The aims of this study were to examine the effect of exogenous prostaglandin on mucin secretion and to determine the role of nonmucin glycoproteins on gallstone formation in the prairie dog model of cholesterol cholelithiasis. The concentration of total glycoprotein and nonmucin glycoproteins was measured in gallbladder bile from four groups of prairie dogs fed a control diet or a diet supplemented with 1.2% cholesterol with or without simultaneous subcutaneous administration of prostaglandin E1. Cholesterol feeding resulted in an increased concentration of concanavalin-A binding–proteins in gallbladder bile associated with an increase in pronucleating activity in vitro. Treatment with prostaglandin E1 and cholesterol feeding was associated with a significant increase in the incidence of cholesterol gallstone formation. Prostaglandin E1 treatment in the cholesterol-fed animals increased biliary concentrations of total glycoprotein and concanavalin-A–binding glycoproteins. Therefore the increased biliary glycoprotein level in cholesterolfed, prostaglandin E1-treated prairie dogs, which reflects higher levels of mucin and nonmucin glycoproteins, appears to be an important factor in gallstone formation. (HEPATOLOGY 1993;17:693–700.)