Isolation of plasma membrane and endoplasmic reticulum fragments from chick embryo fibroblasts

Abstract

Plasma membrane and endoplasmic reticulum were isolated from chick embryo fibroblasts using a derivation of the method of Avruch and Wallach ( Biochim. Biophys. Acta, 233 (1971) 334). The fractions isolated were checked for purity using 5′-nucleotidase and (Na +, K +, Mg 2+)-ATPase as markers for plasma membrane, NADPH:cytochrome c reductase as a marker for endoplasmic reticulum and succinate dehydrogenase as a marker for mitochondrial contamination. The method yielded a plasma membrane fraction containing less than 4 % of the endoplasmic reticulum and an endoplasmic reticulum fraction containing less than 2 % of the plasma membrane. Mitochondrial contamination of both fractions was less than 3 %. Similar yields and purity were obtained for fractions isolated from cells which had been infected with either Semliki Forest virus or with Newcastle Disease virus.