Rapid screening assay of congenital adrenal hyperplasia by measuring 17 alpha-hydroxyprogesterone with high-performance liquid chromatography/electrospray ionization tandem mass spectrometry from dried blood spots.
A rapid, simple, and specific method was developed for the diagnosis of congenital adrenal hyperplasia (CAH) from dried blood spots on newborn screening cards based on high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS). The usefulness of 17 alpha-hydroxyprogesterone (17 OH-P) determination on dried filter-paper blood samples from patients with CAH caused by 21-hydroxylase deficiency was evaluated. The LC/MS/MS detection of 17 OH-P was rapid, <4 min. The intra- and interday accuracy and precision of the method were <7%. Our procedure maintained good linearities (R(2) > 0.992) and recovery rate (>83%). We used this new method to directly determine the 17 OH-P levels in dried blood specimens from abnormal children of various ages, with a detection limit of 20 ng/ml (approximately 240 pg), to avoid the time-consuming derivatization steps required by the gas-chromatography/mass spectrometry (GC/MS) method. Four dried filter-paper blood samples of CAH patients (three girls and one boy, 1-14 years old) were all quantified in an LC/MS/MS study and revealed high 17 OH-P levels (>90 ng/ml). After treatment, all of the elevated 17 OH-P levels either decreased or disappeared. Compared with CAH patients, 17 OH-P was nearly undetectable (<20 ng/ml) in the normal infants by LC/MS/MS. This LC/MS/MS assay is not only useful for both diagnosis and monitoring of treatment of CAH in all other age groups, it also can be used as a screening test for CAH infants. In this study, we provided the first data on 17 OH-P in dried blood specimens affected with CAH using HPLC/ESI-MS/MS.
Lai
Chien-Chen
CC
Department of Medical Genetics and Medical Research, China Medical College Hospital, Taichung, Taiwan, Republic of China.
Tsai
Chang-Hai
CH
Tsai
Fuu-Jen
FJ
Wu
Jer-Yuarn
JY
Lin
Wei-De
WD
Lee
Cheng-Chun
CC
D019326:17-alpha-Hydroxyprogesterone / Q000097:blood*; D000312:Adrenal Hyperplasia, Congenital / Q000175:diagnosis*; D016022:Case-Control Studies; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006801:Humans; D007231:Infant, Newborn; D011863:Radioimmunoassay; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods
D019326:17-alpha-Hydroxyprogesterone