Because of the widespread long-term clinical use of LHRH agonists, the possibility of antibody development to one of these, (D-Ser-(Butt)6) LHRH (1-9) nonapeptide-ethylamide (buserelin), was studied by examining sera from experimental animals [stumptailed monkey, rhesus monkey, rabbit, dog, rat] and humans receiving long-term agonist-treatment for ability to bind 125I-labeled buserelin. None of the sera from monkeys, rabbits, dogs or rats receiving buserelin by s.c. injection or via minipump or from 81 patients receiving the agonist by s.c. injection or nasal spray had detectable antibodies. The immunochemistry of buserelin was investigated by actively immunizing male rabbits against the agonist. This also enabled determination of the possibility of this stimulus to the immune response resulting in the generation of antibodies which could cross-react with endogenous LHRH and neutralize its biological effect. All 41 rabbits immunized against unconjugated buserelin and all 23 immunized against buserelin conjugated to a protein carrier developed antibodies, those in the latter group producing highest titers. Serum from one of the animals immunized against unconjugated buserelin and from 9 of the animals immunized against conjugated buserelin had the capacity to bind 125I-labeled LHRH. These LHRH antibody titers were very low and were insufficient to cause suppression of serum testosterone concentrations or affect testicular weight except in 1 rabbit immunized against unconjugated agonist. The LHRH antibody titer in this rabbit was always < 1:100 which other studies have indicated to be too low to produce a biological effect. The association of low LHRH antibody titer with decreased testes weight in this animal was fortuitous. The possibility of circulating antibodies being produced to buserelin or their interfering with the action of endogenous LHRH is extremely low.