Radiation Inactivation of Biological and Immunological Activities of Beef Thyrotropin.

The molecular weight of bovine thyrotropin has been estimated by several investigators. In 1955, Fels, Simpson, and Evans (1) prepared bovine thyroid-stimulating hormone of 1 to 4 units per mg by a method of trichloroacetic acid fractionation and obtained a sedimentation coefficient of 1.02 by ultracentrifugation. They estimated the molecular weight to be 10,000. In 1956, Pierce and Nyc (2) by means of purification on Amerlite IRC-50 prepared bovine TSHl with a potency of 5 U.S.P. units per mg, and by means of ultracentrifugation estimated the sedimentation coefficient to be 2.5 to 2.9. Condliffe and Bates (3) in 1957 by means of chromatography on both IRC-50 and carboxymethylcellulose prepared bovine TSH with a potency of 15 U.S.P. units per mg, and later (4) with a potency of 20 to 50 units per mg. The sedimentation coefficient was estimated to be 2.6 (5). In 1960, Carsten and Pierce (6) by chromatography on DEAEcellulose and by starch gel electrophoresis, prepared bovine TSH with 30 to 60 units per mg. By using the technique of microelectrodialysis they estimated the molecular weight to be 26,000 to 30,000. Fontaine and Condliffe (7) in 1963 with density gradient centrifugation estimated the sedimentation coefficient to be 2.82 S, and the molecular weight to be 26,000 to 31,000. Since completely purified preparations of bovine TSH have not been prepared, these molecular weights should not be viewed as firmly established (5). The molecular volume of protein and polypeptide hormones may be estimated by means of the rate of inactivation of biological activity on exposure to ionizing radiation of low energy transfer (8). The major advantage of this method is that purification is not required. We have studied the inactivation of bovine TSH by 2.0 m.e.v. electrons. Residual activity was measured biologically in the day-old chick and immunologically by a radioimmunoassay. Molecular volumes and weights were calculated by using each of these modalities. These data form the basis of this report.