A simple, rapid immunometric assay for determination of functional and growth hormone-occupied growth hormone-binding protein in human serum.
We present a sensitive time-resolved fluorometric immunofunctional assay (TR-FIA) for direct quantitation of functional growth hormone-binding protein (GHBP), using an immunoassay kit for growth hormone (GH-DELFIA). In addition to the immobilized GH antibody, one monoclonal antibody against GHBP was used. This anti-GHBP was labelled with the chelate of europium. The assay was performed in one step. The detection limit for GHBP was 0.044 nmol L-1 (NBS + 3 SD). The calibration curve was linear in the interval 0.11-8.03 nmol L-1. Average intra-assay coefficient of variation (CV) was 3.44%. Average interassay CV at GHBP concentrations 0.563 nmol L-1 and 1.40 nmol L-1 were 12% and 6.3% respectively. Analytical recovery in serum ranged from 76% to 127% with a mean of 101 +/- 3.6%. Serum GHBP in 102 normal subjects ranged from 0.513 to 3.772 nmol L-1 and was positively related to body mass index (P < 0.001). In growth hormone-deficient sera GHBP was higher than in control subjects (1.751 +/- 0.179 nmol L-1 and 1.257 +/- 0.140 nmol L-1 respectively, P < 0.001). Acromegalic patients had lower levels of GHBP than controls (0.946 +/- 0.251 and 1.234 +/- 0.144 nmol L-1 respectively, P = 0.005). This assay also allowed detection of GH-complexed GHBP in serum. These results were in agreement with theoretical values calculated from the measured GH and the functional GHBP concentrations. Results were compared with data obtained by a recently reported, validated ligand immunofunctional assay (LIFA), which is fundamentally different. There was a significant linear relationship between the results from the two assays (r = 0.89, P = 0.001). The slope of the regression line was 0.65. In conclusion, this new convenient GHBP TR-FIA provides a sensitive and precise method for detecting total GHBP as well as complexed GHBP in human serum, and allows easy processing of large numbers of samples.
Department of Endocrinology and Diabetes, University Hospital of Aarhus, Denmark.
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Fisker
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Frystyk
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Skriver
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Vestbo
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Ho
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H
Orskov
D000172:Acromegaly / Q000097:blood; D000293:Adolescent; D000328:Adult; D000368:Aged; D002352:Carrier Proteins / Q000097:blood* / Q000378:metabolism; D002648:Child; D005260:Female; D005470:Fluorometry / Q000379:methods*; D019382:Human Growth Hormone / Q000097:blood* / Q000172:deficiency / Q000378:metabolism; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D008297:Male; D008875:Middle Aged; D011485:Protein Binding; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D002352:Carrier Proteins; D019382:Human Growth Hormone; C051815:somatotropin-binding protein