How much insulin-like growth factor I (IGF-I) circulates? Impact of standardization on IGF-I assay accuracy

There is a significant systematic difference between the normal range obtained from ethylenediamine tetraacetate plasma samples using the Genentech total insulin-like growth factor I (IGF-I) RIA and normal ranges for other total IGF-I RIAs. To determine whether the quality of the assay standard was the cause of this systematic difference, we analyzed commercially available preparations of recombinant human IGF-I (rhIGF-I) typical of those used as IGF-I immunoassay standards along with our own well characterized rhIGF-I assay standard. For the commercial standards, high performance liquid chromatography-derived purities were low, and some vendor-assigned protein concentrations were inconsistent with values from quantitative amino acid analysis. The Genentech rhIGF-I assay standard was highly pure and quantitatively correct. However, the poor quality of some commercial rhIGF-I preparations was not the primary reason for the systematic discrepancy between the Genentech total IGF-I RIA normal range and most other normal ranges. Most assays for total IGF-I are calibrated against the WHO International Reference Reagent (IRR) for IGF-I Immunoassays (87/ 518). The Genentech total IGF-I RIA is not calibrated against WHO IRR 87/518. The protein content assigned to WHO IRR 87/518 was a consensus value from a multicenter collaborative study. Physicochemical analyses showed that WHO IRR 87/518 is Met(-1)-IGF-I of low purity (44%), and that the assigned protein content is higher than the value determined by quantitative amino acid analysis. Thus, assays that are calibrated against WHO IRR 87/518 will report total IGF-I concentrations in excess of actual values. We believe that calibration against WHO IRR 87/518 is the cause of the systematic discrepancy between the Genentech IGF-I assay normal range and most other normal ranges, and that much of the plasma IGF-I concentration data in the literature are of questionable accuracy.