Antibody against acid-stable insulin-like growth factor binding protein detects 150,000 mol wt growth hormone-dependent complex in human plasma.
In order to measure the 150,000 mol wt binding complex for insulin-like growth factors in human plasma without the need for acidification to dissociate endogenous ligands, we have purified an acid-stable plasma binding protein (BP) and raised a specific rabbit antiserum against it. When used in RIA, with a covalent complex of binding protein and radioiodinated insulin-like growth factor-I as tracer, pure BP and human plasma samples gave parallel displacement curves. Incubation of plasma with pure insulin-like growth factor-I or -II (5 micrograms/ml) had no effect on the immunoreactivity of the samples. Compared to purified BP standard, the immunoreactive BP content of plasma from normal adults was 9.20 +/- 1.59 micrograms/ml (mean +/- SD, n = 10), from acromegalic subjects, 22.5 +/- 3.63 micrograms/ml (n = 10), and from GH-deficient subjects, 4.10 +/- 1.59 micrograms/ml (n = 7). Although this antibody reacted with a protein of approximately 60,000 mol wt in acidified plasma, gel chromatography of plasma at pH 7 indicated that a GH-dependent protein of mol wt 150,000 was measured almost exclusively in unacidified samples. Direct binding of insulin-like growth factor-II, however, showed the major peak of binding activity in normal plasma in the 40-45,000 mol wt region, whereas in acromegalic plasma most ligand binding was in the 150,000 mol wt region. It is concluded that there is essentially no free immunoreactive acid-stable BP in native plasma, that the unoccupied binding sites of 40-45,000 mol wt, detectable by ligand binding, are present on an immunologically distinct protein, and that the immunoreactive 150,000 mol wt binding complex in native plasma is strongly GH-dependent.
J L
JL
Martin
R C
RC
Baxter
D000906:Antibodies / Q000032:analysis*; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000097:blood / Q000276:immunology*; D002845:Chromatography / Q000379:methods; D013006:Growth Hormone / Q000502:physiology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D018969:Insulin-Like Growth Factor Binding Proteins; D008970:Molecular Weight; D011863:Radioimmunoassay
D000906:Antibodies; D002352:Carrier Proteins; D018969:Insulin-Like Growth Factor Binding Proteins; D013006:Growth Hormone