The free Insulin-like growth factor-I (IGF-I) in plasma from normal adults was directly measured with a newly developed highly sensitive immunoradiometric assay (IRMA) for IGF-I. The capture antibody did not crossreact with IGF-I associated binding proteins which exist in plasma, and the assay was designed not to shift the equilibrium of the IGF-I and binding proteins. Total IGF-I concentration was measured using this assay with preliminary acid-ethanol extraction. Approximately 1 percent of total IGF-I existed in the free form. Gel filtration of plasma was also used to separate the free IGF-I from its bound form. The free/total ratio of IGF-I as determined by gel filtration was similar to that determined directly by IRMA with and without acid-ethanol extraction.

A sensitive enzyme-linked immunosorbent assay for human proinsulin was developed by the modification of the method reported using monoclonal antibodies. In the present method, two monoclonal antibodies, an anti-C-peptide antibody bound to microtiter plate, and a biotin-labeled anti-insulin antibody were uesd. This assay was specific for proinsulin and failed to detect both insulin and C-peptide. The minimal detection limit of this assay was approximately 0.1 pmol/1. lmmunoreactive proinsulin levels in serum of normal subjects, ranged from 1.7 to 8.7 pmol/1 with the mean of 4.6 pmol/l. The ranges for the intraand inter-assay coefficients of variance were 3.1-3.7% and 5.0-14.9%, respectively. Reverse phase HPLC analysis of serum of normal subject, as measured with this assay system, revealed two immunoreactive (IR-) forms. One form eluted at the same position as that of authentic proinsulin and the other was detected in a more hydrophilic part of the chromatogram (shorter retention time). Elution profiles of IR-insulin and IR-C-peptide in human serum were also examined by the present reverse phase HPLC and compared to those of IR-proinsulins.Proinsulin, the biosynthetic precursor of insulin, is

The purpose of this study was to investigate the factors affecting the sensitivity of an immunoradiometric assay (IRMA) for human adrenocorticotropic hormone (ACTH). This method relies on the formation of an immune complex consisting of a 1251-labelled polyclonal antibody, human ACTH, and a solid phase monoclonal antibody. Several factors affecting assay performance were investigated, including preparation and purification of the 1251-labelled polyclonal antibody, standard diluent buffers, and incubation conditions. The optimized assay covered a range of 5-1,200 pg/ml ACTH. Non-specific binding (in the absence of antigen) was consistently less than 0.2% of added 1251-labelled antibody, and this was the primary factor in the detection limit. The minimal detection limit (+2.0 SD of the zero

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