented as means ± SE of triplicate assays and the experiment was repeated at least twice.

    Article Snippet: Cells were cultured in DMEM (Thermo Fisher Scientific, Waltham, MA, United States) containing 10% FBS (Biowest, France), penicillin 100 U/ml, and streptomycin 10 μg/ml.

    Techniques: Chick Chorioallantoic Membrane Assay, Infection, Cell Culture, Concentration Assay, Inhibition, Activity Assay, CCK-8 Assay

    Cell morphology and gene and protein expression of Col2a1, Col1a1, Sox9, and MMP13 in chondrocytes stimulated with RS. A. Rat chondrocytes were cultured with DMEM plus 5% RS (5R) or DMEM plus 10% FBS (10F) for 24 h, and cell cultures were analyzed by microscopy. The chondrocytes dedifferentiated into fibroblastic chondrocytes after normal medium (DMEM plus 10% FBS, 10F) was replaced with DMEM containing 5% RS (5R). B. Col2a1, Col1a1 and MMP13 protein levels were analyzed by western blot. Col1a1 and MMP13 protein expression increased and Col2a1 protein expression decreased in the 5R group when compared to the 10F group. C. In situ expression of Col2a1, Col1a1, and Sox9 were detected in chondrocytes cultured with 5% RS or 10% FBS. The blue color indicates DAPI staining of nuclei. Red indicates Col2a1 expression and green indicates Col1a1 or Sox9 expression. Col1a1 and Sox9 expression increased and Col2a1 expression decreased in the 5R group when compared to the 10F group. D. Chondrocytes were cultured with RS or FBS for 24 h, mRNA was isolated from the chondrocytes, and expression of matrix degradation enzymes and collagen synthesis genes were analyzed by qRT-PCR. Data are expressed as the mean ± SD from three separate experiments. * P

    Journal: American Journal of Translational Research

    Article Title: The MEK-ERK1/2 signaling pathway regulates hyaline cartilage formation and the redifferentiation of dedifferentiated chondrocytes in vitro

    doi:

    Figure Lengend Snippet: Cell morphology and gene and protein expression of Col2a1, Col1a1, Sox9, and MMP13 in chondrocytes stimulated with RS. A. Rat chondrocytes were cultured with DMEM plus 5% RS (5R) or DMEM plus 10% FBS (10F) for 24 h, and cell cultures were analyzed by microscopy. The chondrocytes dedifferentiated into fibroblastic chondrocytes after normal medium (DMEM plus 10% FBS, 10F) was replaced with DMEM containing 5% RS (5R). B. Col2a1, Col1a1 and MMP13 protein levels were analyzed by western blot. Col1a1 and MMP13 protein expression increased and Col2a1 protein expression decreased in the 5R group when compared to the 10F group. C. In situ expression of Col2a1, Col1a1, and Sox9 were detected in chondrocytes cultured with 5% RS or 10% FBS. The blue color indicates DAPI staining of nuclei. Red indicates Col2a1 expression and green indicates Col1a1 or Sox9 expression. Col1a1 and Sox9 expression increased and Col2a1 expression decreased in the 5R group when compared to the 10F group. D. Chondrocytes were cultured with RS or FBS for 24 h, mRNA was isolated from the chondrocytes, and expression of matrix degradation enzymes and collagen synthesis genes were analyzed by qRT-PCR. Data are expressed as the mean ± SD from three separate experiments. * P

    Article Snippet: Chondrocytes were isolated from the articular cartilage of 24-hour-old Sprague-Dawley (SD) rats and dispersed in 0.1% collagenase type II (C6885, Sigma-Aldrich, Switzerland) for 3 h. Chondrocytes were collected and cultured in Dulbecco’s Modified Eagle’s Medium (DMEM, Biowest, France) supplemented with 10% FBS (Biowest, France) and 1% penicillin-streptomycin.

