Solubilization of Lymphocytes

The purification and study of transmembrane proteins requires isolation of these structures from their lipid environment. This isolation is carried out through the use of detergents. In this unit, several approaches to solubilization of membrane proteins are presented. Solubilization of whole lymphocytes, using conditions aimed at minimizing the disruption of protein-protein interactions, is described in the first basic protocol; an optional step is included that may be useful when the disruption of protein interactions is desired as part of a purification protocol. In some situations, it may be desirable to purify membranes prior to their solubilization (see second basic and alternate protocols). To determine the physical relationship between proteins, a cross-linking protocol (see support protocol) is provided.

The protocols presented in this section are intended as a general guide for solubilization of lymphocyte and lymphocyte-membrane proteins. They are intended for use in conjunction with immunoprecipitation of radiolabeled proteins (unit 8.3), immunoaffinity purification (unit 8.2), immunoblotting (unit 8.10), or as the basis for a scale-up in the initial steps of large-scale protein purification. Parameters such as numbers of cells and lysis volume are based on studies using a T cell hybridoma. Variation of the solubilization procedures presented are given in units 8.2, 8.3, & 8.5.

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