GLUCAGON ANTIBODIES AND AN IMMUNOASSAY FOR GLUCAGON*

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Abstract
Uncertainty as to the hormonal status of glucagon can be attributed to the lack of a specific method for its measurement in plasma. The following study was designed to determine if an immunoassay for glucagon, using the radiochromatographic techniques of Berson and Yalow, might be developed. Glucagon antibodies were produced in rabbits. Antibodies of the non-precipitating type were present in the gamma globulin fraction and the kinetics of their reaction with glucagon-I131 was similar to that of the insulin-antibody reaction. Non-radioactive glucagon was found to inhibit competitively the reaction between glucagon-I131 and antibody, providing the basis for an immunoassay. The ratio of anti-body-bound to free glucagon-I131 could be progressively lowered by the addition of increasing amounts of non-radioactive glucagon. By employing a 1:100 dilution of antiserum with 50-100 uugm glucagon-I131 as little as 50 uugm of unlabeled glucagon could be measured. Other peptide hormones, insulin, pitressin, and ACTH did not influence the assay. Injected glucagon could be recovered from human blood and its time disappearance curve, as determined by the immunoassay, was qualitatively similar to the disappearance curve of glucagon-I131. Extracts of the pancreas of dogs and of humans gave extremely high values for glucagon, even in 800-fold dilutions, whereas in extracts of organs known not to contain glucagon, no glucagon could be assayed. These studies reveal the antigenicity of beef-pork glucagon in rabbits and indicate the feasibility of a specific and sensitive method for measuring glucagon. The cross-reactivity of dog and of human glucagon with rabbit antibodies to beef-pork-insulin has been established.