    Techniques: Expressing, Cell Culture, Microscopy, Western Blot, In Situ, Staining, Isolation, Quantitative RT-PCR

    Analysis of GAG content and gene expression in chondrocytes cultured with the MEK inhibitor PD0325901. A. Chondrocytes were cultured in DMEM supplemented with 5% RS (5R), 10% FBS (10F), or 5% RS plus various PD0325901 concentrations for 24 h. Chondrocytes were stained with Safranin O (left), and the GAG content in the chondrocytes was measured using the DMB method (right). B. Gene expression of matrix degradation enzymes and collagen synthesis genes in chondrocytes was evaluated by qPCR after treatment with and without various concentrations of the MEK-ERK1/2 pathway inhibitor PD0325901. Data are expressed as the mean ± SD from three separate experiments. # P

    Journal: American Journal of Translational Research

    Article Title: The MEK-ERK1/2 signaling pathway regulates hyaline cartilage formation and the redifferentiation of dedifferentiated chondrocytes in vitro

    doi:

    Figure Lengend Snippet: Analysis of GAG content and gene expression in chondrocytes cultured with the MEK inhibitor PD0325901. A. Chondrocytes were cultured in DMEM supplemented with 5% RS (5R), 10% FBS (10F), or 5% RS plus various PD0325901 concentrations for 24 h. Chondrocytes were stained with Safranin O (left), and the GAG content in the chondrocytes was measured using the DMB method (right). B. Gene expression of matrix degradation enzymes and collagen synthesis genes in chondrocytes was evaluated by qPCR after treatment with and without various concentrations of the MEK-ERK1/2 pathway inhibitor PD0325901. Data are expressed as the mean ± SD from three separate experiments. # P

    Article Snippet: Chondrocytes were isolated from the articular cartilage of 24-hour-old Sprague-Dawley (SD) rats and dispersed in 0.1% collagenase type II (C6885, Sigma-Aldrich, Switzerland) for 3 h. Chondrocytes were collected and cultured in Dulbecco’s Modified Eagle’s Medium (DMEM, Biowest, France) supplemented with 10% FBS (Biowest, France) and 1% penicillin-streptomycin.

    Techniques: Expressing, Cell Culture, Staining, Real-time Polymerase Chain Reaction

    Release of NO from JS-K . (A) Generation of NO from JS-K in RPMI-1640 medium containing 10% fetal bovine serum was determined indirectly by photorimetric detection of nitrite according to Green et al. [ 31 ]. (B) Detection of nitrotyrosine by fluorescence microscopy in 22Rv1 cells: Generation of intracellular NO was detected indirectly by detection of nitrotyrosine (green). Cell nuclei were stained with DAPI. [A] untreated cells, [B] 2 μM JS-K neg , [C] 2 μM JS-K, [D] 200 μM DETA/NO.

    Journal: BMC Cancer

    Article Title: JS-K, a glutathione/glutathione S-transferase-activated nitric oxide releasing prodrug inhibits androgen receptor and WNT-signaling in prostate cancer cells

    doi: 10.1186/1471-2407-12-130

    Figure Lengend Snippet: Release of NO from JS-K . (A) Generation of NO from JS-K in RPMI-1640 medium containing 10% fetal bovine serum was determined indirectly by photorimetric detection of nitrite according to Green et al. [ 31 ]. (B) Detection of nitrotyrosine by fluorescence microscopy in 22Rv1 cells: Generation of intracellular NO was detected indirectly by detection of nitrotyrosine (green). Cell nuclei were stained with DAPI. [A] untreated cells, [B] 2 μM JS-K neg , [C] 2 μM JS-K, [D] 200 μM DETA/NO.

    Article Snippet: After transfection cells were grown in RPMI 1640 with 5% dextran charcoal treated steroid free, dextran charcoal treated FBS (FBSdcc; Biowest, Nuaillé, France) and treated with 5 nM DHT and different concentrations of JS-K. (2) WNT-signalling : Luciferase reporter plasmids pTopFlash (250 ng/well) and pFopFlash (250 ng/well) were mixed either with pbCAT expression vector (250 ng/well) or insert-free vector (250 ng/well). pRL-tk-luc (80 ng/well) was co-transfected to correct for transfection efficiency.

