Simultaneous determination of androstenedione and testosterone in human serum by liquid chromatography-tandem mass spectrometry.
BACKGROUND
We aimed to develop a sensitive assay to quantitate serum concentrations of both androstenedione and testosterone within the female range simultaneously, using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for use in the routine clinical laboratory and to compare this method with immunoassay.


METHODS
Samples (200 microL) were prepared by liquid-liquid extraction using (1 mL) methyl-tert-butyl-ether. Deuterated androstenedione and testosterone were used as internal standards.


RESULTS
The standard curve was linear to 50 nmol/L, the lower limit of quantitation was 0.25 nmol/L, and intra- and inter-assay coefficients of variation were < 10% for both androgens over the range 0.3-35 nmol/L. There was a poor relationship between the LC-MS/MS and the radioimmunoassay methods for androstenedione with the LC-MS/MS generally giving lower results. For testosterone, the LC-MS/MS and immunoassay methods compared well at all concentrations. However, when female samples only were examined, the agreement deteriorated.


CONCLUSIONS
We have developed a sensitive and precise LC-MS/MS method, which gives more accurate results for all androstenedione measurements and low testosterone concentrations than immunoassay.
Gallagher
L M
LM
Department of Clinical Biochemistry, South Manchester University Hospitals NHS Trust, Manchester M23 9LT, UK.
Owen
L J
LJ
Keevil
B G
BG
D000735:Androstenedione / Q000097:blood*; D002138:Calibration; D002853:Chromatography, Liquid; D005260:Female; D006801:Humans; D012015:Reference Standards; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry; D013739:Testosterone / Q000097:blood*
D013739:Testosterone; D000735:Androstenedione