Twenty-four hours after transfection of the miR-152 mimic or inhibitor, bovine myoblasts were processed using a cell cycle testing kit (Multisciences, Hangzhou, China), and flow cytometry was used to analyze the cell cycle. The detailed operation process was performed according to the manufacturer’s instructions.
Cell cycle testing kit
Manufactured by MultiSciences Biotech
Sourced in China
The Cell Cycle Testing Kit is a laboratory tool designed to analyze the cell cycle progression of various cell types. This kit provides the essential components and protocols to assess the distribution of cells in different phases of the cell cycle, including G1, S, G2, and M phases. The kit utilizes standardized techniques to enable reliable and reproducible cell cycle analysis.
Automatically generated - may contain errors
Lab products found in correlation
Facscanto 2, by BD (13 mentions)
Annexin 5 pe 7 aad apoptosis detection kit, by RiboBio (3 mentions)
Facscalibur, by BD (2 mentions)
Mir 378a 3p mimic, by GenePharma (1 mentions)
Cell light edu dna cell proliferation kit, by RiboBio (1 mentions)
Dm5000b, by Leica (1 mentions)
Annexin 5 fitc, by MultiSciences Biotech (1 mentions)
Accuri c6, by BD (1 mentions)
Accuri c6 flow cytometer, by BD (1 mentions)
20 protocols using cell cycle testing kit
1
Cell Cycle Analysis of Bovine Myoblasts
2
Investigating circLMO7 Role in Cell Cycle
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To gain insights into the effects of circLMO7 in different periods of cell cycle, we analyzed the cell cycle of different treatment groups ((1) miR-378a-3p mimic (2 μg/ml; Genepharma, Shanghai, China), (2) pcDNA-circLMO7 (2 μg/ml), or (3) miR-378a-3p mimic+pcDNA-circLMO7) using the Cell Cycle Testing Kit (Multisciences, Hangzhou, China). We collected cells that had been cultivated in six-well plates and centrifuged them at 800 g/min for 5 min. The supernatant was discarded, and the cells were washed once with cold phosphate buffered saline (PBS, pH=7.4). We resuspended the cells in 1 ml of kit reagent A and 10 μl of reagent B, followed by vortexing for 10 s and incubation for 30 min at room temperature, after which the cell suspension was used for flow cytometry (FACS CantoTM II, BD BioSciences, USA).
3
Cell Cycle Analysis of GMEC
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Cell cycle analysis was performed using Cell Cycle Testing Kit (Multisciences, China). The GMEC were grown in 6-well plates (1 × 106 cells/well). The GMEC were treated with indicated concentration of SB-3CT for 24 h, and subsequently the cells were harvested and centrifuged at 800 g/min for 5 min. The supernatant was discarded, and the cells were washed once by cold PBS. GMEC were resuspended using 1 mL of reagent A and 10 μL of reagent B, subsequently blended by vortexing for 10 s and incubated for 30 min, and then analysis was done using a flow cytometry (FACS CantoTM II, BD BioSciences, USA).
4
Measuring DNA Synthesis and Cell Cycle
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After the transfection of PBMs with the expression vectors, inhibitor and siRNAs, we employed the EdU proliferation assay to measure their influences on DNA synthesis using the Cell Light EdU DNA cell proliferation kit according to the instructions (RiboBio, Guangzhou, China). The stained cells were detected and calculated by fluorescence microscopy (DM5000B, Leica Microsystems). Cell cycle phases were assessed by a cell cycle testing kit (Multisciences, Hangzhou, China) on a flow cytometry instrument (FACS Canto II, BD Biosciences, USA). Briefly, the cells were seeded in 6-well plates and transfected for 24 h after the cells reached 60% confluence. Cold 70% ethanol was used to fix the harvested cells. After staining with 100 μg/mL of the PI master mix at 37 °C for 30 min, the cell suspension was subjected to flow cytometry.
