Solid-phase enzymoimmunoassay of estrone in serum.

Abstract

This rapid solid-phase enzymoimmunoassay for estrone in serum or plasma is done on microtiter plates after the serum is extracted with diethyl ether. No chromatographic or centrifugation steps are involved. The detection limit of the assay is 380 fg per well (28 pmol/L). Intra- and interassay coefficients of variation for the assay of low, medium, and high concentrations of estrone in plasma were respectively 4.4, 9.3; 2.3, 9.1; and 2.0, 6.3 percent. There was a good correlation (correlation coefficient 0.95) between estrone concentrations measured with this assay and with a commercial radioimmunoassay. The analytical procedure is simple, and one person can assay 80 serum samples per working day. We conclude that the assay is very suitable for serum estrone measurements and is more convenient than published radioimmunoassays.

Synthesis of 3-Hydroxyestra-1,3,5(10)-trien-17-one and Estra-1,3,5(10)-triene-3,17β-diol 6α-N-(iε-Biotinyl)caproamide, Tracer Substances for Developing Immunoassays for Estrone and Estradiol

“…Most of the competitive assays use either enzyme-or substratelabeled steroid tracers. These assays often lack analytical sensitivity and specificity due to an altered structure of the respective steroid tracers (5)(6)(7)(8)(9)(10). This effect is observed in cases in which steroids have been derivatized at the ring positions 3 or 17.…”

Synthesis of 17β-Hydroxyandrost-4-en-3-one−7α-(Biotinyl-6-N-hexylamide), a Conjugate Useful for Affinity Chromatography and for Testosterone Immunoassays

“…The absolute configurations of the separated products 17β-hydroxyandrosta-4,6-dien-3-one 17β-acetate-7R/β-(6tert-butyldimethylsiloxyhexyl) 2a/2b were determined by 1 H NMR measurements and nuclear Overhauser effect (NOE) experiments. These experiments established the chemical shifts and the coupling pattern of the AB part of the ABX system of the 6R-H, 6β-H, and 7-H.…”

“…IR (neat) (cm -1 ): 1740 (17-OAc), 1675 (3-one), 1620 (olefin), 1100, 835, 775 (Si-R). 1 ) +26°(MeOH, c ) 3.24). 17β-Hydroxyandrost-4-en-3-one 17β-Acetate-7R-(6-Hexanal) (4a).…”

“…Melting points (uncorrected) were determined on a Reichert thermovar. 1 H NMR spectra were recorded in CDCl 3 on a Bruker ARX 400 (400 MHz), using Me 4 Si for the reference signal (δ 0.00 ppm). The data are reported in the following form: chemical shift (multiplicity, coupling constant in Hz if available, number of protons, position of proton).…”

“…In the case of the determination of steroids in human serum, analytical sensitivity and specificity of such competitive enzyme immunoassays may be severely limited when steroids have been derivatized at ring positions 3 or 17 ( − ), leading to altered epitopes of the respective tracers. The easily derivatized functional groups at position 3 and 17 are the groups that define the identity of testosterone.…”