An Annexin V-APC/7-AAD apoptosis kit (KeyGEN Bio-TECH, China) for flow cytometry was used to detect the early apoptosis and necrosis of cells. Cells were pretreated with 32 μM Fer-1 for 24 h, and then 15 μM of ATO was added and the cells were treated for another 24 h. Next, the cells were harvested, washed twice with cold PBS, and incubated with 5 μL of Annexin V-APC and 5 μL of 7-AAD in 500 μL binding buffer for 10 min in the dark at room temperature. A flow cytometer (BD Biosciences, USA) was used to detect the cellular fluorescence.
Annexin 5 apc 7 aad apoptosis kit
Manufactured by Keygen Biotech
Sourced in China
The Annexin V-APC/7-AAD apoptosis kit is a laboratory reagent used for the detection and quantification of apoptotic cells. It utilizes Annexin V, a protein that binds to phosphatidylserine, and 7-AAD, a dye that stains DNA, to identify cells undergoing various stages of the apoptotic process.
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Lab products found in correlation
Cck 8 kit, by Keygen Biotech (1 mentions)
Fc500 flow cytometer, by Beckman Coulter (1 mentions)
Sdf 1α, by Abcam (1 mentions)
Rpmi 1640 culture medium, by Thermo Fisher Scientific (1 mentions)
Reverse transcription kit, by Takara Bio (1 mentions)
Rna extraction kit, by Takara Bio (1 mentions)
Tunel kit, by Keygen Biotech (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Vcam 1, by Abcam (1 mentions)
One step tb green primescript rt pcr kit 2, by Takara Bio (1 mentions)
P mtor, by Abcam (1 mentions)
Facsverse flow cytometer, by BD (1 mentions)
Modfit 3, by Verity Software House (1 mentions)
Propidium iodide, by Merck Group (1 mentions)
Rnase a, by Roche (1 mentions)
Canto 2 flow cytometer, by BD (1 mentions)
Facscalibur cytometer, by BD (1 mentions)
8 protocols using annexin 5 apc 7 aad apoptosis kit
1
Assessing Apoptosis and Necrosis
2
Cell Viability, Apoptosis, and Cell Cycle Analysis
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Cell viability was measured with a CCK8 kit (KeyGEN BioTECH, China). Briefly, 1 × 104 indicated cells were seeded in 96-well plates and incubated with D347-2761 at different concentrations for a certain time. Then cells were incubated with 5 μl CCK8 for another 2 h, and the absorbance was measured at 450 nm. Apoptotic cells were stained by Annexin V-APC/7-AAD apoptosis kit (KeyGEN BioTECH, China) and subsequently quantified by flow cytometer (BD Biosciences). For cell cycle assay, cells treated by D347-2761 were permeabilized and treated by RNase A, then cells were stained by PI and analyzed by flow cytometry.
3
Investigating CXCR4-Mediated Signaling in Hematological Malignancies
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Cim was purchased from MilliporeSigma, and RPMI 1640 culture medium and fetal bovine serum from Gibco; Thermo Fisher Scientific, Inc. EdU kit, TUNEL kit and Annexin V-APC/7-AAD apoptosis kit were purchased from Jiangsu KeyGEN Biotech Co., Ltd. RNA extraction kit, reverse transcription kit, One Step TB Green PrimeScript RT-PCR kit II (SYBR Green) were purchased from Takara Bio, Inc. Primers were synthesized by Sangon Biotech (Shanghai) Co., Ltd. Rabbit anti-human GAPDH was purchased from Jiangsu KeyGEN Biotech Co., Ltd., and rabbit anti-human C-X-C chemokine receptor type 4 (CXCR4), stromal cell-derived factor-1α (SDF-1α), vascular cell adhesion molecule 1 (VCAM1), leukocyte function-associated antigen-1 (LFA-1), Fms like tyrosine kinase receptor 3 (FLT3), nucleophosmin 1 (NPM-1), CCAAT/enhancer-binding protein alpha (C/EBPA), AKT, phosphorylated (p-)AKT, mTOR and p-mTOR were purchased from Abcam; very-late-antigen-4 (VLA-4) was purchased from ProteinTech Group, Inc. Flow cytometer FC500 was from Beckman Coulter, Inc., ND2000 ultra-microspectrophotometer was from Thermo Fisher Scientific, Inc., and gel image analysis system was produced by GeneGenius. BL8040 (CXCR4 antagonist) was from Roche Diagnostics.
4
Annexin V-APC/7-AAD Apoptosis Assay
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Annexin V-APC/7-AAD Apoptosis Kit (Keygen) was used to detect the apoptosis of cell lines quantitatively according to manufacturer’s instructions. Cells were incubated with anti-Annexin V-APC and 7-AAD for 10 min in the dark. FACS Calibur flowcytometer was used for detection of apoptotic cells. Annexin V-positive and PI-negative cells were considered to be apoptotic cells.
5
Cell Cycle and Apoptosis Analysis
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The stably transfected SW620 and RKO cells were trypsinized (without EDTA) and washed with PBS. For cell cycle analysis, 105 cells/100 µl cells were fixed with 70% ethanol at 4°C for 12 h and resuspended in staining buffer containing 450 µl propidium iodide (PI) and 50 µl RNaseA in the dark for 30 min at room temperature (cat. no. KGA512; Nanjing KeyGen Biotech Co., Ltd.) For apoptosis analysis, the Annexin V APC/7AAD apoptosis kit (cat. no. KGF004; Nanjing KeyGen Biotech Co., Ltd.) was used to stain for early and late apoptotic cells according to the manufacturers' instructions. Subsequently, stained cells were analyzed using a BD FACS-Verse flow cytometer (BD Biosciences). Cell cycle distribution was obtained using ModFit 3.2 software (Verity Software House, Inc.), and the apoptosis rate was calculated using FlowJo 7.6.1 software (FlowJo LLC).
6
Evaluating miR-34a's Impact on Cell Cycle and Apoptosis in MDR-MCF-7 Cells
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After transfection with miR-34a and NC-RNA mimics for 48 h, MDR-MCF-7 breast cancer cells were gently scraped, washed twice with chilled PBS (4°C), and suspended at a concentration of l × 106 cells/mL for flow cytometry detection. For cell cycle analysis, target cells were fixed in 75% ethanol and stained with propidium iodide (Sigma Aldrich) supplemented with RNase A (Roche). The Annexin V-APC/7-AAD apoptosis kit (KeyGEN Biotech, Nanjing, China) was used for apoptosis assays. Each set of two wells was analyzed 3 times with a BD Canto II flow cytometer (BD Biosciences, San Jose, CA, USA). Data were analyzed using CellQuest software.
7
Annexin V-APC/7-AAD Apoptosis Assay
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Apoptosis analysis was performed using the Annexin V-APC/7-AAD apoptosis kit (KeyGEN Biotech, Nanjing, China) according to the manufacturer’s instructions. Cells were analyzed on a FACS Calibur cytometer (BD, USA). Data were analyzed using CellQuest software.
8
Integrin α6 Modulates Cell Apoptosis
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The effect on cell apoptosis of integrin a6 knockdown or overexpression was evaluated using flow cytometry. Briefly, cell apoptosis was analyzed using an Annexin V-APC/7-AAD apoptosis kit (KeyGen Biotech) according to the manufacturer’s instructions.
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