For cortisol analyses serum was gained by centrifugation of blood (15 min, 15°C, 1950× g, Varifuge 3.0, Heraeus, Hanau, Germany) after incubation for 30 min at room temperature and 30 min at 30°C and then stored at -80°C until measurement. The concentration of serum cortisol was measured at selected sampling points (-42 d, -14 d, -7 d, 0.5 h, 2 h, 4 h, 6 h,12 h, 24 h, 48 h, 7 d, 14 d, 21 d, 28 d, 42 d, 56 d, 100 d and 110 d) by radioimmunoassay according to the manufacturer’s protocol (Cortisol RIA Kit, Beckman Coulter, Krefeld, Germany). In brief, 50 µl of the sample or the control were incubated for 1 h at room temperature with 500 µl of 125I-labeld cortisol tracer or for the total count only with 500 µl tracer. Afterwards, bound and total cortisol were determined, and concentrations calculated with a standard curve.
Cortisol ria kit
Manufactured by Beckman Coulter
Sourced in Germany
The Cortisol RIA Kit is a radioimmunoassay (RIA) kit used for the quantitative determination of cortisol levels in human serum, plasma, or urine samples. The kit utilizes the competitive binding principle, where cortisol in the sample competes with a known quantity of radiolabeled cortisol for binding to a limited number of antibody binding sites. The amount of bound radiolabeled cortisol is inversely proportional to the concentration of cortisol in the sample.
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Lab products found in correlation
4 protocols using cortisol ria kit
1
Serum Cortisol Measurement Protocol
2
Biomarkers of Stress and Inflammation
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Fasting blood samples were taken in the morning. Subjects were instructed to restrain from exercise during the day before their examination. Venous blood samples were collected in suitable vacutainers, tubes containing potassium ethylenediaminetetra-acetic acid (K-EDTA) were used for testing cellular blood parameters. Tubes without additives were used to obtain serum for C-reactive protein (CRP) and cortisol tests. After blood collection serum was separated by centrifugation (10 min, room temperature, 1500 rcf). Blood cell parameters were quantified in a multi-parameter automatic hematology analyzer Cell-Dyn 3700 system (Abbott Diagnostics, Abbott Laboratories, Abbott Park, IL, United States). CRP was measured by Cardiac C-Reactive Protein (Latex) High Sensitive turbidimetric immunoassay (Roche Diagnostics) on Cobas 8000 Modular Analyzer (Roche Diagnostics, GmbH, Mannheim, Germany) following the manufacturer’s instructions. Cortisol was measured by Cortisol RIA Kit (Beckman Coulter, Cat.: IM1841) on RIA-mat 280 automated analyzer (Stratec Gmbh, Birkenfeld, Germany) following the manufacturers’ instructions.
3
Plasma Cortisol, UFC, and ACTH Measurement
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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)).
4
Hormonal and Renal Function Analysis
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Plasma and urine samples were stored at 20 °C until analysis. Plasma renin concentration (PRC) was measured using an immunoradiometric assay (Renin III, CisBio, Codolet, France). Plasma aldosterone concentration (PAC) and 24-h urinary excretion of free aldosterone at pH1 (UAE) were measured using a radioimmunoassay (Coat-a-count, Siemens Medical Solutions Diagnostics, Erlangen, Germany). 24-h urinary free cortisol Excretion (UFCE) was measured by radioimmunoassay (Cortisol RIA kit, Beckman Coulter). The ratio of UAE and UFCE to urinary creatinine excretion was used to standardize the 24hour urine collection. Glomerular filtration rate (GFR) was estimated with the simplified modification of diet in renal disease (MDRD) formula. To account for differences in diet between genotypes and visits, if any, 24-hour urinary sodium and potassium excretion were measured. All biochemical and hormonal assessments were performed blind to the genotype.
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