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Development and Evaluation of a Reference Measurement Procedure for the Determination of Estradiol-17 in Human Serum Using Isotope-Dilution Liquid Chromatography-Tandem Mass Spectrometry

Published

Author(s)

Susan S. Tai, Michael J. Welch

Abstract

Estradiol is the most potent natural estrogen and is derived from the ovaries. Its concentration in blood is measured to determine ovarian function. A reference measurement procedure (RMP) for estradiol in serum involving isotope-dilution (ID) coupled with liquid chromatography-tandem mass spectrometry (LC/MS/MS) has been developed and critically evaluated. A deuterated estradiol (estradiol-d3) was used as an internal standard. The estradiol and its internal standard were extracted from serum matrix using solid-phase extractions, and derivatized with dansyl chloride prior to reversed-phase LC/MS/MS. The accuracy of the measurement was evaluated by a comparison of results of this reference method on lyophilized human serum reference materials for estradiol [Certified Reference Materials (CRMs) 576, 577, and 578] with the certified values determined by gas chromatography/mass spectrometry (GC/MS) reference methods, and by a recovery study for the added estradiol. The results of this method for estradiol were within the certified values of CRMs with a mean difference of 1.1% for the three CRMs. The recovery of the added estradiol ranged from 100.7% to 101.8%. This method was applied to the determination of estradiol in frozen serum samples from three single-donors. Excellent reproducibility was obtained with within-set CVs ranging from 0.6% to 2.2%, and between-set CVs ranging from 0.2% to 0.4%. Excellent linearity was also obtained with correlation coefficients of all linear regression lines (measured intensity ratios vs. mass ratios) ranging from 0.998 to 1.000. The detection limit at a signal-to-noise ratio of ~3 was 0.6 pg of estradiol. This well-characterized LC/MS/MS method for serum estradiol, which demonstrates good accuracy and precision, low susceptibility to interferences, and comparability with GC/MS reference methods, qualifies as a reference measurement procedure that can be used to provide an accuracy base to which routine methods for estradiol can be compared and that will serve as a standard of higher order for measurement traceability.
Citation
Journal of Analytical Chemistry
Volume
77 No. 19

Keywords

electrospray ionization, Estradiol-17beta, isotope dilution, LC/MS/MS, liquid chromatography, reference measurement procedure, solid-phase extraction, tandem mass spectrometry

Citation

Tai, S. and Welch, M. (2005), Development and Evaluation of a Reference Measurement Procedure for the Determination of Estradiol-17 in Human Serum Using Isotope-Dilution Liquid Chromatography-Tandem Mass Spectrometry, Journal of Analytical Chemistry (Accessed April 17, 2024)
Created October 1, 2005, Updated February 17, 2017