Abstract
Background: The best index of adrenal dysfunction is urinary free
cortisol (UFC) measurements performed using a 24-h urine collection.
This measurement is also useful in the investigation of Cushing's
syndrome. In this paper, we report a simple and selective method for the
analysis of UFC by liquid chromatography-tandem mass spectrometry
(LC-MS/MS) suitable for use in a high-volume clinical laboratory
routine. The results were compared to those obtained using a commercial
immunoassay method used in our laboratory.
Methods: Urine samples containing 50 ng of internal standard
(Cortisol-9,11,12,12-d(4)) were deproteinized using centrifugal filters
with a molecular weight 10,000 Da cut-off and injected on a reversed
phase column. Cortisol was analyzed in highly selective reaction
monitoring in positive atmospheric pressure chemical ionization mode, at
a resolution of 0.4 amu full width half maximum, and following the
transitions related to the precursor 363.2 for cortisol and 367.2 for
deuterated cortisol. The method validation included analysis of
precision, linearity, sensitivity, recovery and interference from
structurally similar steroids. UFC from 230 subjects was measured using
LC-MS/MS and electrochemiluminescence immunoassay (ECLIA) methods.
Results: The calibration curves exhibited linearity and reproducibility
in the range 7-10,000 nmol/L. Total imprecision was lower than 10\%. The
limit of detection and limit of quantification were 2 and 7 nmol/L,
respectively. Mean recovery was higher than 90\%. Structurally similar
steroids do not interfere in the proposed method, but cause a
significant change in the ECLIA results. Cortisol values obtained using
the ECLIA method were always higher than those obtained using the
LC-MS/MS method, with the bias directly proportional to cortisol
concentrations. The reference values calculated using 180 normal
subjects were 11-70 mu g/day.
Conclusions: The proposed method is sensitive, simple, free from
interferences and reliable for routine use. Clin Chem Lab Med
2010;48:1433-7.
Lingua originale | English |
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pagine (da-a) | 1433-1437 |
Numero di pagine | 5 |
Rivista | Clinical Chemistry and Laboratory Medicine |
Volume | 48 |
DOI | |
Stato di pubblicazione | Pubblicato - 2010 |
Keywords
- mass spectrometry