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PE Annexin V

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Regulatory Status
RUO
Other Names
Annexin A5
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Product Citations
publications
PE_Annexin-V_kit_082412
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- PE for 15 minutes at 37°C in Annexin V Binding buffer. Helix NP Green (Cat. No. 425303 at 1.25 nM) added 5 minutes at room temperature prior to running tubes.
  • PE_Annexin-V_kit_082412
    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- PE for 15 minutes at 37°C in Annexin V Binding buffer. Helix NP Green (Cat. No. 425303 at 1.25 nM) added 5 minutes at room temperature prior to running tubes.
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Cat # Size Price Quantity Check Availability Save
640907 25 tests 79€
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640908 100 tests 181€
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640947 300 tests 356€
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Description

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 (Cat. # 422201) is recommended for use with Annexin V staining. Annexin V binding alone cannot differentiate between apoptotic cells and necrotic. Therefore, we recommend using our Helix NP™ Blue (Cat. No. 425305), Helix NP™ Green (Cat. No. 425303) or Helix NP™ NIR (Cat. No. 425301). Early apoptotic cells will exclude 7-AAD and 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.

Product Details
Technical Data Sheet (pdf)

Product Details

Verified Reactivity
Human, Mouse, Rat
Reported Reactivity
Other Species
Formulation
Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide and BSA (origin USA)
Preparation
The purified protein was conjugated with PE under optimal conditions.
Concentration
Lot-specific (to obtain lot-specific concentration and expiration, please enter the lot number in our Certificate of Analysis online tool.)
Storage & Handling
The Annexin V solution should be stored undiluted between 2°C and 8°C, and protected from prolonged exposure to light. Do not freeze.
Application

FC - Quality tested
Recommended Usage

Each lot of this product is quality control tested by immunofluorescent staining with flow cytometric analysis. For flow cytometric staining, the suggested use of this reagent is 5 µl per 100,000 - million cells in a 100 µl volume of Annexin V Binding Buffer (Cat No. 422201). It is recommended that the reagent be titrated for optimal performance for each application.

Excitation Laser
Blue Laser (488 nm)
Green Laser (532 nm)/Yellow-Green Laser (561 nm)
Application Notes

Annexin V Staining

  1. Wash cells twice with cold BioLegend Cell Staining Buffer (Cat. No. 420201) and then resuspend cells in Annexin V Binding Buffer (Cat. No. 422201) at a concentration of 1x106 cells/mL.
  2. Transfer 100 µL of cell suspension in 5 mL test tube.
  3. Add 5 µL of fluorochrome conjugated Annexin V.
  4. Stain with a viability dye, such as PI (Cat. No. 421301), 7-AAD (Cat. Nos. 420403 & 420404), or Helix NP dyes (Cat. Nos. 425301, 425303, & 425305), if desired.
  5. Gently vortex the cells and incubate for 15 min at RT (25°C) in the dark.
  6. Add 400* µL of Annexin V Binding Buffer (Cat. No. 422201) to each tube. *For more concentrated samples, add a minimum of 200 µl of Annexin V Binding Buffer in this step.
  7. Analyze by flow cytometry.

For a better experience detecting apoptosis, we now recommend Apotracker™. Cell staining with Apotracker™ is Calcium independent. Thus, no special buffers are required, and the protocol can be shortened for single-step co-staining with other reagents.

