Isolation and characterization of human thyroxine-binding globulin (TBG)

A two-step procedure for obtaining a preparation of thyroxine-binding globulin (TBG) of high purity from human serum is described. TBG was isolated from sera of pregnant women by affinity chromatography on a thyroxine-coupled Sepharose 4B column using 8anilino-1-naphthalene sulfonic acid (ANS) as the specific eluent, and then purified in the next step by ion-exchange chromatography on a DEAE Sephadex A-50 column. The purity, molecular weight and immunoreactivity of the obtained TBG preparation were evaluated employing electrophoretic and immunological techniques. A single protein band after disc polyacrylamide gel electrophoresis (PAGE) indicated the high purity of the isolated TBG. The molecular weight, estimated by SDS PAGE was about 55 kDa. Preserved immunoreactivity of the isolated protein was demonstrated by immunoelectrophoresis and immune-blotting using specific antibodies. The TBG preparation obtained by the described isolation procedure was of sufficiently high quality (pure, non-denatured and immunoreactive) to be successfully used as a reagent for radioimmunoassay of TBG in human serum.