Storage stability of five steroids and in dried blood spots for newborn screening and retrospective diagnosis of congenital adrenal hyperplasia

PLoS One. 2020 May 29;15(5):e0233724. doi: 10.1371/journal.pone.0233724. eCollection 2020.

Abstract

Congenital adrenal hyperplasia (CAH) is a severe inherited disorder of cortisol biosynthesis that is potentially lethal or can seriously affect quality of life. For the first time, we aimed to assess the stability of 21-deoxycortisol (21Deox), 11-deoxycortisol (11Deox), 4-androstenedione (4AD), 17-hydroxyprogesterone (17OHP) and cortisol (Cort), diagnostic for CAH, in dried blood spots (DBSs) during a 1 year storage at different temperatures. Spiked DBS samples were stored at room temperature, 4 °C, -20 °C or -70 °C, respectively and analyzed in triplicates using liquid chromatography-tandem mass spectrometry at Weeks 0, 1, 2, 3 and 4, Month 6 and Year 1. Analyte levels within ±15% vs the baseline were considered stable. Our observations show that 21Deox, 4AD and 17OHP were not significantly changed for 1 year even at room temperature at either analyte levels. In contrast, Cort required storage at 4 °C, -20 °C or -70 °C for long-term stability, being significantly decreased at room temperature from Month 6 (p<0.01) in both the 30(60) nM and the 90(180) nM samples. 11Deox was significantly decreased at room temperature at Year 1 (p<0.01) and only in the 30(60) nM samples. Thus, all biomarkers were stable for up to 1 year at 4 °C, -20 °C or -70 °C and at least for 4 weeks at room temperature. These findings have implications for analyses of stored DBS samples in 2nd-tier assays in newborn screening and for retrospective CAH studies.

Publication types

  • Clinical Trial
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adrenal Hyperplasia, Congenital / blood*
  • Adrenal Hyperplasia, Congenital / diagnosis
  • Androstenediol / blood*
  • Dried Blood Spot Testing*
  • Female
  • Humans
  • Infant, Newborn
  • Male
  • Mass Screening*
  • Pregnenediones / blood*
  • Preservation, Biological*

Substances

  • Pregnenediones
  • Androstenediol

Grants and funding

This work was supported by the Hungarian-Romanian Cross Border Cooperation Project (HU RO) 0802/008 SCREENGEN. Article Processing Charges were covered by the Institutional Account Program of the University of Szeged, Szeged, Hungary (Open Access Fund, Grant number: 4581). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.