The human normal glial cell line HEB and glioma cell lines U251, U87, T98-G and A172 were purchased from the Cell Resource Center of Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. DMEM/HG, fetal bovine serum (FBS), Opti-MEM and 0.25% trypsin containing 0.02% EDTA were purchased from Gibco, MTT kit, 5-aza-2'-deoxycytosine (AZA) and histone deacetylase inhibitor (TSA) from Sigma, Trizol, reverse transcription kit from Thermo, SYBR Green Real-time PCR kit from Shanghai Solarbio Bioscience & Technology, QIAamp DNA Mini kit and EpiTect Bisul te kit from Qiagen, Lipofectamine 3000 from Invitrogen, and the cell cycle assay kit from BD. The pcDNA3.1-miR-133a mimic, scrambled miRNA (miR-NC) and the pcDNA3.1-PPARγ overexpression plasmid were obtained from Guangzhou Ruibo Bio. Antibodies against PPARγ, cyclin D1, cyclin D2, CDK4 and β-actin were from Abcam, and the horseradish peroxidase (HRP)-labeled IgG secondary antibody from Guangzhou Jingcai. PPARγ wild type (WT) and mutant (MUT) luciferase reporter plasmids were purchased from Shanghai Jima and the luciferase assay kit from Promega. The PCR primers were synthesized by Shanghai Shenggong. Other reagents were from our laboratory and of analytical grade.
Cell cycle assay kit
Manufactured by BD
Sourced in United States
The Cell Cycle Assay kit is a laboratory equipment product designed to analyze the distribution of cells in different phases of the cell cycle. It provides tools to measure and quantify the percentage of cells in the G0/G1, S, and G2/M phases.
Automatically generated - may contain errors
Lab products found in correlation
Reverse transcription kit, by Thermo Fisher Scientific (9 mentions)
Modfit software, by Verity Software House (4 mentions)
Trypsin, by Thermo Fisher Scientific (3 mentions)
Dmem hg, by Thermo Fisher Scientific (3 mentions)
Mtt kit, by Thermo Fisher Scientific (3 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (3 mentions)
Opti mem, by Thermo Fisher Scientific (3 mentions)
Qiaamp dna mini kit, by Qiagen (3 mentions)
Sybr green real time pcr kit, by Solarbio (3 mentions)
Modfit software version 3, by Verity Software House (3 mentions)
Luciferase assay kit, by Promega (2 mentions)
Epitect bisul te kit, by Qiagen (2 mentions)
5 aza 2 deoxycytosine aza and histone deacetylase inhibitor tsa, by Merck Group (2 mentions)
Trizol, by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Epitect bisulfite kit, by Qiagen (1 mentions)
Dual luciferase reporter assay kit, by Promega (1 mentions)
Pmirglo vector, by Promega (1 mentions)
Facsort, by BD (1 mentions)
Cellquest v5, by BD (1 mentions)
23 protocols using cell cycle assay kit
1
Molecular Mechanisms of miR-133a in Glioma Regulation
2
MicroRNA-133a Regulates Glioma Cell Proliferation
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The human normal glial cell line HEB and glioma cell lines U251, U87, T98-G and A172 were purchased from the Cell Resource Center of Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. DMEM/HG, fetal bovine serum (FBS), Opti-MEM and 0.25% trypsin containing 0.02% EDTA were purchased from Gibco, MTT kit, 5-Azacytidine (AZA) and histone deacetylase inhibitor (TSA) from Sigma, Trizol, reverse transcription kit from Thermo, SYBR Green Real-time PCR kit from Shanghai Solarbio Bioscience & Technology, QIAamp DNA Mini kit and EpiTect Bisulfite kit from Qiagen, Lipofectamine 3000 from Invitrogen, and the cell cycle assay kit from BD. The pcDNA3.1-miR-133a mimic, scrambled miRNA (miR-NC) and the pcDNA3.1-PPARG overexpression plasmid were obtained from Guangzhou Ruibo Bio. Antibodies against PPARG, cyclin D1, cyclin D2, CDK4 and 𝛽-actin were from Abcam, and the horseradish peroxidase (HRP)-labeled IgG secondary antibody from Guangzhou Jingcai. PPARG wild type (WT) and mutant (MUT) luciferase reporter plasmids were purchased from Shanghai Jima and the luciferase assay kit from Promega. The PCR primers were designed and synthesized by Shanghai Sangon Biotechnology. Other reagents were from our laboratory and of analytical grade.
