Simultaneous Measurement of Serum Testosterone and Dihydrotestosterone by Liquid Chromatography-Tandem Mass Spectrometry

Background: Recent reports have described inherent problems with androgen immunoassays compared with mass spectrometry analyses. Methods: We developed a method for measuring serum testosterone (T) and 5α-dihydrotestosterone (DHT) simultaneously via liquid–liquid extraction followed by liquid chromat...

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Veröffentlicht in:Clinical chemistry (Baltimore, Md.) Jg. 54; H. 11; S. 1855 - 1863
Hauptverfasser: Shiraishi, Steve, Lee, Paul W. N, Leung, Andrew, Goh, Victor H. H, Swerdloff, Ronald S, Wang, Christina
Format: Journal Article
Sprache:Englisch
Veröffentlicht: Washington, DC Am Assoc Clin Chem 01.11.2008
American Association for Clinical Chemistry
Oxford University Press
Schlagworte:
Adult
Analytical, structural and metabolic biochemistry
Androgens
Aqueous solutions
Biological and medical sciences
Biomedical research
Calibration
Chromatography, Liquid - methods
Dihydrotestosterone - blood
Female
Females
Fundamental and applied biological sciences. Psychology
Humans
Immunoassays
Investigative techniques, diagnostic techniques (general aspects)
Ionization
Liquid chromatography
Male
Mass spectrometry
Medical sciences
Methods
Middle Aged
Physical examinations
Radioimmunoassay
Reference Values
Research centers
Sensitivity and Specificity
Spectrometry, Mass, Electrospray Ionization
Tandem Mass Spectrometry - methods
Testosterone
Testosterone - blood
Validation studies
Testosterone
Dihydrotestosterone
Androgen
Mass spectrometry MS/MS
Clinical biology
Simultaneous measurement
Serum
Biochemistry
Liquid chromatography
Testicular hormone
Molecular biology
Sex steroid hormone
ISSN:0009-9147, 1530-8561, 1530-8561
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Zusammenfassung:Background: Recent reports have described inherent problems with androgen immunoassays compared with mass spectrometry analyses. Methods: We developed a method for measuring serum testosterone (T) and 5α-dihydrotestosterone (DHT) simultaneously via liquid–liquid extraction followed by liquid chromatography–tandem mass spectrometry (LC-MS/MS) with positive-mode electrospray ionization. Results: The DHT and T calibrators showed a linear response from 0.069 nmol/L to 34.4 nmol/L and 69.3 nmol/L, respectively. T interference in the DHT assay and vice versa were negligible. Within- and between-run imprecision values were <5% for both analytes. Percent recoveries of T and DHT spiked into samples at concentrations spanning the calibration curve were 100%–113% and 98%–107%, respectively. The lower limit of quantification was 0.069 nmol/L for both steroids. Serum T concentrations measured by LC-MS/MS were different from those obtained by RIA, especially at lower T concentrations. Serum DHT concentrations measured by LC-MS/MS were markedly lower than those generated by RIA because of the nonselectivity of the RIA without chromatography. The reference intervals (mean ± 2 SDs) determined for T and DHT were 9.2–33.7 nmol/L and 0.47–2.65 nmol/L, respectively, for 113 healthy adult men and 0.33–2.02 nmol/L and 0.09–0.91 nmol/L, respectively, for 133 healthy premenopausal women. Conclusions: We have developed and validated a selective and precise method for simultaneous measurements of serum T and DHT that can be adopted for routine measurements of these androgens in health and disease in men and women.
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ISSN:0009-9147
1530-8561
1530-8561
DOI:10.1373/clinchem.2008.103846