Study of a lysis medium stabilizing microfilaments and microtubules in vitro and in vivo

Publisher Website
Google Scholar
Abstract
Determination of experimental conditions which allow the evaluation of the variations in the ratio of non-polymerized and polymerized forms of actin and tubulin during the reorganization of the cytoskeletal cell system is of most valuable importance. In order to prepare cell homogenates which would reflect the in vivo situation, a lysis medium which stabilized both microfilaments and microtubulesof Arabacia punctulata and Paracestrotus liwidus, which were determined by DNase inhibition assays and colchicine binding assays respectively were tested in vitro. The lysis medium containing 10 mM K phosphate, 1mM Mg Cl, 5 mM EGTA [ethylene Glycoc-bis-(.beta.-amino ethyl) ether-N,N,N'',N''-tetraacetic acid], 1 M hexylene glycol, 1% Triton X-100, pH 6.4, used at 4.degree. C diffused rapidly into the cells; did not denature actin and tubulin; did not displace the equilibrium between non-polmerized and polymerized forms of actin and tubulin, allowing biochemical assays on cell homogenates; blocked the evolution of the cytoskeletal system and permitted structural studies; and allowed the staining of microfilaments by heavy meromyosin.