Validation of a rapid solid-phase radioimmunoassay for canine, bovine, and equine insulin.

A rapid and convenient commercial radioimmunoassay kit, developed for quantifying hormones in specimens from human beings, was validated for use in measuring insulin in serum of dogs, cattle, and horses. The procedure uses polypropylene assay tubes treated with rabbit anti-porcine insulin serum and porcine [125I]iodoinsulin. Specificity was proven by demonstrating that standard solutions of porcine insulin and serial dilutions of canine, bovine, and equine sera and pancreatic extracts inhibited binding of [125I]iodoinsulin to the antibody in a parallel manner. Gel-filtration chromatography of pancreatic extracts yielded a major peak of immunoreactive material that eluted identically with [125I]iodoinsulin. Immunoreactivity was not associated with fractions that contain larger and smaller molecular weight peptides (eg, proinsulin and C-peptide, respectively). Biological specificity of the assay was shown by demonstrating increased insulin in serum after injection of glucose into heifers and glucagon into dogs and horses. Purified insulin and insulin in pancreatic extracts could be quantitatively recovered from serum, thereby demonstrating accuracy of the assay. Interassay precision of 5 control specimens run in 20 consecutive assays ranged from 6.7% to 20.1% (coefficient of variation) and intra-assay precision of 6 control specimens each assayed 10 times ranged from 4.4% to 10.7% (coefficient of variation). Sensitivity of the assay was 3.2 microIU/ml. This radioimmunoassay for insulin is ideal for veterinary research and diagnosis, because a single set of reagents and procedures can be used for at least 3 species.