    Techniques: Fluorescence, Microscopy, Staining

    Crizotinib-induced apoptosis in Ba/F3 cells transformed by NPM-ALK. Ba/F3 cells were infected with an empty virus (-) and retrovirus expressing NPM-ALK or TEL-JAK2. (A) Whole cell lysates were immunoblotted with an anti-ALK antibody, anti-Flag antibody, anti-JAK2 antibody, anti-phospho-STAT3 antibody, anti-STAT3 antibody, anti-phospho-STAT5 antibody, anti-STAT5 antibody or anti-β-actin antibody. (B) Transduced Ba/F3 cells were incubated with RPMI containing 10% FBS in the absence of IL-3 for 2 days. Viable cell numbers at 24 hr and 48 hr were counted by the Trypan blue exclusion method. Values are the mean ± SD of three independent experiments. (C-F) Transduced Ba/F3 cells were incubated with RPMI containing 10% FBS in the presence of crizotinib (0.25, 0.5, 1 μM) and MMC (0.1, 1, 10 μg/mL) for 24 hr. (C) Cell viability was measured by the WST assay. Values are the mean ± S.D. of four independent experiments. (D) Cells were fixed, treated with propidium iodide, and subjected to a flow cytometric analysis. The percentages of cells in the sub-G1 phase were graphed. Values are the mean ± S.D. of four independent experiments. (E) To evaluate the apoptotic cell death, the chromatin DNA was isolated from cells and subjected to agarose gel electrophoresis. (F) Whole cell lysates were prepared and the phosphorylation of STAT3 and STAT5 was analyzed by immunoblotting. The relative phosphorylation level of STAT3 and STAT5 is shown in the graphs. Values are given as the mean ± SD of three independent experiments. *** P

    Journal: PLoS ONE

    Article Title: Alpha-tocopherol attenuates the anti-tumor activity of crizotinib against cells transformed by NPM-ALK

    doi: 10.1371/journal.pone.0183003

    Figure Lengend Snippet: Crizotinib-induced apoptosis in Ba/F3 cells transformed by NPM-ALK. Ba/F3 cells were infected with an empty virus (-) and retrovirus expressing NPM-ALK or TEL-JAK2. (A) Whole cell lysates were immunoblotted with an anti-ALK antibody, anti-Flag antibody, anti-JAK2 antibody, anti-phospho-STAT3 antibody, anti-STAT3 antibody, anti-phospho-STAT5 antibody, anti-STAT5 antibody or anti-β-actin antibody. (B) Transduced Ba/F3 cells were incubated with RPMI containing 10% FBS in the absence of IL-3 for 2 days. Viable cell numbers at 24 hr and 48 hr were counted by the Trypan blue exclusion method. Values are the mean ± SD of three independent experiments. (C-F) Transduced Ba/F3 cells were incubated with RPMI containing 10% FBS in the presence of crizotinib (0.25, 0.5, 1 μM) and MMC (0.1, 1, 10 μg/mL) for 24 hr. (C) Cell viability was measured by the WST assay. Values are the mean ± S.D. of four independent experiments. (D) Cells were fixed, treated with propidium iodide, and subjected to a flow cytometric analysis. The percentages of cells in the sub-G1 phase were graphed. Values are the mean ± S.D. of four independent experiments. (E) To evaluate the apoptotic cell death, the chromatin DNA was isolated from cells and subjected to agarose gel electrophoresis. (F) Whole cell lysates were prepared and the phosphorylation of STAT3 and STAT5 was analyzed by immunoblotting. The relative phosphorylation level of STAT3 and STAT5 is shown in the graphs. Values are given as the mean ± SD of three independent experiments. *** P

    Article Snippet: These cells were cultured in RPMI-1640 medium (Nacalai Tesque, Tokyo, Japan) supplemented with 10% fetal bovine serum (FBS) (BioWest, Nuaillé, France), 100 units/ml penicillin (Nacalai Tesque), 100 μg/ml streptomycin (Nacalai Tesque), 2 ng/mL IL-3 (PEPROTECH), and 5 μg/mL puromycin (InVivoGen).

    Techniques: Transformation Assay, Infection, Expressing, Incubation, WST Assay, Flow Cytometry, Isolation, Agarose Gel Electrophoresis