Isoelectric Focusing/Western Blotting: A Novel and Practical Method for Quantitation of Carbohydrate‐Deficient Transferrin in Alcoholics

摘要: Carbohydrate-deficient transferrin (CDT) has been described as the single, most accurate marker of chronic alcohol consumption. Rapid, sensitive, and specific measurement serum CDT levels can thus provide important clinical information concerning patient diagnosis treatment. To date, however, methods used for assessing concentrations [e.g., analytical isoelectric focusing combined with immunofixation micro anion-exchange chromatography followed by radioimmunoassay (RIA)] have not practical enough widespread laboratory application. In present study, we examined use a different technique, namely (IEF) Western blotting (IEF/WB). Serum proteins (20–40 μg) were first focused according to points (pl) on highresolution agarose IEF gels (ampholyte pH range 5–8) containing nonionic detergent. The transferred electrophoretically nitrocellulose filters, then stained immunochemically antihuman IgG. IEF/WB completely resolved (focusing at pl 5.7 5.9) from other transferring isoforms, assessed neuramidase-generated standards. Computerized densitometric scanning immunoblots allowed be quantitated directly rather than quotient. content determined was highly correlated (r2= 0.962; n= 17) values previously RIA. larger subject group, (mg/liter) measured 139 ± 54 in recently-drinking alcoholics (n= 58), 81 8 abstaining 7), 68 16 healthy control subjects 16). These are similar those reported using quantitation methods. Importantly, influenced severity liver disease among nor did nondrinkers exhibit elevated values. CDT/total ratios (the latter ELISA) offered no advantage over alone distinguish active heavy drinking. conclusion, developed accurate, importantly, method CDT, highly-reliable Routine implementation this technique laboratories rapidly physician powerful diagnostic tool.