Annexin V Apoptosis Kit [FITC]

Flow Cytometry: Annexin V Apoptosis Kit [FITC] [NBP2-29373] - Controls run before the experimental samples are analyzed, to adjust the fluorescent compensation on the flow cytometer due to the FITC signal from the Annexin V-FITC bleeding into the Propidium Iodide signal.

Flow Cytometry: Annexin V Apoptosis Kit [FITC] [NBP2-29373] - Adherent RAW cells treated with actinomycin-D, detached with a cell detachment solution and stained with the Annexin V Apoptosis Kit [FITC].

• Reactivity: Hu, Mu, Rt
• Applications: Flow, Flow-CS, ICC/IF
• Conjugate: FITC

Annexin V Apoptosis Kit [FITC] Summary

• Description: Annexin V Apoptosis Kit [FITC] can identify apoptosis at an earlier stage than kits based on DNA fragmentation in the nucleus. However, it is like most assays and has limitations. Since Annexin V staining precedes the loss of membrane integrity which accompanies the later stages of cell death resulting from either apoptotic or necrotic processes, staining with Annexin V-FITC is typically used in conjunction with a live/dead dye such as propidium iodide (PI) or 7-Amino-Actinomycin (7-AAD) to allow the investigator to identify early apoptotic cells (PI negative, Annexin V-FITC positive) from dead cells (PI positive, AnnexinV-FITC positive). Viable cells with intact membranes exclude PI, whereas the membranes of dead and damaged cells are permeable to PI. For this reason, Annexin V staining has to be performed on live cells as opposed to assays which require para- formaldehyde/ethanol fixed cells.
  
  The Annexin V assay works in the following manner: Cells that are viable are both Annexin V-FITC and PI negative. While cells that are in early apoptosis are Annexin V-FITC positive and PI negative and cells that are in late apoptosis or already dead are both FITC 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 the dead cells will stain with both Annexin V-FITC and PI (see image). However, when apoptosis is measured over time, cells can be often tracked from Annexin V- FITC and PI negative (viable, or no measurable apoptosis), to Annexin V-FITC positive and PI negative (early apoptosis, membrane integrity is present) and finally to Annexin V- FITC 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 Annexin V-FITC and PI positive, in of itself, reveals less information about the process by which the cells underwent their demise. For this reason, it is a good idea to analyze samples from multiple time points.
• Kit Type: Apoptosis Kit
• Gene: ANXA5

Applications/Dilutions

• Application Notes: Use in Flow cell surface reported in scientific literature (PMID 23955790) Use the Control Cells provided to set up compensation and quadrants. The Control Cells are positive for both Annexin V-FITC and PI.
  The basal level of apoptosis and necrosis varies considerably within a give cell population. Even in the absence of induced apoptosis, most cell populations will contain a minor percentage of cells that are positive for apoptosis ((Annexin V-FITC positive, PI negative) and dead, necrotic, or in the late stages of apoptosis (Annexin V-FITC positive, PI positive). Thus an untreated cell population is used to define the basal level of apoptotic and dead cells.
  Determine the percentage of cells that have been induced to undergo apoptosis by subtracting the percentage of apoptotic cells in the untreated from the treated population. Since cell death is the eventual outcome of cells undergoing apoptosis, cells in the late stages of apoptosis will have a damaged membrane and stain positive for PI as well as for Annexin V-FITC.
  Consequently cells which have undergone necrosis are not distinguishable from those which have undergone apoptosis. Use in Immunocytochemistry/immunofluorescence reported in scientific literature (PMID 25947082). Use in FLOW cytometry reported in scientific literature ( PMID 27448441)

Reactivity Notes

Human reactivity reported in scientific literature (PMID: 23955790) Mouse reactivity reported in scientific literature (PMID: 22566271). Rat reactivity reported in scientific literature (PMID: 25947082)

Packaging, Storage & Formulations

• Storage: Store at 4C. Do not freeze.

