A candidate carrier protein for beta-human chorionic gonadotropin: 54,000-molecular-weight fragment of tetanus toxin

As an alternative to intact tetanus toxoid as a carrier for beta-human chorionic gonadotropin (beta-hCG), a fragment of tetanus toxin was sought that had a relatively low molecular weight, yet was highly immunogenic. Purified culture filtrate tetanus toxin was subjected to limited enzymatic digestion with papain, and the resulting fragments separated by column chromatography on Sephadex G-150. Four fractions were thus identified. Fraction II was found to have a molecular weight of 54,000 by SDS-polyacrylamide gel electrophoresis. This fragment was covalently linked to the beta-subunit of hCG (beta-hCG-TTII) using carbodiimide hydrochloride. The ability of beta-hCG-TTII to stimulate production of anti-hCG sera in rabbits was measured by 125I-hCG radioimmunoassay. At sera dilutions of 1:40,000, an average 125I-hCG binding capacity of 34.7 +/- 5.86% (mean +/- SD) was observed 8 weeks after the final immunization. Tetanus toxin Fragment II has the potential for future application in active immunization studies involving hormone-carrier conjugates.