A domain-specific marker for the hepatocyte plasma membrane. III. Isolation of bile canalicular membrane by immunoadsorption.
Previous immunolabeling studies (Roman, L.M., and A.L. Hubbard, 1983, J. Cell Biol., 96:1548-1558; Roman, L.M., and A.L. Hubbard, 1984, J. Cell Biol., 98:1488-1496, companion paper) established leucine aminopeptidase (LAP) as a specific marker for the bile canalicular (BC) domain of the rat hepatocyte plasma membrane (PM). In this study, we have isolated membrane from a sonicated PM vesicle fraction using anti-LAP-coated Staphylococcus aureus cells as a solid-phase immunoadsorbent. The extent and specificity of the immunoadsorption were assessed by following the behavior of LAP (the BC marker) and 32P-labeled membrane phospholipids (a uniform membrane marker). The BC fraction obtained was significantly enriched in LAP (yield: greater than 70% of PM-LAP). Alkaline phosphatase, 5'-nucleotidase, and a 110,000-dalton glycoprotein, HA-4, were enriched in the BC fraction to the same extent as LAP (enzyme or antigen/LAP = 1.0). However, alkaline phosphodiesterase I was not enriched to the same degree (enzyme/LAP = 0.5). Contamination of this BC fraction by membrane derived from the sinusoidal domain and endoplasmic reticulum, as determined from the distribution of the asialoglycoprotein receptor and NADH cytochrome c reductase, respectively, was small (less than 13%).
D000818:Animals; D000906:Antibodies; D000936:Antigen-Antibody Complex; D002458:Cell Fractionation; D002462:Cell Membrane / Q000648:ultrastructure*; D005455:Fluorescent Antibody Technique; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000648:ultrastructure*; D008297:Male; D008565:Membrane Proteins / Q000302:isolation & purification; D008854:Microscopy, Electron; D008970:Molecular Weight; D010761:Phosphorus Radioisotopes; D051381:Rats; D011919:Rats, Inbred Strains; D013211:Staphylococcus aureus / Q000276:immunology; D014465:Ultrasonics
D000906:Antibodies; D000936:Antigen-Antibody Complex; D008565:Membrane Proteins; D010761:Phosphorus Radioisotopes