Production of anti-glucagon sera with a C-terminal fragment of pancreatic glucagon.

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Abstract
The C-terminal region-specific anti-glucagon sera were raised in rabbits using as immunogen a conjugate of BSA [bovine serum albumin] and a C-terminal fragment of pancreatic glucagon. The hapten was prepared by trypsin digestion of the glucagon, which was a 1:3 mixture of glucagon (18-29) and (19-29). Six rabbits were immunized by s.c. injection of an emulsion of the conjugate with complete Freund''s adjuvant and 5 of the rabbits produced antibodies to the glucagon (GC-1, GC-2, GC-3, GC-5 and GC-6). For comparison, rabbit antisera were also produced against glucagon polymer (GA-10) and syrupy glucagon fibrils (PGA-2). All these antisera and the pancreatic glucagon-specific antiserum 30 K were characterized with dog gut-extract (gut-GLI) and glucagon-related peptide fragments in the radioimmunoassay systems. The assay systems utilized 125I-monosubstituted pancreatic glucagon as tracer and human mono-component glucagon as standard. All sera of the GC-series crossreacted with the dog gut-extract very weakly. Antisera GC-5 and GC-6 exhibited the lowest crossreactivities with the extract, as low as that of 30 K. Characterization of the antiserum GC-5 with purified glucagon-related fragments indicated that the major antigenic determinant located exactly in the C-terminal region of glucagon. High efficiency of the use of the glucagon C-terminal fragment as haptenic immunogen in obtaining the C-terminal region-specific, i.e., pancreatic glucagon-specific antisera was shown.

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