A double antibody adaptation of the glucagon radioimmunoassay is described in detail. Guinea pig antiserum allowing the assay of 2,500 unknowns per milliliter of undiluted serum was induced by biweekly immunization. Glucagon was iodinated with I-125 and purified by elution from a cellulose column, maximal immunoprecipitibility approximating 80 per cent. Standard curves in buffer were sensitive to purified glucagon in amounts of less than 100 μμg.
Rapid degradation of labeled glucagon added to human blood was demonstrated. Trasylol was confirmed as an effective inhibitor of this degradation. Endogenous plasma immunoreactive glucagon (IRG) appeared less labile and may have interfered with the quantitative detection of unlabeled exogenous beef-pork glucagon in plasma recovery studies.
The hypothesis is offered that plasma IRG, herein measured at 400 ± 220 (S.D.) μμg./ml., may be composed of a pancreatic and an extrapancreatic fraction of similar but not identical immunological activity. The cross-reactivity of the two components remains a problem in the interpretation of the glucagon immunoassay as applied to plasma.
