title: Specificity assessment of immunoassay kits for determination of urinary free cortisol concentrations.
abstract: BACKGROUND
  In immunoassay kits for determination of urinary free cortisol (UFC) concentrations, the results vary markedly from kit to kit, so we compared in this study the reaction specificity among 4 commercially available immunoassay kits to determine the applicability of these assays in routine determination of UFC concentrations.
  
  
  METHODS
  Using 4 commercially available kits, cross-reaction was investigated. In addition, urine samples were fractionated by HPLC to investigate endogenous immunoreactive cortisol responses. HPLC fractions were subjected to gas chromatography-mass spectrometry (GCMS) to identify substances causing inter-kit assay discrepancies.
  
  
  RESULTS
  Among the 4 kits, cortisol Kit "TFB" (Immunotech; IOT-RIA method) showed the lowest cross-reaction (2.5%) for prednisolone. Furthermore, on HPLC, 87.8% of the reaction of the entire fraction was seen in the fractions corresponding to the elution position of standard cortisol with the IOT-RIA method; this was the highest percentage among the 4 kits. GCMS revealed that the substance that showed a cross-reaction with the other 3 kits was 5alpha-tetrahydrocortisol (5alpha-THF) glucuronide.
  
  
  CONCLUSIONS
  The IOT-RIA method was found to be the most specific for UFC. The other 3 commercially available kits showed cross-reaction with a conjugate of 5alpha-THF, found to be one of the causes of inter-kit assay discrepancies.
authors.affiliation: Mitsubishi Kagaku Bio-Clinical Laboratories, INC., 3-30-1 Shimura, Itabashi-ku, and Department of Clinical Laboratories, Keio University Hospital, Tokyo, Japan. h-horie@nm.mbcl.co.jp
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Horie
authors.forename: Takeshi
authors.initials: T
authors.lastname: Kidowaki
authors.forename: Yuhei
authors.initials: Y
authors.lastname: Koyama
authors.forename: Tatsuji
authors.initials: T
authors.lastname: Endo
authors.forename: Keiko
authors.initials: K
authors.lastname: Homma
authors.forename: Akira
authors.initials: A
authors.lastname: Kambegawa
authors.forename: Norihiko
authors.initials: N
authors.lastname: Aoki
mesh_terms: D000328:Adult; D002851:Chromatography, High Pressure Liquid; D003429:Cross Reactions; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D006854:Hydrocortisone / Q000276:immunology / Q000652:urine*; D008297:Male; D047748:Pituitary ACTH Hypersecretion / Q000652:urine; D011863:Radioimmunoassay / Q000379:methods*; D011933:Reagent Kits, Diagnostic
chemical_list: D011933:Reagent Kits, Diagnostic; D006854:Hydrocortisone