Each lot of this antibody is quality control tested by our Ki-67 staining protocol below. For flow cytometric staining, the suggested use of this reagent is ≤ 0.25 ?g per million cells in 100 ?l volume. For Western blotting, the suggested use of this reagent is 0.5??g per ml (1:1000).?It is recommended that the reagent be titrated for optimal performance for each application.

Additional reported applications (for the relevant formats) include: immunohistochemical staining of frozen tissue sections¹, Western blotting³, and immunofluorescence microscopy⁴.

Ki-67 Staining Protocol:

1. Prepare 70% ethanol and chill at -20°C.
2. Prepare target cells of interest and wash 2X with PBS by centrifuge at 350xg for 5 minutes.
3. Discard supernatant and loosen the cell pellet by vortexing.
4. Add 3 ml cold 70% ethanol drop by drop to the cell pellet while vortexing.
5. Continue vortexing for 30 seconds and then incubate at -20°C for 1 hour.
6. Wash 3X with BioLegend Cell Staining Buffer and then resuspend the cells at the concentration of 0.5-10 x 10⁶/ml.
7. Mix 100 ?l cell suspension with proper fluorochrome-conjugated Ki-67 antibody and incubate at room temperature in the dark for 30 minutes.
8. Wash 2X with BioLegend Cell Staining Buffer and then resuspend in 0.5 ml cell staining buffer for flow cytometric analysis.

1. Gerdes J, et al. 1983. Int. J. Cancer 31:13. (IHC)
2. Gerdes J, et al. 1984. J. Immunol. 133:1710. (ICFC)
3. Schluter C, et al. 1993 J. Cell Biol. 123:513. (IHC, WB)
4. Bading H, et al. 1989 Exp. Cell. Res. 185:50. (IF)
5. Guha P, et al. 2013. PNAS. 110:5052. PubMed

1. Santos R, et al. 2017. Nat Commun. . 10.1038/s41467-017-01760-5. PubMed
2. Wang L, et al. 2022. Oncol Lett. 23:62. PubMed
3. Mann ER, et al. 2020. Sci Immunol. :5. PubMed
4. Guha P, et al. 2013. Proc Natl Acad Sci U S A. 110:5052. PubMed
5. Junghof J, et al. 2022. NPJ Regen Med. 7:14. PubMed
6. O'Boyle KC, et al. 2020. Methods Mol Biol. 2111:1. PubMed
7. Bruno TC, et al. 2017. Cancer Immunol Res. 0.831944444. PubMed
8. Agrawal N, et al. 2018. Front Immunol. 2.053472222. PubMed
9. Xie S, et al. 2020. Ann Transl Med. 1.384027778. PubMed
10. Eccles JD, et al. 2020. Cell Rep. 30:351. PubMed
11. Tailor D, et al. 2021. Cell Chemical Biology. . PubMed
12. Zhou H, et al. 2017. Leukemia. 31:2065. PubMed
13. Xiong Y, et al. 2019. Onco Targets Ther. 12:993. PubMed
14. Vining KH, et al. 2018. Adv Mater. 30:4. PubMed
15. Wang Z et al. 2019. Br J Pharmacol. 176(17):3390-3406 . PubMed

Expressed in the phases G₁, S, G₂, and M of the cell cycle

1. Byeon IJ, et al. 2005. Nat. Struct. Mol. Biol. 12:987.
2. Yerushalmi R, et al. 2010. Lancet. Oncol. 11:174.
3. Beltrami AP, et al. 2001. N. Engl. J. Med. 344:1750.
4. Sachsenberg N, et al. 1998. J. Exp. Med. 187:1295.
5. Nagy Z, et al. 1997. Acta. Neuropathol. 93:294.