Analysis of Wheat Germ Agglutinin-Binding Glycoproteins of Rat Hepatocyte Plasma Membrane by Monoclonal Antibodies

Five monoclonal antibodies （2D2, 3C3, 6B4, 10B1 and Y-1） directed to the rat hepatocyte plasma membrane were prepared by immunizing mice with a wheat germ agglutinin （WGA）-binding fraction of a solubilized liver plasma membrane. By the indirect immunocytochemical method, 2D2 and 10B1 could label specifically the bile canalicular membrane. On the other hand, 3C3, 6B4 and Y-1 predominantly labeled the sinusoidal surface of the hepatocytes and the surface of endothelium of all tissues, and to a lesser degree the lateral and bile canalicular membrane of hepatocytes. By Western blotting analysis, each antibody was directed to proteins with a different molecular weight. After isolation of hepatocytes by collagenase, the plasma membrane specialization was lost, and all the antibodies reacted to the entire surface of the isolated hepatocytes. When the hepatocytes were cultured, 2D2 （specific for the bile canalicular membrane） labeled the cell-cell adherent sites and the mombrane of bile canaliculus-like structures, indicating that the bile canaliculus specific molecules are redistributed to these sites. Supplement of 3C3, 6B4 and Y-1 （predominantly reacting to the sinusoidal membrane） to the culture medium resulted in a strong inhibition of adhesion of hepatocytes to the substrate. This suggested that those antibodies （3C3, 6B4 and Y-1） inhibit the receptor molecules which may involve in cell substrate adhesion.departmental bulletin pape