Annexin V/PI double-staining analysis was used to detect cell apoptosis. The cells were seeded in triplicate in 6-well plates, cultured overnight and pretreated with 0 or 100 μM I3A for 1 h before 0, 4 or 8 Gy irradiation. After 48 h, the cells were stained with FITC-conjugated Annexin V and PI (KeyGen, Nanjing, China). Apoptosis analysis was performed using a FACSVerse flow cytometer (BD Biosciences, San Diego, CA, USA).
Fitc conjugated annexin 5 and pi
Manufactured by Keygen Biotech
Sourced in China
FITC-conjugated Annexin V and PI is a labeling reagent used in flow cytometric analysis of apoptosis. Annexin V binds to phosphatidylserine, which is exposed on the surface of apoptotic cells. FITC (fluorescein isothiocyanate) is conjugated to Annexin V to allow detection by flow cytometry. Propidium iodide (PI) is included to stain necrotic cells.
Automatically generated - may contain errors
Lab products found in correlation
Facsverse flow cytometer, by BD (1 mentions)
Cellquest software version 5, by BD (1 mentions)
Cell cycle staining kit, by MultiSciences Biotech (1 mentions)
Facscalibur 2 sorter, by BD (1 mentions)
Cell quest research software, by BD (1 mentions)
Rnase a, by Merck Group (1 mentions)
Facs calibar, by BD (1 mentions)
Annexin 5 fitc flow cytometry assay, by BD (1 mentions)
In situ cell death detection kit, by Keygen Biotech (1 mentions)
4 protocols using fitc conjugated annexin 5 and pi
1
Apoptosis Analysis by Annexin V/PI
2
Evaluating CDCA7L siRNA Impact on Glioma Cell Cycle and Apoptosis
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The U87 cells were infected with CDCA7L siRNA or NC siRNA and incubated at 37°C. The transfected glioma cells were then harvested, washed twice with phosphate-buffered saline (PBS), and fixed with 75% ice-cold ethanol. Subsequently, the cells were examined using the Cell Cycle Staining kit (Multi Sciences) and incubated for 30 min in dark accord 37°C according to the manufacturer's instructions. In addition, to detect the effect of CDCA7L siRNA on cell apoptosis, the U87 cells were harvested by trypsinization and incubated with FITC-conjugated Annexin V and PI following the manufacturer's instructions (Keygen Biotech). Finally, the cells were analyzed by flow cytometry and BD CellQuest™ software version 5.1 (BD Biosciences).
3
Apoptosis and Cell Cycle Analysis
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Annexin V-FITC/propidium iodide (PI) double labeling was used to determine the cell cycle distribution or detect apoptosis. U251 cells were infected with the shRNA-CHPF or NC plasmids and incubated at 37˚C. U251 cells were harvested by trypsinization and incubated with FITC-conjugated Annexin V and PI according to the manufacturer's instructions (KeyGen Biotech, Nanjing, China). FACSCalibur II sorter and Cell Quest Research Software (BD Biosciences, San Diego, CA, USA) were used for analysis. To further confirm the changes of DNA content distribution in the shRNA-CHPFtreated cells, we performed PI staining. The cells of different groups were collected and fixed in 70% ethanol at 4̊C overnight. After incubation with 50 µg/ml of PI and RNase A (Sigma-Aldrich) for 30 min at 37̊C in the dark, the cells were subjected to FACS analysis as described above. All experiments were performed in triplicate.
Statistical analysis. All statistical analyses were implemented in the SPSS 20.0 statistical software package. All data are presented as the means ± standard deviations (SD) and the experiments were repeated 3 times. The Student's t-test was used for raw data analysis. A P-value of <0.05 was considered to indicate a statistically significant result.
Statistical analysis. All statistical analyses were implemented in the SPSS 20.0 statistical software package. All data are presented as the means ± standard deviations (SD) and the experiments were repeated 3 times. The Student's t-test was used for raw data analysis. A P-value of <0.05 was considered to indicate a statistically significant result.
4
Assessing Apoptosis with Flow Cytometry and TUNEL
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Propidium iodide (PI) and Annexin V-FITC-flow cytometry assay (BD Pharmingen, CA) was used to detect the apoptosis in the cells. Cells were stained with FITC-conjugated annexin V and PI (KeyGEN BioTECH, China, KGA107) according to the manufacturer’s instructions. After staining, the cells were analyzed by flow cytometry (FACS Calibar; Becton-Dickinson) using Cell Quest software.
TUNEL assay was processed using the in situ cell death detection kit (KeyGEN BioTECH, China, KGA7071). Hepatic tissues were stained according to the manufacturer’s instructions. The apoptosis index was captured using a fluorescence microscope (Olympus IX73, Japan).
TUNEL assay was processed using the in situ cell death detection kit (KeyGEN BioTECH, China, KGA7071). Hepatic tissues were stained according to the manufacturer’s instructions. The apoptosis index was captured using a fluorescence microscope (Olympus IX73, Japan).
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