Rapid screening assay of congenital adrenal hyperplasia by measuring 17α‐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 17α‐hydroxyprogesterone (17OH‐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 17OH‐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 17OH‐P levels in dried blood specimens from abnormal children of various ages, with a detection limit of 20 ng/ml (∼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 17OH‐P levels (>90 ng/ml). After treatment, all of the elevated 17OH‐P levels either decreased or disappeared. Compared with CAH patients, 17OH‐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 17OH‐P in dried blood specimens affected with CAH using HPLC/ESI‐MS/MS. J. Clin. Lab. Anal. 16:20–25, 2002. © 2002 Wiley‐Liss, Inc.