Notes

ADDITIONAL ITEMS REQUIRED (NOT INCLUDED IN THE KIT)
Distilled H2O
Flow Cytometer
Experimental Cells
Centrifuge
Positive Control Cell Line (recommended, not required)

Alternate Names for Annexin V Apoptosis Kit [FITC]

• Anchorin CII
• annexin A5
• Annexin V
• annexin-5
• ANX5
• ANXA5
• ANXV
• Calphobindin I
• Endonexin II
• ENX2CBP-I
• Lipocortin V
• PAP-I
• Placental anticoagulant protein 4
• Placental anticoagulant protein I
• PP4
• Thromboplastin inhibitor
• VAC-alpha
• Vascular anticoagulant-alpha

Background

Apoptosis is the term that describes programmed cell death. It is believed to take place in the majority of animal and plant cells. It is a distinct event that triggers characteristic morphological and biological changes in the cellular life cycle. One of the early events that happens in the apoptotic pathway is the loss of plasma membrane asymmetry. 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 when labeled with a fluorescent tag, such as FITC, can be used with flow cytometry to measure this event. 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.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Kits are guaranteed for 6 months from date of receipt.

Publications for Annexin V Kit (NBP2-29373)(20)

Publications using NBP2-29373

• Hartung F, Pardo LA. Guiding TRAIL to cancer cells through Kv10.1 potassium channel overcomes resistance to doxorubicin Eur. Biophys. J. 2016 Jun 27 [PMID: 27350552] (ICC/IF, Human)
• Skivka LM, Fedorchuk OG, Bezdeneznykh NO. The effect of antineoplastic drug NSC631570 on immunogenicity of B16 melanoma. J Exp Integr Med 2014 Mar 24 (Flow-CS, Mouse)
  
  Details:
  Metastatic mlanoma cells, Fig 2.
• Song W, Wang F, Lotfi P et al. 2-Hydroxypropyl-B-Cyclodextrin Promotes TFEB-mediated Activation of Autophagy: Implications for Therapy. J. Biol. Chem. 2014 Feb 20 [PMID: 24558044] (Flow-CS, Human)
• Kumar A, Kant S, Singh SM. Targeting monocarboxylate transporter by alpha-cyano-4-hydroxycinnamate modulates apoptosis and cisplatin resistance of Colo205 cells: implication of altered cell survival regulation. Apoptosis 2013 Dec [PMID: 23955790] (Flow-CS, Human)
• Li FJ, Schreeder DM, Li R et al. FCRL3 promotes TLR9-induced B-cell activation and suppresses plasma cell differentiation. Eur J Immunol 2013 Nov [PMID: 23857366] (FLOW, Human)
  
  Details:
  Purified human blood B cells, Fig 2C.
• Badr CE, Niers JM, Morse D et al. Suicidal gene therapy in an NF-kB-controlled tumor environment as monitored by a secreted blood reporter. Gene Ther. 2011 May [PMID: 21150937] (Human)
  
  Details:
  Human glioma (Gli36) cells infected with lenti-NF-CU-IGluc and treated with TNF-alpha, 5-FC, or 5FC+TNF alpha, Fig 2D.
• Khurana S, Hollingsworth A, Piche M. Antiapoptotic Actions of Methyl Gallate on Neonatal Rat Cardiac Myocytes Exposed to H2O2. Oxidative Medicine and Cellular Longevity. 2014 Jan 12 (Rat)
  
  Details:
  Neonatal rat cardiac myocytes, Fig 3.
• Vishvakarma NK, Kumar A, Singh V et al. Hyperglycemia of tumor microenvironment modulates stage-dependent tumor progression and multidrug resistance: implication of cell survival regulatory molecules and altered glucose transport. Mol Carcinog 2013 Dec [PMID: 22566271] (Flow-CS, Mouse)
• Lu L, Wang S, Zheng L et al. Amyotrophic lateral sclerosis-linked mutant SOD1 sequesters Hu antigen R (HuR) and TIA-1-related protein (TIAR): implications for impaired post-transcriptional regulation of vascular endothelial growth factor. J Biol Chem. 2009 Dec 4 [PMID: 19805546]
• Narendrula R. rRNA Disruption: A Predictive Marker of Response to Taxane Chemotherapy. Thesis 2014 (Flow-CS, Human)
  
