Determination of insulin-like growth factor-II in human serum

Insulin-like growth factor-II (IGF-II) is a polypeptide mitogen present at relatively high concentrations in the human peripheral circulation, The aim of the present investigation was to develop a reliable assay system for determining total IGF-II, for use in studies of the IGF system. A competitive binding assay was set up using a specific monoclonal anti-IGF-II antibody and radioactively labelled recombinant human IGF-II as the tracer. Displacement curves within the IGF-II concentration range from 0.08 to 5.35 nmol/L were obtained. Cross reactivity with IGF-I at 50% inhibition of tracer binding was only 0.4%. Human serum samples were subjected to acid-ethanol extraction followed by neutralisation and cryoprecipitation in order to remove interfering binding proteins before analysis. This was confirmed by the finding that an excess of IGF-I added at a level below cross reacting concentrations to sequester residual binding proteins did not affect the values obtained for IGF-II in relation to the standard curve. The serum concentrations of total IGF-II determined in three groups of healthy subjects (n = 53) were similar to those recorded by other authors and ranged from 48 to 115 nmol/L. Mean (SD) recovery of IGF-II added to seven sera before extraction was 102 (7.4)%. Intra-assay precision and inter-assay reproducibility were satisfactory. While IGF-II concentrations in sera stored at -20 degrees C were relatively stable for several months, it was necessary to assay the extracts within 48 h of preparation.