Abstract
Although protein is the third most abundant solid in bile and is important in cholesterol crystal formation, methods for quantitating the concentration of total protein in bile have not been systematically evaluated. To establish a reliable protein assay for bile, we evaluated three protein assays (Lowry's method and the fluorescamine and Coomassie blue methods), and employed amino acid analysis as a reference technique. Large protein-to-protein variations were observed with the fluorescamine and Coomassie blue methods. Although all assays were affected by interfering substances, Lowry's method and the fluorescamine technique (after trichloroacetic acid precipitation and delipidation of bile) and the Coomassie blue method with native bile showed excellent correlations (P less than 0.0001) with those obtained by amino acid analysis. Using these reliable protein assays, we examined gallbladder bile obtained at surgery from subjects with and without gallstones. No differences in the concentrations of total biliary proteins were observed among patients with cholesterol (n = 23) or pigment (n = 7) gallstones and subjects without gallstones (n = 10). Protein values obtained by amino acid analysis also did not differ among groups. As expected, bile from patients with cholesterol gallstones was supersaturated with cholesterol while bile from nongallstone subjects and those with pigment stones was unsaturated. These results indicate that it is not possible to separate patients with and without gallstones on the basis of the total protein concentration of gallbladder bile.
Highlights
Protein is the third most abundant solid in bile and is important in cholesterol crystal formation, methods for quantitating the concentration of total protein in bile have not been systematically evaluated
Supersaturation of gallbladder bile in cholesterol is the first step in cholesterol gallstone formation [1]
The mechanism of cholesterol crystal formation and the precise role of nucleation modifiers in this process have not been well characterized, recent evidence suggests that the nucleation modifiers in bile are proteins [5,6,7,8]
Summary
Protein is the third most abundant solid in bile and is important in cholesterol crystal formation, methods for quantitating the concentration of total protein in bile have not been systematically evaluated. All assays were affected by interfering substances, Lowry’s method and the fluorescamine technique (after trichloroacetic acid precipitation and delipidation of bile) and the Coomassie blue method with native bile showed excellent correlations (P < 0.0001) with those obtained by amino acid analysis Using these reliable protein assays, we examined gallbladder bile obtained at surgery from subjects with and without gallstones. Proteins are the third most abundant solid in bile [9] and include specific glycoproteins [5] and apolipoproteins [10, 11] that, respectively, can promote [5] or inhibit cholesterol crystal nucleation [7] For these and other reasons, we [10] and others [4] have proposed that quantitative and/or qualitative abnormalities in biliary proteins may be important in the pathogenesis of cholesterol gallstones. The aims of this study were to establish a reliable, precise, and accurate method for measuring total protein concentration in bile and to compare the total protein concentrations of gallbladder bile obtained from subjects with and without gallstones
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