Purification of tubulin from limited volumes of cultured cells

A method was designed to purify tubulin from limited volumes of cultured cells, which can be performed in less than 4 h. The method is based on the preservation of intact microtubule arrays during cell lysis in a large volume of buffer, followed by disassembly of microtubules in a small volume of cold buffer. This allows a good enrichment in tubulin, which is then purified by one cycle of polymerisation/depolymerisation and a cation exchange chromatography. Such a procedure has been employed successfully on suspension-cultured and on adherent HeLa cells. Tubulin obtained was >= 90% pure, assembly-competent and composed of alpha/beta I and alpha/ beta IV isotypes. Microtubules made with this tubulin displayed specific properties such as resistance to dilution, maybe related to their specific dynamic behaviour. (C) 2005 Elsevier Inc. All rights reserved.