Noncompetitive ELISA for human serum insulin-like growth factor-I.
Measurement of serum insulin-like growth factor I (IGF-I) is important in assessing the growth hormone/IGF axis. Competitive immunoassay methods for IGF-I are complicated by substantial interference from IGF-binding proteins (IGFBPs) due to the relatively low ratio of specific antibody to IGFBPs in the sample, even after standard acid-ethanol sample extraction. We report of development of a noncompetitive ELISA for human IGF-I that avoids this problem by utilizing excess amounts of capture and detection antibodies. Serum samples were prepared by using an abbreviated acid-ethanol extraction method. Neutralized supernatant was added to microwells coated with IGF-I capture antibody; horseradish peroxidase-labeled detection antibody was then added, incubated for 2 h, and then developed. Compared with the "gold standard" method of acid-column chromatography, the simplified acid-ethanol extraction yields a mean +/- SD recovery of 103% +/- 5.5% despite the presence of residual IGFBPs in the extracted sample. Comparisons with a centrifugal filtration sample extraction method are also shown. The ELISA is specific for IGF-I with an absolute sensitivity of 0.03 microgram/L and inter- and intraassay CVs of 3.9-8.8% and 2.6-6.7%, respectively. The availability of a rapid IGF-I ELISA combined with a simple and reliable sample preparation procedure should facilitate clinical and research studies of this important growth factor.
Khosravi
M J
MJ
Diagnostic Systems Laboratories (Canada) Inc., Toronto, ON, Canada.
Diamandi
A
A
Mistry
J
J
Lee
P D
PD
D002498:Centrifugation; D002845:Chromatography; D002851:Chromatography, High Pressure Liquid; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000706:statistics & numerical data; D000431:Ethanol; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007231:Infant, Newborn; D018970:Insulin-Like Growth Factor Binding Protein 1 / Q000097:blood; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000097:blood; D007334:Insulin-Like Growth Factor I / Q000032:analysis*; D008970:Molecular Weight; D011247:Pregnancy; D012680:Sensitivity and Specificity
D018970:Insulin-Like Growth Factor Binding Protein 1; D018972:Insulin-Like Growth Factor Binding Protein 3; D000431:Ethanol; D007334:Insulin-Like Growth Factor I