title: Measurement of amniotic fluid steroids of midgestation via LC-MS/MS.
abstract: BACKGROUND
  Analysis of steroids by mass spectrometry (MS) has evolved into a reliable tool for the simultaneous detection of multiple steroids. As amniotic fluid (AF) and fetal serum composition of early pregnancy are closely related, the analysis of AF can yield information on the physiological status of the developing fetus. We evaluated the use of liquid-chromatography tandem mass spectrometry (LC-MS/MS) for AF steroid analysis, including the analysis of its sensitivity and accuracy for gender verification in healthy subjects.
  
  
  METHODS
  AF of 78 male and 94 female healthy newborns was analyzed by LC-MS/MS at 16 weeks of gestation. The levels of androstenedione, corticosterone, cortisol, cortisone, deoxycorticosterone, 11-deoxycortisol, dehydroepiandrosterone (DHEA), dehydroepiandrosterone-sulfate (DHEA-S), 17-hydroxyprogesterone, progesterone (17-OHP) and testosterone were measured. Steroid levels were compared to RIA and GC-MS levels of midgestation from the literature. Cross-validated logistic regression was used to obtain statistical predictions of gender at birth from testosterone and the above steroids.
  
  
  RESULTS
  LC-MS/MS analysis of AF steroids yielded comparable results with published GC-MS data. Gender specific differences were found for androstenedione and testosterone concentrations with higher levels in the male fetus. In contrast to published RIA data no gender specific differences were observed for 17-hydroxyprogesterone and dehydroepiandrosterone AF concentrations. Testosterone concentrations yielded highly accurate predictions for male gender at birth. Additional analysis of further steroids did neither increase the accuracy, sensitivity nor specificity of this prediction. The estimated optimal cut-off value for amniotic testosterone level was 0.074 μg/L for healthy male newborns.
  
  
  CONCLUSIONS
  LC-MS/MS is a reliable method for the determination of steroids in amniotic fluid. The determination of testosterone in amniotic fluid by LC-MS/MS in early pregnancy of healthy subjects can be used to offer a reliable prediction of fetal gender at birth.
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University of Erlangen-Nürnberg, Erlangen, Germany. Electronic address: fabian.fahlbusch@uk-erlangen.de.
authors.forename: Fabian B
authors.initials: FB
authors.lastname: Fahlbusch
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University of Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Heussner
authors.affiliation: Institute of Medical Biometry, Informatics and Epidemiology (IMBIE), Rheinische Friedrich-Wilhelms-University, Bonn, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Schmid
authors.affiliation: Department of Obstetrics and Perinatal Medicine, Diakoniekrankenhaus Friederikenstift, Hannover, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Schild
authors.affiliation: Department of Gynecology and Obstetrics, University of Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Ruebner
authors.affiliation: Department of Gynecology and Obstetrics, University of Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Hanna
authors.initials: H
authors.lastname: Huebner
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University of Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Rascher
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University of Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Helmuth-Guenther
authors.initials: HG
authors.lastname: Doerr
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University of Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Manfred
authors.initials: M
authors.lastname: Rauh
mesh_terms: D000305:Adrenal Cortex Hormones / Q000032:analysis*; D000328:Adult; D000653:Amniotic Fluid / Q000737:chemistry*; D002853:Chromatography, Liquid; D005260:Female; D005312:Fetal Blood / Q000737:chemistry; D006801:Humans; D007231:Infant, Newborn; D008297:Male; D011247:Pregnancy; D013256:Steroids / Q000032:analysis*; D053719:Tandem Mass Spectrometry
chemical_list: D000305:Adrenal Cortex Hormones; D013256:Steroids
keywords: Amniocentesis; Amniotic fluid; Gender; LC–MS/MS; Steroid profile