Application References

(PubMed link indicates BioLegend citation)
  1. Koopman G, et al. 1994. Blood 84:1415.
  2. Vermes I, et al. 1995. J. Immunol. Methods 184:39.
  3. Dachary-Prigent J, et al. 1993. Blood 81:2554.
  4. Sekine C, et al. 2009. Int Immunol. PubMed
  5. Grujic M, et al. 2010. J. Immunol. 185:1730. PubMed
Product Citations
  1. Roblek M, et al. 2022. Front Oncol. 12:777634. PubMed
  2. Desikan SA, et al. 2022. STAR Protoc. 3:101388. PubMed
  3. Li DD, et al. 2022. J Cell Mol Med. 26:2438. PubMed
  4. Chen Q, et al. 2022. Cell Rep. 39:110990. PubMed
  5. Hao J, et al. 2022. Cell Rep. 41:111804. PubMed
  6. Kulkarni PP, et al. 2023. FASEB J. 37:e22768. PubMed
  7. Lam W, et al. 2023. Front Pharmacol. 13:1095186. PubMed
  8. de Oliveira TD, et al. 2023. Front Med (Lausanne). 9:1059028. PubMed
  9. Yen M, et al. 2022. Cell. 185:1414. PubMed
  10. Wenes M, et al. 2022. Cell Metab. 34:731. PubMed
  11. Knelson EH, et al. 2022. Cancer Immunol Res. 10:947. PubMed
  12. Manolakou T, et al. 2022. Sci Adv. 8:eabo5840. PubMed
  13. Luan H, et al. 2023. Molecules. 28:. PubMed
  14. Donadei C, et al. 2023. J Clin Med. 12:. PubMed
  15. Lan Y, et al. 2023. iScience. 26:106817. PubMed
  16. Symeonidou V, et al. 2021. Cell Rep. 37:109900. PubMed
  17. Chiu Y, et al. 2017. Nat Commun. 8:14324. PubMed
  18. Kulkarni PP, et al. 2021. Haematologica. . PubMed
  19. Liu Y, et al. 2022. Cancer Discov. :. PubMed
  20. Hinterbrandner M, et al. 2021. JCI Insight. 6:e151797. PubMed
  21. Carriere PP, et al. 2018. Int J Oncol. 53:1442. PubMed
  22. Liu J, et al. 2021. Sci Adv. 7: . PubMed
  23. Herman KD, et al. 2022. Front Immunol. 13:956991. PubMed
  24. Bartleson JM, et al. 2020. Nat Immunol. 1384:21. PubMed
  25. MacDonald A, et al. 2021. Front Immunol. 12:755995. PubMed
  26. Lei L, et al. 2021. J Cell Mol Med. . PubMed
  27. Paterson N, et al. 2022. Elife. 11:. PubMed
  28. Fuhrmann F et al. 2016. eLife. 5 pii: e20616. PubMed
  29. Blümel E, et al. 2020. Oncoimmunology. 9:1751561. PubMed
  30. Grinnemo KH, et al. 2019. Stem Cell Res Ther. 0.674305556. PubMed
  31. Krauss PL, et al. 2021. Front Immunol. 12:730672. PubMed
  32. Blümel E, et al. 2019. Oncoimmunology. 8:e1641387. PubMed
  33. Malouf C, et al. 2021. Blood. 138:2066. PubMed
  34. Jakic B, et al. 2021. Cell Death Dis. 12:187. PubMed
  35. Sun H, et al. 2017. J Autoimmun. 10.1016/j.jaut.2017.08.004. PubMed
  36. Hills LB, et al. 2021. J Immunol. 206:89. PubMed
  37. Verma N, et al. 2020. Sci Adv. 6:00. PubMed
  38. Ryan P, et al. 2016. Proc Natl Acad Sci U S A. 113(50):14378-14383. PubMed
  39. Olson WJ, et al. 2019. Cell Rep. 28:2878. PubMed
  40. Roy AG, et al. 2021. Int J Mol Sci. 22:. PubMed
  41. Zhang X, et al. 2021. Front Pharmacol. 12:629513. PubMed
  42. Kamali K, et al. 2021. Front Immunol. 12:647900. PubMed
  43. Lobastova L, et al. 2021. Frontiers in Cell and Developmental Biology. 9:698503. PubMed
  44. Jeong S, et al. 2021. Cell Metab. 33:145. PubMed
  45. Lei L, et al. 2021. Front Cell Dev Biol. 659744:9. PubMed
  46. Mesnieres M, et al. 2021. Cell Reports. 36(8):109618. PubMed
  47. Cochran AM, et al. 2021. Front Cell Dev Biol. 9:698639. PubMed
  48. Ruan Y, et al. 2022. Exp Ther Med. 23:47. PubMed
  49. Stevens M, et al. 2016. Nat Biotechnol. 34:1161-1167. PubMed
  50. Dregalla RC, et al. 2021. J Orthop Res. 39:2744. PubMed
  51. Ruffo E, et al. 2016. Sci Transl Med. 8: 321ra7. PubMed
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  53. Ye Z, et al. 2022. Front Oncol. 12:1018154. PubMed
  54. Guan H, et al. 2015. Oncogenesis. 4: 166. PubMed
RRID
not an antibody (BioLegend Cat. No. 640907)
AB_2561298 (BioLegend Cat. No. 640908)
not an antibody (BioLegend Cat. No. 640947)

Antigen Details

Biology Area
Apoptosis/Tumor Suppressors/Cell Death, Cell Biology, Neuroscience
Gene ID
308 View all products for this Gene ID
UniProt
View information about Annexin V on UniProt.org

Related FAQs

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

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

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.

What type of PE do you use in your conjugates?
We use R-PE in our conjugates.
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.

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.

Can I freeze Annexin V conjugates?

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

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.

Go To Top Version: 8    Revision Date: 10/22/2024

For Research Use Only. Not for diagnostic or therapeutic use.

 

This product is supplied subject to the terms and conditions, including the limited license, located at www.biolegend.com/terms) ("Terms") and may be used only as provided in the Terms. Without limiting the foregoing, BioLegend products may not be used for any Commercial Purpose as defined in the Terms, resold in any form, used in manufacturing, or reverse engineered, sequenced, or otherwise studied or used to learn its design or composition without express written approval of BioLegend. Regardless of the information given in this document, user is solely responsible for determining any license requirements necessary for user’s intended use and assumes all risk and liability arising from use of the product. BioLegend is not responsible for patent infringement or any other risks or liabilities whatsoever resulting from the use of its products.

 

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This data display is provided for general comparisons between formats.
Your actual data may vary due to variations in samples, target cells, instruments and their settings, staining conditions, and other factors.
If you need assistance with selecting the best format contact our expert technical support team.

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