Annexin A5 (ANXA5) protein (FITC)

Name: Annexin A5 (ANXA5)

Synonyms: PP4, ANX5, ENX2, Anx5, R74653, LC5, MGC93655, ANXA5, MGC126971, anx, anx5, ANX V, anxa5, cb989, wu:fa98f06, wu:fj10f10, MGC89158

Host: Please enquire

Reactivity: Please enquire

Conjugate: FITC

Application: Flow Cytometry (FACS), Fluorescence Microscopy (FM)

Catalog no.: ABIN412007

Additional Information

Description: Reagent for detecting early stages of apoptosis. During apoptosis, phosphatidylserine (PS) is translocated from the cytoplasmic face of the plasma membrane to the cell surface. Annexin V has a strong, Ca 2+ -dependent affinity for PS and therefore is used as a probe for detecting apoptosis. The Annexin V-FITC conjugate can be used for detection of apoptosis by fluorescence microscopy or by flow cytometry.

Application Details

Protocol: A. Incubation of cells with Annexin V-FITC: 1. Induce apoptosis by desired methods. 2. Collect 1 x 10 5 cells by centrifugation. 3. Resuspend cells in 500 μ l of 1X Annexin V Binding Buffer (Cat.#: 1035-100). 4. Add 1 μ l of Annexin V-FITC and 1 μ l Propidium Iodide (PI, Cat.#: 1056-1). 5. Incubate at room temperature for 5 min in the dark. Proceed to B or C below depending on method of analysis. B. Quantification by Flow Cytometry: Analyze cells by flow cytometry (Ex. = 488 nm, Em. = 530 nm) using FL1 channel for detecting Annexin V-FITC staining and FL2 channel for detecting PI staining. For adherent cells, trypsinize and gently wash cells with serum-containing medium before incubation with Annexin V-FITC (A.3-5). C. Detection by Fluorescence Microscopy: 1. Place the cell suspension from Step A.5 on a glass slide, and cover with a glass coverslip. For analyzing adherent cells, grow cells directly on a coverslip. Following incubation (A.5), invert coverslip on a glass slide and visualize cells. The cells can also be washed with 1X Annexin V Binding Buffer and fixed in 2% formaldehyde before visualization. (Cells must be incubated with Annexin V-FITC before fixation because any cell membrane disruption can cause nonspecific binding of annexin V to PS on the inner surface of the cell membrane.) 2. Observe the cells under a fluorescence microscope using a dual filter set for FITC and rhodamine, or separate filters. Cells that have bound Annexin V-FITC will show green staining on the plasma membrane. Cells that have lost membrane integrity will show red PI staining throughout the nuclei and a halo of green staining (FITC) on the plasma membrane. TESTING RESULTS: Jurkat cells (treated with 2 μ M camptothecin for 6 hours) were collected for annexin V assay according to the kit instructions. Results show 40-60% apoptotic cells as analyzed by flow cytometry.

Buffer: 0.15 mg/ml in PBS containing 1% BSA, 0.02% sodium azide.

Storage: Store at 4 °C. Do not freeze. Stable for 1 year.

Restrictions: For Research Use only

Publications

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Grad, Bahlis, Reis et al.: "Ascorbic acid enhances arsenic trioxide-induced cytotoxicity in multiple myeloma cells." in: Blood, Vol. 98, Issue 3, pp. 805-13, 2001 (PubMed).

Oshiro, Landowski, Catlett-Falcone et al.: "Inhibition of JAK kinase activity enhances Fas-mediated apoptosis but reduces cytotoxic activity of topoisomerase II inhibitors in U266 myeloma cells." in: Clinical cancer research : an official journal of the American Association for Cancer Research, Vol. 7, Issue 12, pp. 4262-71, 2001 (PubMed).

Lin, LeCouter, Kowalski et al.: "Characterization of endocrine gland-derived vascular endothelial growth factor signaling in adrenal cortex capillary endothelial cells." in: The Journal of biological chemistry, Vol. 277, Issue 10, pp. 8724-9, 2002 (PubMed).

Breckenridge, Nguyen, Kuppig et al.: "The procaspase-8 isoform, procaspase-8L, recruited to the BAP31 complex at the endoplasmic reticulum." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 99, Issue 7, pp. 4331-6, 2002 (PubMed).

