Mass spectrometric and physiological validation of a sensitive, automated, direct immunoassay for serum estradiol using the Architect®

BackgroundMeasurement of estradiol (E2) plays a critical role in the diagnosis and clinical management of reproductive disorders. The challenge for all currently available direct methods for measuring E2 is to provide accuracy and precision across a wide dynamic range.MethodsWe describe the development and multi-site performance evaluation of a direct E2 assay on the Architect i2000®. Assay performance and method comparisons were performed by testing specimens from men, healthy women with regular menstrual cycles, and post-menopausal women using the Architect assay and isotope dilution, gas chromatography-mass spectrometry (ID/GC-MS). Reference intervals were established by testing prospectively collected daily blood draws from 42 healthy women, 72 postmenopausal women and 101 males.ResultsNo unexpected cross-reactivity or interference was observed for over 40 compounds tested. Recovery was 100 ± 10% in the presence of estrone and estriol. Functional sensitivity (%CV < 20%) was < 15 pg/ml.1 The imprecision of the assay was < 7.1% (total CV), < 2.5%, and < 2.3% for control sera containing 45, 190, and 600 pg/ml estradiol, respectively. The assay had a correlation of y = 1.033 x + 0.3156, r2 = 0.99, n = 131 compared to ID/GC-MS. Reference intervals for the current Architect Estradiol assay are reported.ConclusionsFormat changes resulted in dramatic improvement in the performance and accuracy of this direct, fully automated assay. The assay is standardized by ID/GC-MS. The assay is clinically useful for serum concentrations from 15 to > 4000 pg/ml.