title: Concurrent Confirmation and Differential Diagnosis of Congenital Adrenal Hyperplasia from Dried Blood Spots: Application of a Second-Tier LC-MS/MS Assay in a Cross-Border Cooperation for Newborn Screening.
abstract: OBJECTIVE
  Newborn screening for congenital adrenal hyperplasia (CAH) is generally performed using 17- hydroxyprogesterone dissociation-enhanced, lanthanide fluorescence immunoassay (DELFIA®). The primary screening results must be confirmed due to high false-positive rates; however, the need to obtain a separate specimen can hamper early recognition, differential diagnosis and treatment. We aimed to develop a single liquid chromatography-tandem mass spectrometry (LC-MS/MS) method that allows both the confirmation and differential diagnosis of CAH using the same dried blood spot (DBS) as in primary screening.
  
  
  METHODS
  An LC-MS/MS assay for cortisol, 21-deoxycortisol, 11-deoxycortisol, 4-androstenedione and 17-hydroxyprogesterone was developed, validated and applied to a total of 163 DBS samples tested positive in primary newborn screening in a cross-border cooperation.
  
  
  RESULTS
  Excellent baseline resolution and reliable determination of all analytes were achieved in DBS samples following simple sample preparation without derivatization. Of a total of 163 DBS samples tested positive in primary screening, the 21-hydroxylase-deficient form of CAH was confirmed in 1 sample.
  
  
  CONCLUSIONS
  The present LC-MS/MS assay was successfully applied as a second-tier test in a cross-border cooperation for newborn screening. The assay allows concurrent confirmation and differential diagnosis of CAH and can be performed on the same DBS samples as in primary screening, enabling early diagnosis and treatment.
authors.affiliation: Department of Paediatrics, University of Szeged, Szeged, Hungary.
authors.forename: Péter
authors.initials: P
authors.lastname: Monostori
authors.forename: Pál
authors.initials: P
authors.lastname: Szabó
authors.forename: Otilia
authors.initials: O
authors.lastname: Marginean
authors.forename: Csaba
authors.initials: C
authors.lastname: Bereczki
authors.forename: Eszter
authors.initials: E
authors.lastname: Karg
mesh_terms: D000312:Adrenal Hyperplasia, Congenital / Q000097:blood* / Q000175:diagnosis* / Q000191:economics; D002851:Chromatography, High Pressure Liquid; D059788:Dried Blood Spot Testing / Q000191:economics / Q000379:methods*; D005189:False Positive Reactions; D005260:Female; D006801:Humans; D006814:Hungary; D007231:Infant, Newborn; D007391:International Cooperation; D008297:Male; D013058:Mass Spectrometry; D015997:Neonatal Screening / Q000191:economics / Q000379:methods*; D011786:Quality Control; D012015:Reference Standards; D015203:Reproducibility of Results; D012383:Romania; D013255:Steroid 21-Hydroxylase / Q000097:blood
chemical_list: D013255:Steroid 21-Hydroxylase