Dalton: "Targeting the mitochondria: an exciting new approach to myeloma therapy. Commentary re: N. J. Bahlis et al., Feasibility and correlates of arsenic trioxide combined with ascorbic acid-mediated depletion of intracellular glutathione for the treatment of re" in: Clinical cancer research : an official journal of the American Association for Cancer Research, Vol. 8, Issue 12, pp. 3643-5, 2002 (PubMed).

Guzey, Kitada, Reed: "Apoptosis induction by 1alpha,25-dihydroxyvitamin D3 in prostate cancer." in: Molecular cancer therapeutics, Vol. 1, Issue 9, pp. 667-77, 2002 (PubMed).

Alonso, Tamasdan, Miller et al.: "Flavopiridol induces apoptosis in glioma cell lines independent of retinoblastoma and p53 tumor suppressor pathway alterations by a caspase-independent pathway." in: Molecular cancer therapeutics, Vol. 2, Issue 2, pp. 139-50, 2003 (PubMed).

LeCouter, Lin, Tejada et al.: "The endocrine-gland-derived VEGF homologue Bv8 promotes angiogenesis in the testis: Localization of Bv8 receptors to endothelial cells." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 100, Issue 5, pp. 2685-90, 2003 (PubMed).

Korfali, Ruchaud, Loegering et al.: "Caspase-7 gene disruption reveals an involvement of the enzyme during the early stages of apoptosis." in: The Journal of biological chemistry, Vol. 279, Issue 2, pp. 1030-9, 2004 (PubMed).

Tyurina, Serinkan, Tyurin et al.: "Lipid antioxidant, etoposide, inhibits phosphatidylserine externalization and macrophage clearance of apoptotic cells by preventing phosphatidylserine oxidation." in: The Journal of biological chemistry, Vol. 279, Issue 7, pp. 6056-64, 2004 (PubMed).

Beaupre, Cepero, Obeng et al.: "R115777 induces Ras-independent apoptosis of myeloma cells via multiple intrinsic pathways." in: Molecular cancer therapeutics, Vol. 3, Issue 2, pp. 179-86, 2004 (PubMed).

Chen, Van der Sluis, Boulware et al.: "The FA/BRCA pathway is involved in melphalan-induced DNA interstrand cross-link repair and accounts for melphalan resistance in multiple myeloma cells." in: Blood, Vol. 106, Issue 2, pp. 698-705, 2005 (PubMed).

Okamoto, Kawamura, Kanbe et al.: "Suppression of serum IgE response and systemic anaphylaxis in a food allergy model by orally administered high-dose TGF-beta." in: International immunology, Vol. 17, Issue 6, pp. 705-12, 2005 (PubMed).

Obeng, Boise: "Caspase-12 and caspase-4 are not required for caspase-dependent endoplasmic reticulum stress-induced apoptosis." in: The Journal of biological chemistry, Vol. 280, Issue 33, pp. 29578-87, 2005 (PubMed).

Terashima, Kojima, Fujimiya et al.: "The fusion of bone-marrow-derived proinsulin-expressing cells with nerve cells underlies diabetic neuropathy." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 102, Issue 35, pp. 12525-30, 2005 (PubMed).

Kono, Dunn, Durda et al.: "Role of the mitogen-activated protein kinase signaling pathway in the regulation of human melanocytic antigen expression." in: Molecular cancer research : MCR, Vol. 4, Issue 10, pp. 779-92, 2006 (PubMed).

Chung, Kim, Lee et al.: "Ghrelin inhibits apoptosis in hypothalamic neuronal cells during oxygen-glucose deprivation." in: Endocrinology, Vol. 148, Issue 1, pp. 148-59, 2006 (PubMed).

Del Gaizo Moore, Brown, Certo et al.: "Chronic lymphocytic leukemia requires BCL2 to sequester prodeath BIM, explaining sensitivity to BCL2 antagonist ABT-737." in: The Journal of clinical investigation, Vol. 117, Issue 1, pp. 112-21, 2007 (PubMed).

Vempati, Diaz, Barrientos et al.: "Role of cytochrome C in apoptosis: increased sensitivity to tumor necrosis factor alpha is associated with respiratory defects but not with lack of cytochrome C release." in: Molecular and cellular biology, Vol. 27, Issue 5, pp. 1771-83, 2007 (PubMed).

Hirai, Nelson, Yamazaki et al.: "Non-classical export of epimorphin and its adhesion to alphav-integrin in regulation of epithelial morphogenesis." in: Journal of cell science, Vol. 120, Issue Pt 12, pp. 2032-43, 2007 (PubMed).