3
Insights into Glioma Cell Regulation
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The human normal glial cell line HEB and glioma cell lines U251, U87, T98-G and A172 were purchased from the Cell Resource Center of Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. DMEM/HG, fetal bovine serum (FBS), Opti-MEM and 0.25% trypsin containing 0.02% EDTA were purchased from Gibco, MTT kit, 5-aza-2'-deoxycytosine (AZA) and histone deacetylase inhibitor (TSA) from Sigma, Trizol, reverse transcription kit from Thermo, SYBR Green Real-time PCR kit from Shanghai Solarbio Bioscience & Technology, QIAamp DNA Mini kit and EpiTect Bisul te kit from Qiagen, Lipofectamine 3000 from Invitrogen, and the cell cycle assay kit from BD. The pcDNA3.1-miR-133a-5p mimic, scrambled miRNA (miR-NC) and the pcDNA3.1 PGC-1α overexpression plasmid were obtained from Guangzhou Ruibo Bio. Antibodies against PGC-1α, cyclin D1, cyclin D2, Cdk4 and β-actin were from Abcam, and the horseradish peroxidase (HRP)-labeled IgG secondary antibody from Guangzhou Jingcai. QuickChange Site-Directed Mutagenesis kit was purchased from Stratagene. Dual-Luciferase Reporter Assay kit and pmirGLO vector were from Promega. The PCR primers were synthesized by Shanghai Shenggong. Other reagents were from our laboratory and of analytical grade.
4
Cell Cycle Analysis of Transfected Cells
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At 24 hrs after transfection, cells were collected. Cell Cycle Assay Kit (BD Biosciences, Franklin Lakes, NJ, USA) was used to analyze the cell cycle. Briefly, the cells were incubated with 200 μL liquid A for 10 mins, and 150 μL liquid B for another 10 mins. Then, the cells were incubated with 120 μL liquid C in dark for 10 mins before flow cytometry analysis on FACSort (BD Biosciences, Franklin Lakes, NJ, USA). The result was analyzed using ModFit software version 3.2 (Verity Software House, Topsham, ME, USA).
5
Flow Cytometry Analysis of NK Cell Cycle
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NK cells (1×105) were washed with PBS twice, and subjected to flow cytometry using the Cell Cycle Assay Kit (BD Biosciences, Franklin Lakes, NJ, USA) for the detection of cell cycles. Briefly, the cells were incubated with 200 μl liquid A at room temperature for 10 min, and then mixed with 150 μl liquid B before incubation at room temperature for 10 min. Then, 120 μl liquid C was added before incubation at room temperature in the dark for 10 min. Finally, the cells were used for flow cytometry and the results were analyzed using ModFit software (Verity Software House, Topsham, ME, USA).
6
Cell Cycle and Apoptosis Analysis
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At 24 h after transfection, cells (1x106) from each group were washed with pre-cooled PBS (2X) and subjected to flow cytometry using the Cell Cycle Assay kit (BD Biosciences) to detect the cell cycle distribution according to the manufacturer's instructions. The data were analyzed using ModFit software (v4.1; Verity Software House, Inc.).
After treatment with serum from healthy subjects or sepsis patients for 24 h, HUVECs (1x106) in each group were washed with pre-cooled PBS (2X) and subjected to flow cytometry using the ANXN V-FITC Apoptosis Detection kit I (BD Biosciences) to detect apoptosis according to the manufacturer's instructions. Annexin V-positive cells were early apoptotic, PI-positive cells were necrotic and double-positive cells were late apoptotic (CellQuest v5.1; BD Biosciences).
