Annexin V — Purified

Apoptosis, the physiological process of cell death, is a naturally occurring phenomenon. It is normally a tightly regulated process; however, if deregulated, tumor growth can result. Apoptotic processes can be measured in the laboratory easily and most effectively by using a FITC-labeled Annexin V protein to stain cells and analyze them via flow cytometry. It is a useful method to screen and analyze cells in a high throughput manner, resulting in a clearer understanding of the cellular response to various stimuli.

When cells are undergoing cell death, phosphatidylserine (PS), a phospholipid which makes up a portion of the cell membrane, will translocate to the outside-facing side of the membrane. This translocation in the cell death mechanism exposes PS to the extracellular environment. Annexin V, a 35-36 kDa phospholipid-binding protein, has a high affinity for PS (KDa=7nmol/l). Once Annexin V is bound to the apoptotic cell it acts as a beacon for cells that have undergone apoptosis. Annexin V is often conjugated to FITC so the amount of signal can be measured and analyzed from labeled cells using a flow cytometer. The translocation of PS to the outside or exposed side of the membrane is an early event in the apoptotic process. Therefore, Annexin V staining is considered a marker for early stage apoptosis. Often times, Annexin V is paired with Propidium Iodide (PI) stain in order to analyze for late-apoptotic or dead cells. Early stage apoptotic cells will only take up the Annexin V stain but will remain PI negative, and late-stage apoptotic cells will be positive for both Annexin V and PI.

Application Guidance:

Custom Conjugation: Because this product is azide-free, tag-free, and lacks carrier proteins, it is highly optimized for conjugation with FITC, PE, APC, or custom near-IR dyes using standard amine-reactive chemistry.

Flow Cytometry / Imaging: Once conjugated, Annexin V is typically paired with a vital dye such as Propidium Iodide (PI) or 7-AAD.

Lyophilized

Lyophilized from a 0.22 µm filtered solution in 1x PBS (pH 7.4) containing 5% Trehalose (w/v) as a lyoprotectant.

Briefly centrifuge the vial at low speed before opening to ensure all lyophilized powder is at the bottom. Reconstitute using sterile-filtered Phosphate Buffered Saline (PBS) to your desired concentration (0.1 - 1 mg /ml mg/mL}. Mix gently by pipetting or rolling. Do not vortex vigorously.

Short-Term Storage: The formulation is stable for up to three (3) months when stored in the dark at 2°C to 8°C.

Long-Term Storage: For long-term storage of the lyophilized material, storage between -20°C and -35°C is recommended.

Next Day Blue Ice

1. Piazza, GA. et al. (2016) Oncotarget. 7(5):5353-65.PubMed
2. Gassman, NR. et al. (2018) Environ Toxicol. 33(3):333-342.PubMed
3. Gelles, JD. and Chipuk, JE. (2016) Cell Death Dis. 7(12): e2493.