Loading...

FITC Annexin V Apoptosis Detection Kit with 7-AAD

Pricing & Availability
Regulatory Status
RUO
Other Names
Annexin A5 Apoptosis Detection Kit
Ave. Rating
Submit a Review
Product Citations
publications
FITC_AnnexinV_Apoptosis_Detection_Kit_7AAD_1_103012
Human T-cell leukemia cell line, Jurkat, treated (top) or non-treated (bottom) with BioLegend's LEAF™ purified anti-human CD95 (clone EOS9.1) mAb (Cat. No. 305704) for 4 hours, then stained with FITC Annexin V Apoptosis Detection Kit with 7-AAD (Cat. No. 640922).
  • FITC_AnnexinV_Apoptosis_Detection_Kit_7AAD_1_103012
    Human T-cell leukemia cell line, Jurkat, treated (top) or non-treated (bottom) with BioLegend's LEAF™ purified anti-human CD95 (clone EOS9.1) mAb (Cat. No. 305704) for 4 hours, then stained with FITC Annexin V Apoptosis Detection Kit with 7-AAD (Cat. No. 640922).
  • FITC_AnnexinV_Apoptosis_Detection_Kit_7AAD_2_103012
See FITC spectral data
Cat # Size Price Quantity Check Availability Save
640922 100 tests 203€
Check Availability


Need larger quantities of this item?
Request Bulk Quote
Description

BioLegend's FITC Annexin V Apoptosis Detection Kit with 7-AAD 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 7-amino-actinomycin D (7-AAD) solution. Early apoptotic cells will exclude 7-AAD, while late stage apoptotic cells will stain positively, due to the passage of these dyes into the nucleus where they bind to DNA.

7-AAD (7-amino-actinomycin D) has a high DNA binding constant and is efficiently excluded by intact cells. It is useful for DNA analysis and dead cell discrimination during flow cytometric analysis. When excited by 488 laser light, 7-AAD fluorescence is detected in the far red range of the spectrum (650 nm long-pass filter).
 

Product Details
Technical Data Sheet (pdf)

Product Details

Verified Reactivity
Human, Mouse, Rat
Reported Reactivity
Other Species
Concentration
Lot-specific (to obtain lot-specific concentration and expiration, please enter the lot number in our Certificate of Analysis online tool.)
Storage & Handling
Store between 2°C and 8°C. Do not freeze.

Caution: 7-AAD is a potential carcinogen. It is recommended that the user wear protective clothing, gloves, and eye/face protection in order to avoid contact with skin and eyes.
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 5 µL of 7-AAD Viability Staining 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

