Antibodies were raised against the carboxy terminus of the insulin A chain in sheep, goat and rabbit using as antigen the synthetic octapeptide YQLENYCN conjugated to bovine serum albumin (BSA). All of the antisera obtained cross-reacted with molecules containing this peptide sequence. Using these antibodies, we developed a radioimmunoassay that could detect the insulin A chain itself, both denatured and natural mammalian and avian insulins, as well as proinsulins and insulin fusion proteins from microorganisms.