Measurement of plasma cortisol was performed using Siemens Atellica IM Cortisol kit (intra-assay variation coefficient ≤ 2.3% for plasma, inter-assay variation coefficient ≤ 7.7% for plasma). Reported cross-reactivity with steroid precursors: 11-deoxycortisol ≤ 25%, 21-deoxycortisol ≤ 15%, 17α-hydroxyprogesterone ≤ 2%, pregnenolone ≤ 0,5%, and progesterone ≤ 1%.
The 24 h UFC was measured by a Beckman Coulter Cortisol RIA kit (intra-assay variation coefficient ≤ 8.9% for urine, inter-assay variation coefficient ≤ 13.3% for urine). Blood samples for ACTH measurement were kept on ice, temporarily frozen at −20°C and analysed with a Thermo Scientific BRAHMS ACTH RIA kit (intra-assay variation coefficient 3.5%, inter-assay variation coefficient 4.7%).
Plasma steroids including cortisol were measured by two-dimensional liquid chromatography–tandem mass spectrometry (Agilent technologies 1260 Infinity II, Agilent 6470 triple quadrupole) using pre-mixed sets of steroids and their internal standards (Chromsystems Instruments & Chemicals GmbH) with in-house developed method for a multiplex quantitative analysis of steroid hormones (3 (link)).
The 24 h UFC was measured by a Beckman Coulter Cortisol RIA kit (intra-assay variation coefficient ≤ 8.9% for urine, inter-assay variation coefficient ≤ 13.3% for urine). Blood samples for ACTH measurement were kept on ice, temporarily frozen at −20°C and analysed with a Thermo Scientific BRAHMS ACTH RIA kit (intra-assay variation coefficient 3.5%, inter-assay variation coefficient 4.7%).
Plasma steroids including cortisol were measured by two-dimensional liquid chromatography–tandem mass spectrometry (Agilent technologies 1260 Infinity II, Agilent 6470 triple quadrupole) using pre-mixed sets of steroids and their internal standards (Chromsystems Instruments & Chemicals GmbH) with in-house developed method for a multiplex quantitative analysis of steroid hormones (3 (link)).
