FIRST DIRECT ASSAY FOR INTACT HUMAN PROINSULIN

We describe a sensitive two-site sandwich enzyme-linked immunosorbent assay for the measurement of intact human proinsulin in 100 mu L. Of S erum or plasma. The assay is based on the use of two monoclonal antibo dies specific for epitopes at the C-peptide/insulin A chain junction a nd at the insulin B chain/C-peptide junction, respectively. Cross-reac tivities with insulin, C-peptide, and the four proinsulin conversion i ntermediates were negligible. The detection limit in buffer was 0.2 pm ol/L (3 standard deviations from zero). The working range was 0.2-100 pmol/L. The mean intra- and interassay coefficients of variation were 2.4% and 8.9%, respectively. The mean recovery of added proinsulin was 103%. Dilution curves of 40 serum samples are parallel to the proinsu lin calibration curve. Proinsulin concentrations in 20 fasting healthy subjects were all above the limit of detection: median (range), 2.7 p mol/L (1.1-6.9 pmol/L). Six fasting, non-insulin-dependent diabetes me llitus and five insulinoma patients had proinsulin concentrations sign ificantly higher than healthy subjects: median (range), 7.7 pmol/L (3. 2-18 pmol/L) and 153 pmol/L (98-320 pmol/L), respectively.