Elabscience® Annexin V-FITC/7-AAD Cell Apoptosis Detection kit is developed to identify apoptotic and necrotic cells.
Annexin V is a member of the annexin family, which binds to phosphatidylserine (PS) in a calcium-dependent manner. Annexin V-FITC, the FITC-conjugated format, binds specifically to the PS on the outer leaflet of apoptotic cell membrane and can be detected with flow cytometry or fluorescence microscopy.
7-AAD (7-amino-actinomycin D) has a high DNA binding constant and is efficiently excluded by intact cells. It is useful for DNA analysis and dead cell discrimination during flow cytometric analysis. Due to the loss of integrity of membrane, 7-AAD can enter late apoptotic or necrotic cells to stain DNA. Cells at different apoptotic stages can be distinguished by using Annexin V and 7-AAD.
Jurkat cells were treated with 2 μM Camptothecin and detected with this kit.
Jurkat cells were cultured with (Right) or without (Left) 2 μM Camptothecin for 4 h. Annexin V-FITC single-positive cells were early apoptotic cells, Annexin V-FITC and 7-AAD double-positive cells were necrotic or late apoptotic cells, and 7-AAD single-positive cells were naked nuclei.
The Annexin V Binding Buffer (10 ×)[E-CK-A151] is a 10 × concentrated solution. Dilute with DI water to 1 × working solution before use.
For example: Take 1 mL Annexin V Binding Buffer (10 ×), dilute with DI water to 10 mL.
Annexin V-FITC and 7-AAD Viability Staining Solution should be stored in dark.
7-AAD Viability Staining Solution should be split into small tubes and stored at -20°C.
Other reagents should be stored at 2~8°C.
1. For maximal assay performance, this kit should be used within 12 months. Avoid freeze / thaw cycles.
2. For FCM analysis, please set untreated cells stained with both Annexin V-FITC and 7-AAD as negative control. As for compensation controls, please use drug-treated cells stained with either Annexin V-FITC or 7-AAD.
3. Annexin V-FITC can be detected in FITC channel while 7-AAD can be detected in PerCP/Cy5.5 channel.
4. Detect apoptosis as soon as possible after staining to avoid the increase number of apoptosis or necrosis.
5. Avoid extended exposure of the samples to direct light to protect the fluorophores from quenching.
6. For your safety and health, please wear the lab coat and disposable gloves before the experiments.