UNIT 18.2 Labeling Cultured Cells with 32Pi and Preparing Cell Lysates for Immunoprecipitation
Published Online: 1 MAY 2001
Copyright © 2003 by John Wiley and Sons, Inc.
Lab Protocol Title
Current Protocols in Molecular Biology
How to Cite
Sefton, B. M. 2001. Labeling Cultured Cells with 32Pi and Preparing Cell Lysates for Immunoprecipitation. Current Protocols in Molecular Biology. 40:18.2:18.2.1–18.2.8.
- Published Online: 1 MAY 2001
- Published Print: OCT 1997
This unit describes 32Pi labeling and lysis of cultured cells to be used for subsequent immunoprecipitation of proteins. The approach is appropriate, however, for labeling any cellular constituent with 32P. This procedure is suitable for insect, avian, and mammalian cells and can be used with both adherent and nonadherent cultures. The general approach involves biosynthetic labeling with 32Pi in medium containing a reduced concentration of phosphate. This approach can also be modified to label any cellular constituents with 32Pi. The first procedure described is 32Pi labeling of adherent or nonadherent (e.g., hematopoietic) cells with subsequent lysis in a detergent buffer. More rigorous lysis conditions to be used for working with proteins that are difficult to solubilize are also described.