Neutralizing and non-neutralizing antibodies to bovine thyroid-stimulating hormone and its subunits

To test the possibility that the long-acting thyroid stimulator (LATS) might represent an immune complex either of thyroid-stimulating hormone (TSH) with anti-TSH or of a subunit of TSH with an appropriate antibody, we immunized rabbits with bovine TSH (bTSH), bLH (luteinizing hormone), and their alpha and beta subunits (bTSHalpha and bTSHbeta). Binding, neutralizing, and nonneutralizing antibodies were demonstrated in the antisera obtained. First, antisera to TSH, TSHbeta, and TSHalpha all bound [(125)I]TSH and [(125)I]TSHbeta. Anti-bTSHbeta antisera bound [(125)I]bTSHbeta better than did anti-TSH sera, while the binding of [(125)I]bTSH was similar with both types of antiserum. Second, the thyroid-stimulating activity (McKenzie bioassay) of TSH could be neutralized by incubation with various dilutions of anti-TSH or anti-TSHbeta. Finally, when incubation mixtures containing TSH and dilutions of anti-TSHbeta antisera that only partially neutralized TSH were treated with an antiserum against rabbit immunoglobulins to precipitate immune complexes, the bioassay response of the TSH was abolished. This phenomenon was not observed when antiserum to the intact hormone was substituted in the incubation mixture. The removal of TSH biological activity from a mixture of TSH and anti-bTSHbeta by addition of an anti-immunoglobulin indicated that biologically active immune complexes were formed between TSH and anti-TSHbeta but not between TSH and anti-TSH. The time-course of the bioactivity and several other characteristics of these complexes differentiate them from LATS.