MII oocytes were transferred to Annexin-V (Vazyme, Nanjing, China). The Annexin-V was diluted in the dilution ratio of 1 to 9 by the binding buffer. Oocytes were incubated in a 38.5 °C, 5% CO2 incubator for 30 min, then washed 3 times with DPBS + 0.3% PVP for 5 min each time. Observation was performed with a confocal microscope (Olympus, Tokyo, Japan). Each group consisted of 20~30 oocytes.
Annexin 5
Manufactured by Vazyme
Sourced in China
Annexin V is a protein that binds to phosphatidylserine, a phospholipid found on the surface of cells undergoing apoptosis (programmed cell death). It is commonly used in laboratory techniques to detect and quantify apoptotic cells.
Automatically generated - may contain errors
Lab products found in correlation
Pi apoptosis detection kit, by Vazyme (2 mentions)
Facscalibur, by BD (1 mentions)
Ab44976, by Abcam (1 mentions)
Ly294002, by Cell Signaling Technology (1 mentions)
P mtor ser2448, by Cell Signaling Technology (1 mentions)
Ab182858, by Abcam (1 mentions)
Ab191606, by Abcam (1 mentions)
Ab191866, by Abcam (1 mentions)
Ab32064, by Abcam (1 mentions)
Ab32503, by Abcam (1 mentions)
Beclin 1, by Cell Signaling Technology (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Ab81283, by Abcam (1 mentions)
Propidium iodide, by Vazyme (1 mentions)
Facsort flow cytometer, by Beckman Coulter (1 mentions)
Propidium iodide staining solution, by Vazyme (1 mentions)
7 protocols using annexin 5
1
Annexin-V Staining of Oocytes
2
Annexin V-FITC/PI Apoptosis Assay
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Annexin V-FITC/PI staining (CAT#: A211-01/02; Vazyme Biotech Co., Ltd., Nanjing, China) was used to evaluate A498 cell apoptosis. A498 cells underwent various treatments before being harvested and resuspended with 500 μL of binding buffer. Subsequently, the A498 cells were stained with 5 μL of Annexin V and 10 μL of PI (CAT#: A211-01/02; Vazyme Biotech Co., Ltd., Nanjing, China) for 30 min in the dark. Then, apoptotic cells were detected using a flow cytometer (BD FACSCalibur).
3
Isoliquiritigenin-Induced Apoptosis Pathway
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Isoliquiritigenin (ISL, purity ≥98%) was purchased from Chengdu Desite Chemical Company Limited, and a stock solution of 10 mM was dissolved in dimethyl sulfoxide (Sigma-Aldrich; Merck KGaA). Other reagents were purchased as follows: chloroquine (HCQ), 3-(4,5-dimetrylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) (Beyotime Biotechnology, Haimen, China), Annexin V, and PI apoptosis Detection Kit (Vazyme Biotech, China). Primary antibodies against Bcl-2 (1:2000, ab182858), Bax (1:5000, ab32503), Caspase-3 (1:500, ab44976), PARP (1:5000, ab32064), p-PI3K (1:1000, ab191606), Akt, P-Akt (Ser473), p-Akt (1:5000, ab81283), and secondary antibodies (1:5000, ab191866) were purchased from Abcam (UK). LC-3 (1:1000, #4108), P-mTOR (Ser2448) (1:1000, #5536), Beclin1 (1:1000, #4122), and LY294002 (#9901 S) were obtained from Cell Signaling Technology (Danvers, MA, USA).
4
Annexin V-based Apoptosis Assay
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Neutrophils from each group were collected and washed twice with cold PBS. Annexin V was added to the cells and gently vortexed. Annexin V (Vazyme Biotech, Jiangsu, China) is a Ca2+-dependent phospholipid-binding protein with high affinity for PS, and it binds to exposed apoptotic cell surface PS. Cells were incubated for 15 min at room temperature in the dark and analyzed using FC within 1 h.
5
Apoptosis Rate Determination by Flow Cytometry
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A total of 48 h after transfection, 5 × 105 GCs were collected and stained for 10 min using 5 μL Annexin V (Vazyme, Nanjing, China) and 5 μL Propidium Iodide (PI) (Vazyme, China), a d the apoptosis rate was detected by flow cytometry (Becton Dickinson, Franklin Lakes, USA) and analyzed using FlowJo v7.6 software.
6
Annexin V and PI Apoptosis Assay
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The B16F10 cells treated with PBS, Dox (1 μM) and Dox-coated hydrogel with or without R837 (10 μg/mL) were suspended in 1 × 105 cells/mL, and 5 μL Annexin V (Vazyme, Nanjing, China) and 5 μL propidium iodide staining solution (Vazyme, Nanjing, China) were added to 100 μL of the cell suspension. 400 μL binding buffer was next added to the cell suspension. After incubated at room temperature for 10min in the dark, the stained cells were assayed and quantified using a FACSort Flow Cytometer (Beckman Coulter, Brea, CA, USA). Cell debris was excluded from the analysis by an appropriate forward light scatter threshold setting. What's more, compensation was used wherever necessary.
7
Apoptosis Dynamics in A172 and U251 Cells
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The apoptosis capacity of A172 and U251 cells was tested with Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) Apoptosis Detection Kit (Vazyme) in line with the manufacturers’ instructions. The results were evaluated with flow cytometry (Beckman Coulter, Atlanta, GA, USA).
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