Isolation of sinusoidal and canalicular liver plasma membranes: Effects of frozen storage of human material

Abstract

Sinudoisal and canalicular plasma membranes were prepared from rat and fresh or frozen human liver and characterized using plasma membrane marker enzymes. The distributions of these enzymes were used as a means of comparison between the plasma membrane fractions and homogenates from the rat and fresh and frozen human livers. Compared with their homogenate, all the sinusoidal plasma membrane preparations were poorly enriched with Na +-K +-ATPase (2.6–8.4-fold), a sinusoidal marker enzyme. Activities of Na +-K +-ATPase and Mg 2+-ATPase were much greater in rat membranes whereas leucine aminopeptidase and alkaline phosphatase activities were much greater in the human membranes. Consideration of the extent of enrichment showed that Mg 2+-ATPase and alkaline phosphatase are better markers in rat canalicular plasma membranes and leucine aminopeptidase in human membranes from both fresh and frozen tissue. Intracellular organelle marker enzymes were not enriched in the plasma membranes, thus indicating no significant contamination. Importantly, the relative enrichments of the marker enzymes showed no difference between fresh and frozen human liver. Absolute activities of the enzymes were decreased, however (except alkaline phosphatase), after frozen storage. This work indicates that human liver tissue may be frozen and stored in liquid nitrogen before plasma membranes are isolated with no effect on the distribution of marker enzymes in the membranes.