  Details:
  A2789 human ovarian carcinoma cell line, Fig 12.
• Basset C, Bonnet-Magnaval F, Navarro MG, Touriol C et al. Api5 a new cofactor of estrogen receptor alpha involved in breast cancer outcome Oncotarget 2017 Aug 08 [PMID: 28881748] (Flow, Human)
• Esposito CL, Nuzzo S, Kumar SA et al. A combined microRNA-based targeted therapeutic approach to eradicate glioblastoma stem-like cells J Control Release 2016 Jul 21 [PMID: 27448441] (FLOW, Human)
• Chen Y, Huang H, Yao C et al. Antitumor activity of combined endostatin and thymidine kinase gene therapy in C6 glioma models Cancer Med 2016 Jul 01 [PMID: 27366865]
• Huang CL, Chao CC, Lee YC et al. Paraquat Induces Cell Death Through Impairing Mitochondrial Membrane Permeability. Mol. Neurobiol. 2015 May 07 [PMID: 25947082] (ICC/IF, FLOW, Rat)
  
  Details:
  Annexin V-FITC-conjugated apoptosis kit was used for analyzing apoptotic cells in experiments involving PC12 cells. The cells with or without 24 hours of Paraquat/PQ treatments (+ dibucaine or cyclosporin A treatments) were subjected to Annexin V-FITC-staining followed by microscopgy imaging or flow cytometric analysis (Fig 1D).
• Xue Y, Wu L, Liu Y et al. ENTPD5 Induces Apoptosis in Lung Cancer Cells via Regulating Caspase 3 Expression PLoS ONE 2015 Mar 21 [PMID: 25794010] (FLOW, Human)
• Song W, Soo Lee S, Savini M et al. Ceria nanoparticles stabilized by organic surface coatings activate the lysosome-autophagy system and enhance autophagic clearance. ACS Nano. 2014 Oct 28 [PMID: 25315655] (FLOW, Human)
  
  Details:
  Annexin V - FITC Apoptosis Kit (Imgenex's CytoGLO Annexin V-FITC Apoptosis Detection Kit 10085K) used for detecting - Annexin V binding affinity change (%) in HeLa/TFEB cells treated or not with CeO2 nanoparticles or taxol for 24 h (Figure 3A).
• Wang F, Segatori L. Remodeling the proteostasis network to rescue glucocerebrosidase variants by inhibiting ER-associated degradation and enhancing ER folding. PLoS One. 2013 [PMID: 23620750] (FLOW, Human)
  
  Details:
  Annexin V - FITC Apoptosis Kit (Imgenex's CytoGLO Annexin V-FITC Apoptosis Detection Kit ) used for detecting - Annexin V binding in lacidipine and EerI treated GD cells, Gaucher?s disease /GD patient-derived fibroblasts homozygous for the L444P (1448T>C) mutation (GM10915). FLOW data is shown in Figure 3A.
• Wang F, Song W, Brancati G, Segatori L. Inhibition of endoplasmic reticulum-associated degradation rescues native folding in loss of function protein misfolding diseases. J Biol Chem. 2011 [PMID: 22006919] (FLOW, Human)
  
  Details:
  Annexin V - FITC Apoptosis Kit (Imgenex's CytoGLO Annexin V-FITC Apoptosis Detection Kit ) used for detecting apoptosis induction in L444P GC patient-derived fibroblasts treated or not with ER-associated degradation / ERAD inhibitors (FIGURE 8)
• Wang F, Chou A, Segatori L. Lacidipine remodels protein folding and Ca 2+ homeostasis in Gaucher's disease fibroblasts: a mechanism to rescue mutant glucocerebrosidase. Chem Biol. 2011 [PMID: 21700212] (FLOW, Human)
  
  Details:
  Annexin V - FITC Apoptosis Kit (Imgenex's CytoGLO Annexin V-FITC Apoptosis Detection Kit ) used for detecting Annexin V binding in experiments involving L444P Gaucher's disease patient-derived fibroblasts.
• Saydam O, Senol O, Wurdinger T et al. miRNA-7 attenuation in Schwannoma tumors stimulates growth by upregulating three oncogenic signaling pathways. Cancer Res. 2011 Feb 1 [PMID: 21156648] (Flow - CS, Human)
  
  Details:
  Flow (cell surface): Human primary Schwannoma and HEI-193 cell lines, S1.

Bioinformatics

• Gene Symbol: ANXA5