Radioimmunoassay of a basic somatomedin: comparison of various assay techniques and somatomedin levels in various sera

Abstract

Purification of a basic somatomedin (SM), with similarity to SM-C and insulin-like growth factor, from human plasma Cohn fraction IV-1 enabled development of a RIA based on this SM.SM antiserum was produced by immunizing rabbits with partially purified SM. This antiserum (final dilution, 1:50,000) specifically bound approximately 40% of added [125I]-SM in this RIA. The RIA sensitivity was 2 x 10(-4) U immunoreactive SM (IRSM). Highly purified SM-C, insulin-like growth factor 1, and our SM revealed parallel and approximately equipotent dose-response curves in this RIA; rat SM and multiplication stimulation activity revealed less cross-reactivity. IRSM was detected in sera of all species tested except fish. Acidification of sera, without subsequent chromatography, before assay permitted measurement of total IRSM with either an equilibrium or nonequilibrium RIA technique. Acidification of serum appears to increase SM-binding capacity while decreasing binding affinity of the 20,000--50,000 mol wt proteins in serum. The mean (+/- SEM) IRSM concentrations in sera from normals and patients with acromegaly, hypopituitarism, GH deficiency before/after treatment, and Laron dwarfism were 1.45 +/- 0.17, 5.49 +/- 0.48, 0.19 +/- 0.07, 0.10 +/- 0.02/0.64 +/- 0.45, and 0.25 +/- 0.11 U/ml, respectively, compared to a pooled normal human serum reference standard which was designated to contain 1 IRSM U/ml. Measurements of total IRSM (bound and free) in serum may not accurately reflect SM bioactivity and will require interpretative caution.