C-peptide immunochemiluminometric assay developed from two seemingly identical polyclonal antisera

Two goats were immunized with synthetic C-peptide. Their antisera were immunopurified separately on a C-peptide affinity column. The purified antibodies from one goat were labeled with acridinium ester; the other goat's antibodies were immobilized on plastic beads. Standards were synthetic C-peptide. The new immunochemiluminometric assay (ICMA) for C-peptide was compared with our routine radioimmunoassay (RIA) by using Novo's 125I-labeled C-peptide and Cambridge Medical's antiserum. Good correlations were found between the new ICMA and the RIA (r = 0.951; ICMA = 1.07 RIA + 147; n = 112). The new ICMA showed good recoveries (91 percent to 108 percent) of added C-peptide and parallelism of diluted specimens. The incubation time was shortened from 48 hours for RIA to five hours for ICMA. In addition, it was shown that polyclonal antisera from two animals immunized by the same antigen and purified by the same affinity column can be used to prepare immunometric assays.

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