5
Cell Cycle Analysis via Flow Cytometry
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Cell cycle was analyzed by using a cell cycle testing kit (Multisciences, Hangzhou, China). The bovine myoblasts were treated with 1 mL of DNA staining solution and 10 μL of permeabilization solution in dark conditions for 30 min, then detected using a FACSCalibur flow ytometer (Becton Dickinson, USA). For the detailed procedure, refer to the manufacturer’s instructions.
6
Cell Cycle Analysis by Flow Cytometry
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The cell cycle was assessed using a Cell Cycle Testing Kit (Multisciences). ICP1 cells that had been cultivated in six‐well plates were harvested and centrifuged at 800 g for 5 min. The supernatant was discarded, and the cells were washed once with cold PBS. The cells were resuspended in 1 mL of kit reagent A and 10 μL of reagent B, followed by vortexing for 10 s and incubation for 30 min at room temperature, after which the cell suspension was used for flow cytometry (FACS Canto™ II; BD Biosciences, San Jose, CA, USA).
7
Cell Cycle Analysis of Transfected Myoblasts
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Myoblasts were transfected with siRNAs, mimics, and plasmids, and the cell cycle was assessed using the cell cycle testing kit (Multisciences, Hangzhou, China). After transfecting for 24 h, cells were harvested and washed with cold phosphate buffered saline (PBS). Then, the staining solution and permeabilization reagent were added to the cells. After incubating for 30 min, the cell cycle was analyzed by flow cytometry (FACS Canto II, BD Biosciences, USA).
8
Cell Cycle and Apoptosis Analysis
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The cell cycle of different treatment groups was measured using the cell cycle testing kit (Multisciences, Hangzhou, China). After incubation, the cell suspension obtained was used for flow cytometry (FACS Canto II, BD BioSciences, USA), and each treatment group had three independent replicates. In a flow cytometer, the cell apoptosis of MuSCs was also measured by flow cytometry using FITC-labeled Annexin V and propidium iodide (PI).
9
Cell Cycle Analysis by Flow Cytometry
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The cell cycle was assessed using a Cell Cycle Testing Kit (Multisciences, Hangzhou, China). When the density reached 5 × 106, ICP2 cells were collected and centrifuged at 800 rpm for 5 min. After being washed once with cold PBS, the cells were resuspended in 500 μL of propidium iodide staining solution and incubated at room temperature for 30 min in the dark. Then, the cell suspension was used for flow cytometry detection (Becton, Dickinson and Company, Franklin Lake, NJ, USA).
10
Cell Cycle and Apoptosis Analysis
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We analyzed the cell cycle of different treatment groups using the cell cycle testing kit (Multisciences, Hangzhou, China). Myoblasts were grown in six-well plates (1 × 106 cells/well) with 2 mL culture medium. After treatment for 24 hr, we harvested cells and washed them in PBS buffer. Then, 1 mL of DNA strain solution and 10 μL of permeabilization solution were added to the resuspended cells. After incubating for 30 min in the dark at room temperature, the cell suspension was used for flow cytometry (FACS Canto II, BD BioSciences, USA), and each treatment group had three independent replicates.
Cell apoptosis was measured by annexin V-FITC/PI staining assay. After incubation, cells with different treatment were washed three times with PBS buffer and then resuspended in 500 μL 1× binding buffer. Then, cells were incubated for 10 min in the dark at room temperature in the presence of annexin V-FITC (5 μL) and PI (10 μL; Multisciences, Hangzhou, China). Afterward, cells were analyzed using flow cytometry, and each treatment group had three independent replicates. Cells stained with only annexin V were evaluated as being in early apoptosis; cells stained with both annexin V and PI were evaluated as being in late apoptosis stage.
Cell apoptosis was measured by annexin V-FITC/PI staining assay. After incubation, cells with different treatment were washed three times with PBS buffer and then resuspended in 500 μL 1× binding buffer. Then, cells were incubated for 10 min in the dark at room temperature in the presence of annexin V-FITC (5 μL) and PI (10 μL; Multisciences, Hangzhou, China). Afterward, cells were analyzed using flow cytometry, and each treatment group had three independent replicates. Cells stained with only annexin V were evaluated as being in early apoptosis; cells stained with both annexin V and PI were evaluated as being in late apoptosis stage.
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