title: Development of a rapid assay for the analysis of serum cortisol and its implementation into a routine service laboratory.
abstract: BACKGROUND
  LC-MS/MS is rapidly becoming the technology of choice for measuring steroid hormones. We have developed a rapid LC-MS/MS assay for the routine analysis of serum cortisol. We have used this assay to investigate the effects of gender and exogenous steroid interference on the immunoassay measurement of serum cortisol.
  
  
  METHODS
  Zinc sulphate (40 µL) was added to 20 µL of sample. This was vortexed for 10 s followed by the addition of 100 µL of internal standard in methanol. Following mixing and centrifugation, 10 µL of sample was injected into an Acquity LC system coupled to a Quattro Premier tandem mass spectrometer. Serum samples (n = 149) were analysed by LC-MS/MS and two commercial immunoassays. Results were then compared for all samples and for gender differences. A further set of serum samples (n = 171) was analysed by the LC-MS/MS assay and a GC-MS assay.
  
  
  RESULTS
  Cortisol had a retention time of 0.98 min and the assay had an injection-to-injection time of 2.6 min per sample. Mean recovery was 99% and mean CV was 8%. The immunoassays gave comparisons of: Roche = 1.23 × LC-MS/MS -1.12 nmol/L and Abbott = 0.94 × LC-MS/MS + 11.97. The comparison with GC-MS showed LC-MS/MS = 1.11 × GC-MS - 22.90.
  
  
  CONCLUSIONS
  We have developed an LC-MS/MS assay for serum cortisol analysis that is suitable for routine clinical use and has been in use in our laboratory for 12 months. The availability of this assay will give more reliable results in patients receiving exogenous steroid therapy.
authors.affiliation: Manchester Academic Health Science Centre, University Hospital of South Manchester, The University of Manchester, Manchester, UK. laura.owen@uhsm.nhs.uk
authors.forename: Laura J
authors.initials: LJ
authors.lastname: Owen
authors.forename: Joanne E
authors.initials: JE
authors.lastname: Adaway
authors.forename: Sarah
authors.initials: S
authors.lastname: Davies
authors.forename: Sarah
authors.initials: S
authors.lastname: Neale
authors.forename: Nadia
authors.initials: N
authors.lastname: El-Farhan
authors.forename: David
authors.initials: D
authors.lastname: Ducroq
authors.forename: Carol
authors.initials: C
authors.lastname: Evans
authors.forename: D Aled
authors.initials: DA
authors.lastname: Rees
authors.forename: Finlay
authors.initials: F
authors.lastname: MacKenzie
authors.forename: Brian G
authors.initials: BG
authors.lastname: Keevil
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods / Q000639:trends; D019411:Clinical Laboratory Techniques / Q000379:methods / Q000639:trends; D003955:Diagnostic Tests, Routine / Q000379:methods / Q000639:trends*; D006801:Humans; D006854:Hydrocortisone / Q000097:blood*; D053719:Tandem Mass Spectrometry / Q000379:methods / Q000639:trends*
chemical_list: D006854:Hydrocortisone
keywords: LC-MS/MS; Serum cortisol