Purification of testosterone‐oestradiol‐binding globulins from mammalian sera by anion‐exchange high performance liquid chromatography

Abstract

Testosterone‐oestradiol‐binding globulin (TeBG) has been isolated from serum or plasma of several species using procedures that yielded highly purified protein, but which required multiple and tedious chromatographic steps. In this report we describe a procedure for the isolation of TeBG which involves two chromatographic steps: androgen affinity chromatography followed by anion‐exchange high performance liquid chromatography (anion‐exchange HPLC). The purity of the final product was confirmed by silver staining following fractionation on sodium dodecyl sulphate‐containing polyacrylamide gels. The size heterogeneity and specific binding activity of TeBGs purified from human, rabbit, or bull serum (or plasma) by this technique was indistinguishable from preparations obtained by conventional chromatography. The present technique shortened the entire purification procedure to about 5 working days and yielded milligram quantities of highly purified protein. Bases on our experience with serum or plasma from the human, rabbit, and bull, this approach should be suitable for isolation of TeBG from a wide range of species.