Protocol

Lysing Tissue-Culture Cells for Immunoprecipitation

This protocol was adapted from "Immunoprecipitation," Chapter 7, in Using Antibodies by Ed Harlow and David Lane. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 1999.

INTRODUCTION

For cells grown in tissue culture, the most useful method of lysis is treating with detergents, as described in this protocol. Non-ionic detergents, such as NP-40, solubilize the plasma and intracellular membranes, break many weak intermolecular bonds, and solubilize most of the commonly studied protein antigens. RIPA lysis buffer, which contains non-ionic detergents mixed with ionic detergents, may be used as a more rigorous extraction buffer to release all but the insoluble proteins of the cell and to break most weak noncovalent interactions. The resulting lysate is ready for preclearing.