Mancao, Hammerschmidt: "Epstein-Barr virus latent membrane protein 2A is a B-cell receptor mimic and essential for B-cell survival." in: Blood, Vol. 110, Issue 10, pp. 3715-21, 2007 (PubMed).

Del Gaizo Moore, Schlis, Sallan et al.: "BCL-2 dependence and ABT-737 sensitivity in acute lymphoblastic leukemia." in: Blood, Vol. 111, Issue 4, pp. 2300-9, 2008 (PubMed).

Chen, Xie, Kuhn et al.: "Targeting the p27 E3 ligase SCF(Skp2) results in p27- and Skp2-mediated cell-cycle arrest and activation of autophagy." in: Blood, Vol. 111, Issue 9, pp. 4690-9, 2008 (PubMed).

Zhang, Wang, Liu et al.: "Regulation of tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis by DJ-1 in thyroid cancer cells." in: Endocrine-related cancer, Vol. 15, Issue 2, pp. 535-44, 2008 (PubMed).

Sumi, Ishida, Okumura et al.: "Activation of tumour-necrosis factor-related apoptosis-inducing ligand receptor enhances the severity of murine allergic conjunctivitis." in: The British journal of ophthalmology, Vol. 93, Issue 1, pp. 110-5, 2008 (PubMed).

Kim, Shu, Xu et al.: "Chemical biology investigation of cell death pathways activated by endoplasmic reticulum stress reveals cytoprotective modulators of ASK1." in: The Journal of biological chemistry, Vol. 284, Issue 3, pp. 1593-603, 2009 (PubMed).

Kuhn, Hunsucker, Chen et al.: "Targeted inhibition of the immunoproteasome is a potent strategy against models of multiple myeloma that overcomes resistance to conventional drugs and nonspecific proteasome inhibitors." in: Blood, Vol. 113, Issue 19, pp. 4667-76, 2009 (PubMed).

Faber, Krivtsov, Stubbs et al.: "HOXA9 is required for survival in human MLL-rearranged acute leukemias." in: Blood, Vol. 113, Issue 11, pp. 2375-85, 2009 (PubMed).

Tu, Ren, Wang et al.: "The p53-cathepsin axis cooperates with ROS to activate programmed necrotic death upon DNA damage." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 106, Issue 4, pp. 1093-8, 2009 (PubMed).

Radke, Reynolds, Melo et al.: "Mature human eosinophils express functional Notch ligands mediating eosinophil autocrine regulation." in: Blood, Vol. 113, Issue 13, pp. 3092-101, 2009 (PubMed).

Krenning, Dankers, Drouven et al.: "Endothelial progenitor cell dysfunction in patients with progressive chronic kidney disease." in: American journal of physiology. Renal physiology, Vol. 296, Issue 6, pp. F1314-22, 2009 (PubMed).

Matulis, Morales, Yehiayan et al.: "Darinaparsin induces a unique cellular response and is active in an arsenic trioxide-resistant myeloma cell line." in: Molecular cancer therapeutics, Vol. 8, Issue 5, pp. 1197-206, 2011 (PubMed).

Snyder, Reynolds, Hollingsworth et al.: "Clara cells attenuate the inflammatory response through regulation of macrophage behavior." in: American journal of respiratory cell and molecular biology, Vol. 42, Issue 2, pp. 161-71, 2010 (PubMed).

Anguissola, Köhler, OByrne et al.: "Bid and calpains cooperate to trigger oxaliplatin-induced apoptosis of cervical carcinoma HeLa cells." in: Molecular pharmacology, Vol. 76, Issue 5, pp. 998-1010, 2009 (PubMed).

Chen, Wu, Hung et al.: "Unique epitopes on C epsilon mX in IgE-B cell receptors are potentially applicable for targeting IgE-committed B cells." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 184, Issue 4, pp. 1748-56, 2010 (PubMed).

Tsujii, Eguchi, Chenchik et al.: "Screening of cell death genes with a mammalian genome-wide RNAi library." in: Journal of biochemistry, Vol. 148, Issue 2, pp. 157-70, 2010 (PubMed).

Wu, Zhou, Robertson et al.: "Identification of a bone marrow-derived CD8αα+ dendritic cell-like population in inflamed autoimmune target tissue with capability of inducing T cell apoptosis." in: Journal of leukocyte biology, Vol. 88, Issue 5, pp. 849-61, 2010 (PubMed).

