Unit

UNIT 18.2 Labeling Cultured Cells with 32Pi and Preparing Cell Lysates for Immunoprecipitation

  1. Bartholomew M. Sefton

Published Online: 1 MAY 2001

DOI: 10.1002/0471142727.mb1802s40

Current Protocols in Molecular Biology

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.

Author Information

  1. The Salk Institute, San Diego, California

Publication History

  1. Published Online: 1 MAY 2001
  2. Published Print: OCT 1997

Abstract

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.