FITC Annexin V Apoptosis Detection Kit with PI

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Other Names
Annexin A5 Apoptosis Detection Kit
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FITC_Annexin-V_kit_052318
Human T leukemia cell line Jurkat, treated (left) or non-treated (right) with BioLegend’s anti-human CD95 (EOS9.1) mAb (Cat. No. 305704) for 4 hours at 37°C, then stained with Annexin V- FITC for 15 minutes at 37°C in Annexin V Binding buffer. Propidium Iodide (PI) (cat. 421301 at 0.03 µg/Test) was added 5 minutes prior to running tubes.
  • FITC_Annexin-V_kit_052318
    Human T leukemia cell line Jurkat, treated (left) or non-treated (right) with BioLegend’s anti-human CD95 (EOS9.1) mAb (Cat. No. 305704) for 4 hours at 37°C, then stained with Annexin V- FITC for 15 minutes at 37°C in Annexin V Binding buffer. Propidium Iodide (PI) (cat. 421301 at 0.03 µg/Test) was added 5 minutes prior to running tubes.
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640914 100 tests 172€
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Description

BioLegend's FITC Annexin V Apoptosis Detection Kit with PI has been specifically designed for the identification of apoptotic and necrotic cells.

Annexin V (or Annexin A5) is a member of the annexin family of intracellular proteins that binds to phosphatidylserine (PS) in a calcium-dependent manner. PS is normally only found on the intracellular leaflet of the plasma membrane in healthy cells, but during early apoptosis, membrane asymmetry is lost and PS translocates to the external leaflet. Fluorochrome-labeled Annexin V can then be used to specifically target and identify apoptotic cells. Annexin V Binding Buffer is recommended for use with Annexin V staining. Annexin V binding alone cannot differentiate between apoptotic and necrotic cells. To help distinguish between the necrotic and apoptotic cells we recommend use of our Propidium Iodide Solution (PI). Early apoptotic cells will exclude PI, while late stage apoptotic cells and necrotic cells will stain positively, due to the passage of these dyes into the nucleus where they bind to DNA.

Propidium iodide is a fluorescent dye that binds to DNA. When excited by 488 nm laser light, it can be detected with in the PE/Texas Red® channel with a bandpass filter 610/10. It is commonly used in evaluation of cell viability or DNA content in cell cycle analysis by flow cytometry.
 

Product Details
Technical data sheet

Product Details

Reactivity
All mammalian species
Concentration
Lot-specific (please contact technical support for concentration and total µg amount, or use our Lookup tool if you have a lot number.)
Storage & Handling
Store between 2°C and 8°C. Do not freeze.

Caution: Propidium Iodide Solution is toxigenic and mutagenic; handle with care.
Application

FC - Quality tested

Recommended Usage

Staining Procedure:
1. Wash cells twice with cold BioLegend's Cell Staining Buffer, and then resuspend cells in Annexin V Binding Buffer at a concentration of 0.25-1.0 x 107 cells/ml.
2. Transfer 100 µl of cell suspension in a 5 ml test tube.
3. Add 5 µl of FITC Annexin V.
4. Add 10 µl of Propidium Iodide Solution.
5. Gently vortex the cells and incubate for 15 min at room temperature (25°C) in the dark.
6. Add 400 µl of Annexin V Binding Buffer to each tube. Analyze by flow cytometry with proper machine settings.

Application Notes

Materials Provided:
0.5 ml of FITC Annexin V
1 ml of Propidium Iodide Solution
50 ml of Annexin V Binding Buffer

Materials Not Included:
Cell Staining Buffer (Cat. No. 420201)

