Enzyme immunoassay of pancreatic glucagon at the picogram level using beta-D-galactosidase as a label [dog]

An enzyme immunoassay of pancreatic glucagon was established by using E. coli β-D-galactosidase [EC 3.2.1.23] as a marker. In order to increase the sensitivity of the immunoassay, different peptides obtained from glucagon fragments were used to produce the enzyme conjugate and the immunogen. Antiserum N6E raised against C-terminal fragment peptide (15–29) could be diluted to more than 1: 100,000 in the assay and was highly specific for pancreatic glucagon. The antiserum reacted well with the C-terminal fragment peptide (21–29) as well as another fragment peptide (15–29) and pancreatic glucagon. The enzyme immunoassay using antiserum N6E and fragment peptide (21–29)-enzyme conjugate could detect as little as 1 to 2 pg of glucagon. The mean recovery of glucagon added to serum specimens was 104% and the coefficients of variation were 3.7–14.5% (within assay) and 9.0–18.5% (between assay).