Measurement of dimeric inhibin in porcine serum: evidence for low concentrations and existence of binding proteins.
An immunoradiometric assay and serum extraction procedure were developed to measure dimeric inhibin in porcine serum with minimal interference by putative inhibin-binding proteins. Assay sensitivity was 50 pg/tube, and it incorporated antibodies against the N-terminal region of inhibin's alpha-subunit, alpha-(1-25)-Ab, and against the C-terminal region of inhibin's beta A-subunit. To determine whether inhibin-binding proteins were present in porcine serum, serum was incubated with [125I]-recombinant human (rh)-inhibin and then chromatographed by gel filtration. Radioiodinated rh-inhibin was associated with protein(s) > 600 kDa. Radioiodinated rh-inhibin also was incubated with alpha 2-macroglobulin, an inhibin-binding protein in human and rat serum. Elution profiles were similar for serum and alpha 2-macroglobulin. Serum- and alpha 2-macroglobulin-[125I]rh-inhibin complexes dissociated upon exposure to 8 M urea. Porcine serum was treated with urea, after which inhibin was isolated and concentrated. The recovery of rh-inhibin added to starting serum was 28%. Concentrations of endogenous dimeric inhibin were < 28 pg/ml in serum collected from sows at random stages of the estrous cycle and were < 21 pg/ml in serum collected from sows 2 d postweaning. Results demonstrate that 1) concentrations of dimeric inhibin are low in porcine serum, and 2) an inhibin-binding protein(s), consistent with alpha 2-macroglobulin, is present in porcine serum.
Meyer
R L
RL
Department of Animal Science, University of Minnesota, St. Paul 55108, USA.
Wheaton
J E
JE
D000818:Animals; D002352:Carrier Proteins / Q000097:blood*; D006801:Humans; D015592:Immunoradiometric Assay; D007265:Inhibins / Q000097:blood* / Q000302:isolation & purification; D007457:Iodine Radioisotopes; D046911:Macromolecular Substances; D011994:Recombinant Proteins / Q000097:blood; D013552:Swine / Q000097:blood*; D000511:alpha-Macroglobulins / Q000378:metabolism
D002352:Carrier Proteins; D007457:Iodine Radioisotopes; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D000511:alpha-Macroglobulins; D007265:Inhibins