Application References

(PubMed link indicates BioLegend citation)
  1. Ranhanathan P, et al. 2013. J AM Soc Nephrol. PubMed
  2. Yu J, et al. 2014. PloS One. 9:114650. PubMed
  3. Wang Y, et al. 2014. Biochem Biophys Res Commun. 456:656. PubMed
  4. Jaishy B, et al. 2015. J Lipid Res. 56:546. PubMed
Product Citations
  1. Yu J, et al. 2014. PLoS One. 9:114650. PubMed
  2. Wang Y, et al. 2014. Biochem Biophys Res Commun. 456:656. PubMed
  3. Jaishy B, et al. 2015. J Lipid Res. 56:546. PubMed
  4. Yuan L, et al. 2015. Endocr Relat Cancer. 22: 577 - 591. PubMed
  5. Blue E, et al. 2015. Diabetes. 64: 2664 - 2675. PubMed
  6. Liu J, et al. 2015. Biochem Biophys Res Commun. 463: Pags 262-267. PubMed
  7. Levy G, et al. 2015. PLoS One. 10: 0136070. PubMed
  8. Belo H, et al. 2015. PLoS One. 10: e0139740. PubMed
  9. Ma H, et al. 2015. Sci Rep. 5: 15121. PubMed
  10. Weiss R, et al. 2015. Innate Immunity. 21: 813 - 826. PubMed
  11. Kan L, et al. 2015. PLoS One. 10: 0144513. PubMed
  12. Kongkavitoon P, et al. 2016. PLoS One. 11: 0146696. PubMed
  13. Chang T, et al. 2016. Sci Rep. 6:23403. PubMed
  14. Sareddy G, et al. 2016. Sci Rep. 6: 24185. PubMed
  15. Cao W, et al. 2016. Nat Commun. 7:11687. PubMed
  16. Saboor-Maleki S, et al. 2016. Technol Cancer Res Treat. 10.1177/1533034616650119. PubMed
  17. Mahmoud A, et al. 2016. Int J Toxicol. 35: 429 - 437. PubMed
  18. Ho J, et al. 2016. PLoS Biol. 14:e2000117. PubMed
  19. Balachandran C, et al. 2017. Biomed Pharmacother. 10.1016/j.biopha.2017.06.096. PubMed
  20. Guo Q, et al. 2018. Front Immunol. 9:1197. PubMed
  21. Yao J, et al. 2018. Onco Targets Ther. 11:3793. PubMed
  22. Shao M, et al. 2018. Evid Based Complement Alternat Med. 2018:6049498. PubMed
  23. Sohrabi Y, et al. 2019. Front Immunol. 9:3155. PubMed
  24. Ryva B, et al. 2019. Front Oncol. 9:100. PubMed
  25. Flegkas A, et al. 2019. Medicines (Basel). 6:1. PubMed
  26. Virumbrales–Muñ oz M, et al. 2019. Sci Rep. 9:6199. PubMed
  27. Tatsuno K, et al. 2019. Cell Death Dis. 10:578. PubMed
  28. Soonthornchai W, et al. 2019. Asian Pac J Allergy Immunol. N/A. PubMed
  29. Ali DM, et al. 2019. Cell Death Discov. 5:128. PubMed
  30. Coelho CC, et al. 2019. Sci Rep. 9:11050. PubMed
  31. Korencak M, et al. 2019. JCI Insight. 4. PubMed
  32. Ouyang J, et al. 2019. J Surg Res. 233:221. PubMed
  33. Fei X, et al. 2019. Cancer Manag Res. 11:4781. PubMed
  34. Allur Subramaniyan S, et al. 2018. Cell Commun Adhes. 24:19. PubMed
Publication Library

Antigen Details

Biology Area
Apoptosis/Tumor Suppressors/Cell Death, Cell Biology, Neuroscience
Gene ID
308 View all products for this Gene ID

Related FAQs

Can I freeze Annexin V conjugates?

It should not be frozen as it will lead to loss of biological activity due to dimerization.

Can I use RPMI during Annexin V staining?

It is best to follow protocol as described on the product data sheet. Moreover, RPMI 1640 has a relatively high concentration of phosphate and low calcium ion concentration, which negatively impacts Annexin binding to its target phosphatidylserine (PS). Measurement of cell death by using Annexin V may also be significantly affected by time of incubation on ice, calcium concentration, and type of medium.

How does pH and staining temperature affect Annexin V-Phosphatidylserine binding?

Annexin-Phosphatidylserine binding is lost below pH 5.2 and with prolonged incubation over a temperature of 42°C.

How is your Annexin made and what sequence does it cover?

It is made in E. coli, covering human aa Met1-Asp320.

Is Annexin V suitable for conjugation with the Maxpar® kit for CyTOF®?

Maxpar® Labeling kits require the protein to be partially reduced, so the metal chelate can be introduced through an SH group in the hinge region of the reduced antibody. Human Annexin V contains only one Cysteine which was reported to be chemically inactive. Thus, the Maxpar® labeling protocol would not work with Annexin V, unless a free –SH group can be introduced to Annexin V.  For more information regarding SH-mediated conjugation of Annexin V please consult published papers such as this one.

Why do I need to use Annexin V Binding Buffer?

Annexin V binding requires the presence of calcium in the solution.  So, we provide Annexin V Binding Buffer (cat # 422201), which is optimized for the best performance of Annexin V staining.

Go To Top Version: 2    Revision Date: 05.23.2018

For research use only. Not for diagnostic use. Not for resale. BioLegend will not be held responsible for patent infringement or other violations that may occur with the use of our products.

 

*These products may be covered by one or more Limited Use Label Licenses (see the BioLegend Catalog or our website, www.biolegend.com/ordering#license). BioLegend products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products, reverse engineer functionally similar materials, or to provide a service to third parties without written approval of BioLegend. By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses. Unless otherwise indicated, these products are for research use only and are not intended for human or animal diagnostic, therapeutic or commercial use.

 

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Toll-Free Phone: 1-877-Bio-Legend (246-5343) Phone: (858) 768-5800 Fax: (877) 455-9587

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