Sharmeen, Skrtic, Sukhai et al.: "The antiparasitic agent ivermectin induces chloride-dependent membrane hyperpolarization and cell death in leukemia cells." in: Blood, Vol. 116, Issue 18, pp. 3593-603, 2010 (PubMed).

Gao, Liu, Du et al.: "Implication of oxygen-regulated protein 150 (ORP150) in apoptosis induced by proteasome inhibitors in human thyroid cancer cells." in: The Journal of clinical endocrinology and metabolism, Vol. 95, Issue 11, pp. E319-26, 2010 (PubMed).

Kilbride, Farrelly, Bonner et al.: "AMP-activated protein kinase mediates apoptosis in response to bioenergetic stress through activation of the pro-apoptotic Bcl-2 homology domain-3-only protein BMF." in: The Journal of biological chemistry, Vol. 285, Issue 46, pp. 36199-206, 2010 (PubMed).

Herrmann, Kortylewski, Kujawski et al.: "Targeting Stat3 in the myeloid compartment drastically improves the in vivo antitumor functions of adoptively transferred T cells." in: Cancer research, Vol. 70, Issue 19, pp. 7455-64, 2010 (PubMed).

Gao, Wei, Lai et al.: "Nucleolar follistatin promotes cancer cell survival under glucose-deprived conditions through inhibiting cellular rRNA synthesis." in: The Journal of biological chemistry, Vol. 285, Issue 47, pp. 36857-64, 2010 (PubMed).

Robbins, Tersey, Ogihara et al.: "Inhibition of deoxyhypusine synthase enhances islet {beta} cell function and survival in the setting of endoplasmic reticulum stress and type 2 diabetes." in: The Journal of biological chemistry, Vol. 285, Issue 51, pp. 39943-52, 2010 (PubMed).

Sumitomo, Nakata, Higashino et al.: "Streptolysin S contributes to group A streptococcal translocation across an epithelial barrier." in: The Journal of biological chemistry, Vol. 286, Issue 4, pp. 2750-61, 2011 (PubMed).

Mao, Hou, Cao et al.: "The tricyclic antidepressant amitriptyline inhibits D-cyclin transactivation and induces myeloma cell apoptosis by inhibiting histone deacetylases: in vitro and in silico evidence." in: Molecular pharmacology, Vol. 79, Issue 4, pp. 672-80, 2011 (PubMed).

Du, Yan, Zhang et al.: "Proteasome inhibition induces a p38 MAPK pathway-dependent antiapoptotic program via Nrf2 in thyroid cancer cells." in: The Journal of clinical endocrinology and metabolism, Vol. 96, Issue 5, pp. E763-71, 2011 (PubMed).

Chen, Overcash, Green et al.: "The tumor suppressor activity of the transmembrane protein with epidermal growth factor and two follistatin motifs 2 (TMEFF2) correlates with its ability to modulate sarcosine levels." in: The Journal of biological chemistry, Vol. 286, Issue 18, pp. 16091-100, 2011 (PubMed).

Neri, Ren, Azab et al.: "Integrin β7-mediated regulation of multiple myeloma cell adhesion, migration, and invasion." in: Blood, Vol. 117, Issue 23, pp. 6202-13, 2011 (PubMed).

Peters, Raghavan: "Endoplasmic reticulum calcium depletion impacts chaperone secretion, innate immunity, and phagocytic uptake of cells." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 187, Issue 2, pp. 919-31, 2011 (PubMed).

Neri, Ren, Gratton et al.: "Bortezomib-induced "BRCAness" sensitizes multiple myeloma cells to PARP inhibitors." in: Blood, Vol. 118, Issue 24, pp. 6368-79, 2011 (PubMed).

Dondeti, Wubbenhorst, Lal et al.: "Integrative genomic analyses of sporadic clear cell renal cell carcinoma define disease subtypes and potential new therapeutic targets." in: Cancer research, Vol. 72, Issue 1, pp. 112-21, 2012 (PubMed).

E, Cao, Bhattacharya et al.: "Endogenous vascular endothelial growth factor-A (VEGF-A) maintains endothelial cell homeostasis by regulating VEGF receptor-2 transcription." in: The Journal of biological chemistry, Vol. 287, Issue 5, pp. 3029-41, 2012 (PubMed).