[Obtaining immune sera to high-titre thyroxine for immunoassay]

A method of synthesis of L-thyroxine-protein conjugates was described. It included an additional step of acylation of thyroxine by succinic anhydride. The acylated derivative was activated by carbodiimide to produce a compound which could react with the protein amino-groups. Protein conjugates with a high thyroxine content (over 20 thyroxine residues per molecule) were synthesized with minimal protein cross-linking using this method. The excess of acylated thyroxine was easily dialized against alkaline buffer. The rabbits immunized monthly with such conjugates gave antisera with high titers (1:2000-1:4000 by the ELISA method) in 9-13 weeks after the first injection. The method can be applied to other thyroid hormones and some-molecular-weight antigens.