After treatment with serum from healthy subjects or sepsis patients for 24 h, HUVECs (1x106) in each group were washed with pre-cooled PBS (2X) and subjected to flow cytometry using the ANXN V-FITC Apoptosis Detection kit I (BD Biosciences) to detect apoptosis according to the manufacturer's instructions. Annexin V-positive cells were early apoptotic, PI-positive cells were necrotic and double-positive cells were late apoptotic (CellQuest v5.1; BD Biosciences).
7
Cell Cycle Analysis by Flow Cytometry
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At 48 h following transfection, cells were digested with trypsin and rinsed twice with pre-cold PBS. Cell cycle was detected by flow cytometry, using a Cell Cycle Assay kit (BD Biosciences, Franklin Lakes, NJ, USA), according to the manufacturer's instructions. Cells (2×106 cells/ml) were incubated with 200 µl solution A at room temperature for 10 min, and then incubated with 150 µl solution B at room temperature for 10 min, followed by incubation at room temperature with 120 µl solution C in the dark for 10 min. Fluorescence was detected using a flow cytometer, and the results were analyzed with Modfit software (version 3.2; Verity Software House, Inc., Topsham, ME, USA).
8
Apoptosis and Cell Cycle Analysis in HUVECs
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At 48 h after transfection, HUVECs were trypsinized, collected and washed twice with precooled phosphate-buffered saline. Cellular apoptosis was measured by flow cytometry using a FITC Annexin V apoptosis detection kit I (BD Biosciences, Franklin Lakes, NJ, USA), following the manufacturer's instructions. Early apoptotic cells were identified by single-positive staining with Annexin V, necrotic cells were identified by single-positive staining with propidium iodide and late apoptotic cells were identified by double-positive staining with propidium iodide and Annexin V. To determine the cell cycle, a cell cycle assay kit (BD Biosciences) was employed. Cells were incubated with 200 µl liquid A at room temperature for 10 min, followed by 150 µl liquid B at room temperature for 10 min and finally 120 µl liquid C at room temperature in the dark for 10 min. A flow cytometer was used for detection and the results were analyzed using ModFit v3.2 software (http://www.vsh.com ).
9
Endothelial Cell Proliferation and Cell Cycle
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The proliferation activity of the vascular endothelial cells was detected with the CCK-8 methods (Beyotime, Haimen, Jiangsu, China), according to the manufacturer’s instructions. Briefly, the vascular endothelial cells were planted onto the 96-well plate at the density of 2×103 cells/well. The 20 μl CCK-8 (5 g/L) was added into the wells at 0 h, 24 h, 48 h, and 72 h, respectively. On the final day, 150 μl CCK-8 reaction solution was added to incubate the cells at 37°C for 2 h. The absorbance at 490 nm was detected and the proliferation curves were plotted.
Cell cycle was detected with flow cytometry. Totally 1×106 HUVEC cells were collected from each group. After washing with PBS, the cell cycle was detected with the Cell Cycle Assay Kit (BD company). The cells were subsequently incubated with 200 μl solution A at room temperature for 10 min, 150 μl solution B at room temperature for 10 min, and then 120 μl solution C in the dark for 10 min. The detection data from flow cytometry were analyzed by Modfit software.
Cell cycle was detected with flow cytometry. Totally 1×106 HUVEC cells were collected from each group. After washing with PBS, the cell cycle was detected with the Cell Cycle Assay Kit (BD company). The cells were subsequently incubated with 200 μl solution A at room temperature for 10 min, 150 μl solution B at room temperature for 10 min, and then 120 μl solution C in the dark for 10 min. The detection data from flow cytometry were analyzed by Modfit software.
10
Cell Cycle Analysis by Flow Cytometry
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Cell cycle distribution was examined by flow cytometry using a cell cycle assay kit (BD Biosciences, San Jose, CA, USA) according to the manufacturer’s protocol. After propidium iodide (PI) staining, the cellular DNA content was used to determine cell cycle profile (G0/G1, G2/M and S phases), and the distribution of cell cycle was analyzed using Cell-FIT software (BD Biosciences, San Jose, CA, USA).
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