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. Maciel E, et al. 2014. Arch Biochem Biophys. 548:38. PubMed
Product Citations
  1. Ferriz M, et al. 2023. STAR Protoc. 4:102079. PubMed
  2. Barrios-Bernal P, et al. 2023. Int J Mol Sci. 24:. PubMed
  3. Ribeiro MS, et al. 2023. Methods Mol Biol. 2618:289. PubMed
  4. Mohammadpour H, et al. 2023. Cell Rep. 42:112250. PubMed
  5. Park SA, et al. 2023. Int J Mol Sci. 24:. PubMed
  6. Groß E, et al. 2023. Cancers (Basel). 15:. PubMed
  7. Delangre E, et al. 2021. Cell Death Dis. 12:1136. PubMed
  8. Zhu Y, et al. 2022. Oxid Med Cell Longev. 2022:3182931. PubMed
  9. Dubiella C, et al. 2021. Nat Chem Biol. 17:954. PubMed
  10. Silveira DB, et al. 2022. Cell Biochem Funct. 40:163. PubMed
  11. Jang HJ, et al. 2022. Cancer Res. 82:2593. PubMed
  12. Bellanti F, et al. 2023. Int J Mol Sci. 24: . PubMed
  13. Lee H, et al. 2019. Cancer Res. 79:2839. PubMed
  14. Staresinic B, et al. 2018. Sci Rep. 8:9412. PubMed
  15. Sima LE, et al. 2021. J Immunother Cancer. 9:. PubMed
  16. Khodeer S, et al. 2022. J Cell Sci. 135: . PubMed
  17. Karabicici M, et al. 2021. Mol Oncol. 15:2185. PubMed
  18. Witney T, et al. 2015. Clin Cancer Res. 21: 3896-3905. PubMed
  19. Maeda J, et al. 2016. PLoS One. 11: 0156689. PubMed
  20. Wang Z, et al. 2020. Cancer Manag Res. 3.563194444. PubMed
  21. Deng H, et al. 2020. Nat Commun. 3.896527778. PubMed
  22. Zhang Q, et al. 2021. Theranostics. 11:8301. PubMed
  23. Koolivand M, et al. 2022. Cell J. 24:44. PubMed
  24. Liu M, et al. 2016. Cell Mol Gastroenterol Hepatol. 2:63-76. PubMed
  25. Demelash A, et al. 2015. J Biol Chem. 290: 21962-21975. PubMed
  26. Saunders IT, et al. 2019. Cancer Cell Int. 19:98. PubMed
  27. Znidar K, Bosnjak M 2015. Mol Ther Nucleic Acids. 5:e322. PubMed
  28. Tsuchiya N, et al. 2020. Cell Reports. 29(1):162-175.e9.. PubMed
  29. Zietzer A, et al. 2020. J Extracell Vesicles. 9:1786967. PubMed
  30. Khan H, et al. 2019. Cell Rep. 29:3009. PubMed
  31. Ruivo CF, et al. 2022. Gut. . PubMed
  32. Ma J, et al. 2021. Curr Protoc. 1:e144. PubMed
  33. Ma L, et al. 2015. Blood. 126: 247 - 256. PubMed
  34. Martínez-Montemayor MM, et al. 2019. Front Pharmacol. 10:115. PubMed
  35. Davies SP, et al. 2019. Cell Rep. 29:1610. PubMed
  36. Davenne T, et al. 2020. Cell Rep. 31:107640. PubMed
  37. Yan C, et al. 2019. Cancer Biol Med. 16:109. PubMed
  38. Zhang H, et al. 2020. Proc Natl Acad Sci U S A. 117:29862. PubMed
  39. Wang L, et al. 2022. Am J Transl Res. 14:150. PubMed
  40. Oshima R, et al. 2016. Sci Rep. 6: 24997. PubMed
  41. Kim S, et al. 2021. J Clin Invest. 131:. PubMed
  42. Yang Q, et al. 2016. Mol Biol Cell. 27: 197 - 208. PubMed
  43. Lee HS, et al. 2019. Interact Cardiovasc Thorac Surg. 28:353. PubMed
  44. Ito T, et al. 2021. Commun Biol. 4:694. PubMed
  45. Maciel E, et al. 2014. Arch Biochem Biophys. 548:34. PubMed
  46. Zhang D, et al. 2020. Stem Cell Res Ther. 0.816666667. PubMed
  47. Chen Z, et al. 2019. J Exp Med. 216:152. PubMed
  48. Parv K, et al. 2021. Front Endocrinol (Lausanne). 12:606175. PubMed
  49. Boemi I, et al. 2021. Front Cell Dev Biol. 9:555248. PubMed
  50. Gudikote JP, et al. 2021. J Biol Chem. 297:101163. PubMed
  51. Chen SM, et al. 2021. Biomolecules. 11:. PubMed
  52. Perkhofer L, et al. 2016. Stem Cell Res. 17: 367-378. PubMed
  53. Osei-Amponsa V, et al. 2020. Mol Cell Biol. 40:00:00. PubMed
  54. Wright JA, et al. 2021. Front Immunol. 12:705307. PubMed
  55. Ma S, et al. 2016. Cell Death Dis. 7:e2307. PubMed
  56. Karabicici M, et al. 2021. Front Cell Dev Biol. 9:639779. PubMed
  57. Krohn S, et al. 2022. Front Immunol. 13:908093. PubMed
  58. Pham DV, et al. 2022. J Exp Clin Cancer Res. 41:9. PubMed
  59. Zietzer A, et al. 2021. Cell Mol Life Sci. 79:48. PubMed
  60. Bianchi M, et al. 2021. Innate Immun. 27:89. PubMed
  61. Pan Y, et al. 2017. Nature. 543:252-256. PubMed
  62. Sim CK, et al. 2019. J Immunol Res. 2019:7596786. PubMed
  63. Lang L, et al. 2017. J Exp Clin Cancer Res. 10.1186/s13046-017-0583-4. PubMed
  64. Yang L, et al. 2021. Front Immunol. 12:722273. PubMed
  65. Wei L, et al. 2015. PLoS One. 10: 0131763. PubMed
  66. Artru F, et al. 2020. J Hepatol. 72:1052. PubMed
  67. Urdinguio RG, et al. 2019. Nucleic Acids Res. 47:5016. PubMed
  68. Aggarwal N, et al. 2021. Cell Rep. 37:110170. PubMed
  69. Balani DH, et al. 2021. J Bone Miner Res. 36:757. PubMed
  70. Simula L, et al. 2020. Cell Death Differ. 27:2749. PubMed
  71. Barbu EA, et al. 2020. Front Immunol. 1.385416667. PubMed
  72. Notara M, et al. 2015. Stem Cell Res. 15: 643-654. PubMed

Antigen Details

Biology Area
Apoptosis/Tumor Suppressors/Cell Death, Cell Biology, Cell Proliferation and Viability, Neuroscience
Molecular Family
Cytokines/Chemokines
Gene ID
308 View all products for this Gene ID

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.

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.

Why is washing not recommended after the addition 7-AAD or PI addition when assessing viability?

These dyes bind in equilibrium with DNA. Therefore, external dye concentration must be maintained during analysis and the dye should not be washed out.

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: 4    Revision Date: 11/01/2023

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.

 

BioLegend, the BioLegend logo, and all other trademarks are property of BioLegend, Inc. or their respective owners, and all rights are reserved.

 

8999 BioLegend Way, San Diego, CA 92121 www.biolegend.com
Toll-Free Phone: 1-877-Bio-Legend (246-5343) Phone: (858) 768-5800 Fax: (877) 455-9587

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.

ProductsHere
Login / Register
Remember me
Forgot your password? Reset password?
Create an Account