TY - JOUR T1 - Development and evaluation of a candidate reference method for the determination of total cortisol in human serum using isotope dilution liquid chromatography/mass spectrometry and liquid chromatography/tandem mass spectrometry. AU - Tai,Susan S-C, AU - Welch,Michael J, PY - 2004/2/14/pubmed PY - 2004/8/6/medline PY - 2004/2/14/entrez SP - 1008 EP - 14 JF - Analytical chemistry JO - Anal Chem VL - 76 IS - 4 N2 - Cortisol is an important diagnostic marker for the production of steroid hormones, and accurate measurements of serum cortisol are necessary for proper diagnosis of adrenal function. A candidate reference method involving isotope dilution coupled with liquid chromatography/mass spectrometry (LC/MS) and liquid chromatography/tandem mass spectrometry (LC/MS/MS) has been developed and critically evaluated. An isotopically labeled internal standard, cortisol-d(3), was added to serum, followed by equilibration and solid-phase and ethyl acetate extractions to prepare samples for liquid chromatography/mass spectrometry electrospray ionization (LC/MS-ESI) and liquid chromatography/tandem mass spectrometry electrospray ionization (LC/MS/MS-ESI) analyses. (M + H)(+) ions at m/z 363 and 366 for cortisol and its labeled internal standard were monitored for LC/MS. The transitions of (M + H)(+) --> [(M + H)(+) - 2H(2)O] at m/z 363 --> 327 and 366 --> 330 were monitored for LC/MS/MS. The accuracy of the measurement was evaluated by a comparison of results of this candidate reference method on lyophilized human serum reference materials for cortisol [Certified Reference Materials 192 and 193] with the certified values determined by gas chromatography/mass spectrometry reference methods and by a recovery study for the added cortisol. The results of this method for total cortisol agreed with the certified values within 1.1%. The recovery of the added cortisol ranged from 99.8% to 101.0%. This method was applied to the determination of cortisol in samples of frozen serum pools. Excellent precision was obtained with within-set CVs of 0.3%-1.5% and between-set CVs of 0.04%-0.4% for both LC/MS and LC/MS/MS analyses. The correlation coefficients of all linear regression lines ranged from 0.998 to 1.000. The detection limits (at a signal-to-noise ratio of approximately 3-5) were 10 and 15 pg for LC/MS and LC/MS/MS, respectively. This method, which demonstrates good accuracy and precision, and is free from interferences from structural analogues, qualifies as a candidate reference method and can be used as an alternative reference method to provide an accuracy base to which the routine methods can be compared. SN - 0003-2700 UR - https://www.unboundmedicine.com/medline/citation/14961732/Development_and_evaluation_of_a_candidate_reference_method_for_the_determination_of_total_cortisol_in_human_serum_using_isotope_dilution_liquid_chromatography/mass_spectrometry_and_liquid_chromatography/tandem_mass_spectrometry_ DB - PRIME DP - Unbound Medicine ER -