Radioimmunoprecipitation assay buffer

Radioimmunoprecipitation assay buffer (RIPA buffer) is a lysis buffer used to lyse cells and tissue, for radio immunoprecipitation assay (RIPA).^[1][2] This buffer is more denaturing than NP-40 or Triton X-100 lysis buffer because it contains the ionic detergents SDS and sodium deoxycholate as active constituents and is particularly useful for disruption of nuclear membranes in the preparation of nuclear extracts. The RIPA buffer gives low background but can denature kinases.

Recipe

RIPA buffer recipe for total protein extract is as follows:

• 25mM Tris (10 mM sodium phosphate may be used instead), pH 7–8
• 150 mM NaCl
• 0.1% SDS (optional)
• 0.5% sodium deoxycholate
• 1% Triton X-100 or NP-40
• Protease inhibitors such as PMSF or cocktail protease inhibitors which are commercially available (use according to the manufacturer's guide)

References

1. Alcaraz C, De Diego M, Pastor MJ, Escribano JM (July 1990). "Comparison of a radioimmunoprecipitation assay to immunoblotting and ELISA for detection of antibody to African swine fever virus". J. Vet. Diagn. Invest. 2 (3): 191–6. doi:10.1177/104063879000200307. PMID 2094444.
2. Ngoka LC (October 2008). "Sample prep for proteomics of breast cancer: proteomics and gene ontology reveal dramatic differences in protein solubilization preferences of radioimmunoprecipitation assay and urea lysis buffers". Proteome Sci. 6 (1): 30. doi:10.1186/1477-5956-6-30. PMC 2600628. PMID 18950484.