Analysis of 17 α-hydroxyprogesterone in bloodspots by liquid chromatography tandem mass spectrometry.
BACKGROUND
Monitoring of treatment for patients diagnosed with congenital adrenal hyperplasia (CAH) can be performed by measuring the concentration of 17α-hydroxyprogesterone (17OHP) in bloodspots collected on filter papers. A method is described here for measuring 17OHP by liquid chromatography tandem mass spectrometry (LCMSMS).


METHODS
17OHP was extracted by liquid-liquid extraction and analysed by LCMSMS. The method was validated for sensitivity, specificity, linearity, recovery, ion suppression, precision and bias.


RESULTS
The standard curve was linear from 0 to 400 nmol/L. Intra-assay %CVs were <10 and inter-assay %CVs were <15 over the range 10-200 nmol/L. Limit of quantitation was 6 nmol/L. No ion suppression was detected. The only interfering compound detected was deoxycorticosterone, an intermediate steroid with the same molecular weight as 17α-hydroxyprogesterone. The method was more accurate and precise than an existing radioimmunoassay. There was poor correlation between the two assays.


CONCLUSIONS
We have developed a sensitive and specific assay suitable for quantitation of 17OHP in bloodspots. This method performs better than radioimmunoassay and allows smaller samples to be used.
Royal Free London NHS Foundation Trust, London, UK.
Simon J
SJ
Salter
University Hospital Southampton NHS Foundation Trust, Southampton, UK.
Paul
P
Cook
University Hospital Southampton NHS Foundation Trust, Southampton, UK.
Justin H
JH
Davies
University Hospital Southampton NHS Foundation Trust, Southampton, UK Annie.armston@uhs.nhs.uk.
Annie E
AE
Armston
D019326:17-alpha-Hydroxyprogesterone / Q000097:blood*; D000312:Adrenal Hyperplasia, Congenital / Q000097:blood / Q000175:diagnosis*; D015982:Bias; D002853:Chromatography, Liquid / Q000592:standards*; D003900:Desoxycorticosterone / Q000097:blood; D059788:Dried Blood Spot Testing; D006801:Humans; D059625:Liquid-Liquid Extraction; D011863:Radioimmunoassay; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000592:standards*
D003900:Desoxycorticosterone; D019326:17-alpha-Hydroxyprogesterone
Steroid hormones; endocrinology; mass spectrometry