THE PURIFICATION OF HUMAN α1-T GLYCOPROTEIN FROM SERUM AND ITS CLINICAL APPLICATION

Abstract

Human α1-T glycoprotein (α1-T) was isolated from serum and its clinical significance was investigated. Pooled normal sera were saturated with 40% ammonium sulfate and its supernatant was further saturated to 75%. Precipitates were used as a purification source. Based on the initial procedure by 5 steps of chromatographies we finally developed a simpler modified one with good recovery by reducing 3 steps with the use of monoclonal antibody which consisted of Concanavalin A affinity chromatography, a mouse monoclonal antibody coupled affinity chromatography, and gel-filtration on Sephadex G-200. Purified material was homogenous by sodium dodecyl sulfate Polyacrylamide gel electrophoresis, Polyacrylamide gel electrophoresis and isoelectric focusing. The molecular weight was 77,500 daltons and its isoelectric point was pI 4.5. Amino acid composition was similar to that reported previously by Schwick and Haupt. It contained a trace amount of tryptophan. Using a rabbit antibody and a mouse monoclonal antibody two assays of single radial immunodiffusion and enzyme immunoassay were established. Serum α1-T level was 6.8 mg/dl on the average in 170 normal individuals and urinary excretion was 2.5 mg/day in 14 normal individuals. Serum value was decreased in various disorders and elevated in some cases of acute hepatitis and acute myelogenous leukemia. Its changes in serum had a good correlation with cholinesterase activity and albumin level. These results indicated that α1-T is synthesized mainly by the liver cell and determination of serum α1-T may be a new indicator for evaluating liver cell function.