Annexin V Protein (Biotin)

Catalog No. ABIN2690962

Product Details

Target: Annexin V (ANXA5) (Annexin A5 (ANXA5))

Protein Type: Recombinant

Origin: Human

Source: Escherichia coli (E. coli)

Purification tag / Conjugate: This Annexin V protein is labelled with Biotin.

Application: Flow Cytometry (FACS)

Brand: BD Pharmingen™

Characteristics: Apoptosis is a normal physiologic process which occurs during embryonic development as well as in maintenence of tissue homeostasis. The apoptotic program is characterized by certain morphologic features, including loss of plasma membrane asymmetry and attachment, condensation of the cytoplasm and nucleus, and internucleosomal cleavage of DNA. Loss of plasma membrane is one of the earliest features. In apoptotic cells, the membrane phospholipid phosphatidylserine (PS) is translocated from the inner to the outer leaflet of the plasma membrane, thereby exposing PS to the external cellular environment. Annexin V is a 35-36 kDa Ca2+ dependent phospholipid-binding protein that has a high affinity for PS, and binds to cells with exposed PS. Annexin V may be conjugated to biotin. This format retains its high affinity for PS and thus serves as a sensitive probe for flow cytometric analysis of cells that are undergoing apoptosis. Since externalization of PS occurs in the earlier stages of apoptosis, Biotin Annexin V staining can identify apoptosis at an earlier stage than assays based on nuclear changes such as DNA fragmentation. Biotin Annexin V staining precedes the loss of membrane integrity which accompanies the latest stages of cell death resulting from either apoptotic or necrotic processes. Therefore, staining with Biotin Annexin V is typically used in conjunction with a vital dye such as propidium iodide (PI) or 7-Amino-Actinomycin (7-AAD) to allow the investigator to identify early apoptotic cells (PI negative, Biotin Annexin V positive). Viable cells with intact membranes exclude PI, whereas the membranes of dead and damaged cells are permeable to PI. For example, cells that are considered viable are both Biotin Annexin V and PI negative while cells that are in early apoptosis are Biotin Annexin V positive and PI negative, while cells that are in late apoptosis or already dead are both Biotin Annexin V and PI positive. This assay does not distinguish between cells that have undergone apoptotic death versus those that have died as a result of a necrotic pathway because in either case, the dead cells will stain with both Biotin Annexin V and PI. However, when apoptosis is measured over time, cells can be often tracked from Biotin Annexin V and PI negative (viable, or no measurable apoptosis), to Biotin Annexin V positive and PI negative (early apoptosis, membrane integrity is present) and finally to Biotin Annexin V and PI positive (end stage apoptosis and death). The movement of cells through these three stages suggests apoptosis. In contrast, a single observation indicating that cells are both Biotin Annexin V and PI positive, in of itself, reveals less information about the process by which the cells underwent their demise. Biotin Annexin V is routinely tested by flow cytometric analysis. Other applications were tested during antibody development only or reported in the literature.

Application Details

Comment:

BD Pharmingen™ Biotin Annexin V - Biotin - Clone NA

Sample Volume: 5 μL

Restrictions: For Research Use only

Handling

Buffer: Aqueous buffered solution containing BSA and ≤0.09 % sodium azide.

Preservative: Sodium azide

Precaution of Use: This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Storage: 4 °C

Storage Comment: Store undiluted at 4° C and protected from prolonged exposure to light. Do not freeze. Page 1 of 4556417 Rev. 10 Biotin Annexin V: A tool for identifying cells that are undergoing apoptosis. Jurkat T cells were left untreated (upper left & lower left panels), treated for 5 hours (upper middle & lower middle panels) or 12 hours (upper right & lower right panels) with anti-human Fas antibody (clone DX2, Cat. No. 555670) and Protein G. Cells were incubated with Biotin Annexin V, followed by incubation with SAv-FITC in a buffer containing Propidium Iodide (PI). Cells were then analyzed by flow cytometry. Untreated cells were primarily Biotin Annexin V and PI negative, indicating that they were viable and not undergoing apoptosis. After a 5 hr treatment with DX2, there were two populations of cells: cells undergoing apoptosis (Biotin Annexin V positive and PI negative), and cells that were viable and not undergoing apoptosis (Biotin Annexin V and PI negative). After a 12 hr treatment with DX2, three populations of cells were identified: cells that had already died or were in late stage of apoptosis (Biotin Annexin V and PI positive), cells undergoing apoptosis (Biotin Annexin V positive and PI negative), and cells that were viable and not undergoing apoptosis (Biotin Annexin V and PI negative). The addition of Protein G enhances the ability of DX2 to induce apoptosis, presumably by cross-linking the Fas receptor.

References

van Engeland, Ramaekers, Schutte, Reutelingsperger: "A novel assay to measure loss of plasma membrane asymmetry during apoptosis of adherent cells in culture." in: Cytometry, Vol. 24, Issue 2, pp. 131-9, (1996) (PubMed).

Martin, Reutelingsperger, McGahon, Rader, van Schie, LaFace, Green: "Early redistribution of plasma membrane phosphatidylserine is a general feature of apoptosis regardless of the initiating stimulus: inhibition by overexpression of Bcl-2 and Abl." in: The Journal of experimental medicine, Vol. 182, Issue 5, pp. 1545-56, (1995) (PubMed).

Homburg, de Haas, von dem Borne, Verhoeven, Reutelingsperger, Roos: "Human neutrophils lose their surface Fc gamma RIII and acquire Annexin V binding sites during apoptosis in vitro." in: Blood, Vol. 85, Issue 2, pp. 532-40, (1995) (PubMed).

Koopman, Reutelingsperger, Kuijten, Keehnen, Pals, van Oers: "Annexin V for flow cytometric detection of phosphatidylserine expression on B cells undergoing apoptosis." in: Blood, Vol. 84, Issue 5, pp. 1415-20, (1994) (PubMed).

Raynal, Pollard: "Annexins: the problem of assessing the biological role for a gene family of multifunctional calcium- and phospholipid-binding proteins." in: Biochimica et biophysica acta, Vol. 1197, Issue 1, pp. 63-93, (1994) (PubMed).

Andree, Reutelingsperger, Hauptmann, Hemker, Hermens, Willems: "Binding of vascular anticoagulant alpha (VAC alpha) to planar phospholipid bilayers." in: The Journal of biological chemistry, Vol. 265, Issue 9, pp. 4923-8, (1990) (PubMed).

Target Details

Target: Annexin V (ANXA5) (Annexin A5 (ANXA5))

Alternative Name: Annexin V (ANXA5 Products)

Synonyms: anx Protein, anx5 Protein, ANX V Protein, anxa5 Protein, cb989 Protein, wu:fa98f06 Protein, wu:fj10f10 Protein, MGC89158 Protein, ANX5 Protein, ENX2 Protein, PP4 Protein, RPRGL3 Protein, Anx5 Protein, R74653 Protein, LC5 Protein, enx2 Protein, annexin A5 Protein, annexin A5b Protein, Annexin A5 Protein, annexin A5 L homeolog Protein, ANXA5 Protein, anxa5b Protein, anxa5 Protein, Anxa5 Protein, anxa5.L Protein

Pathways: Apoptosis