Clinical Chemistry
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Clinical Chemistry 36: 1774-1778, 1990;
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Clinical Chemistry, Vol 36, 1774-1778, Copyright © 1990 by American Association for Clinical Chemistry

Time-resolved fluoroimmunoassay for unconjugated estrogen in urine: comparison with a fluorometric assay for total estrogen and application in an in vitro fertilization program

CM Thomas, RJ van den Berg, MF Segers, LM Geelen, JM Hollanders, WH Doesburg and PC Houx
Department of Obstetrics & Gynaecology, Sint-Radboud University Hospital, Nijmegen, The Netherlands.

A time-resolved fluoroimmunoassay (TR-FIA) for unconjugated estrogens in human urine is described. 6-Keto-17 beta-estradiol-6-(O- carboxymethyl)oxime:bovine serum albumin is immobilized onto microtiter strip wells and the coated wells are incubated with 17 beta-estradiol standard preparations or unknowns with a polyclonal antiserum to 17 beta-estradiol-16,17-monosuccinyl:albumin. The antiserum-bound estrogen is detected by incubation with a europium-labeled anti-rabbit IgG that serves as both second antibody and tracer. After the immunoreactions, the bound portion of the labeled antiserum is quantified by dissociating the Eu3+ in a fluorescence-enhancement solution and measuring its fluorescence with a time-resolved fluorometer. The detection limit of the TR-FIA is 24 pmol of 17 beta-estradiol per liter; the analytical range extends to 1.8 nmol/L. This assay is a convenient alternative to radioimmunoassay and to the automated Kober- Ittrich fluorometry of total estrogen. Its advantages include short counting times; use of nonradioactive, stable reagents, all of which are commercially available; and more nearly complete automation. We conclude that this TR-FIA, compared with the Kober-Ittrich fluorometric assay (J Endocrinol 1957; 16:49-56), provides the clinician with equivalent information during follicular development therapy as part of an in vitro fertilization program.




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Copyright © 1990 by the American Association for Clinical Chemistry.