A Simple Protocol for Separation and Assay of µ-Calpain, m-Calpain and Calpastatin from Small Tissue Samples

Abstract

In many cases separation of calpains and calpastatin has to be performed with a relatively small amount of tissue, so that large-scale and complicated multiple-column procedures are not appropriate. In designing a method suitable for small amounts of tissue, it is important that the extraction and separation methods should be rapid in order to avoid postmortem changes in the subcellular distribution of the enzymes, and also to avoid autolysis or degradation of the enzymes themselves. The method presented here uses relatively simple and inexpensive equipment and reagents, although the assay of calpain activity is greatly simplified by access to a fluorometer with the ability to read microtiter plates.