title: Monoclonal antibody-based time-resolved fluorescence immunoassays for daidzein, genistein, and equol in blood and urine: application to the Isoheart intervention study.
abstract: BACKGROUND
  Time-resolved fluorescence immunoassays (TR-FIAs) for phytoestrogens in biological samples are an alternative to mass spectrometric methods. These immunoassays were used to test urine and plasma samples from individuals in a dietary intervention trial aimed at determining the efficacy of dietary isoflavones in reducing the risk of coronary heart disease in postmenopausal women.
  
  
  METHODS
  We established murine monoclonal TR-FIA methods for daidzein, genistein, and equol. These assays could be performed manually or adapted to an automated analyzer for high throughput and increased accuracy. Analysis of urine was conducted on nonextracted samples. Blood analysis was performed on nonextracted samples for daidzein, whereas genistein and equol required diethyl-ether extraction.
  
  
  RESULTS
  Comparison of monoclonal TR-FIA, commercial polyclonal antibody-based TR-FIA, and gas chromatography-mass spectrometry showed correlations (r, 0.911-0.994) across the concentration range observed in the Isoheart study (50 mg/day isoflavones). The concentrations of urinary daidzein and genistein observed during intervention demonstrated good compliance, and a corresponding increase in serum daidzein and genistein confirmed bioavailability of the isoflavone-rich foods; 33 of the 117 volunteers (28.2%) were classified as equol producers on the basis of their urinary equol concentration (>936 nmol/L), and significant differences in the numbers of equol producers were observed between Berlin and the 3 other European cohorts studied.
  
  
  CONCLUSIONS
  The validated monoclonal TR-FIA methods are applicable for use in large-scale human phytoestrogen intervention studies and can be used to monitor compliance, demonstrate bioavailability, and assess equol producer status.
authors.affiliation: Unilever Corporate Research, Sharnbrook, United Kingdom. Duncan.Talbot@Unilever.com
authors.forename: Duncan C S
authors.initials: DC
authors.lastname: Talbot
authors.forename: Richard M
authors.initials: RM
authors.lastname: Ogborne
authors.forename: Tony
authors.initials: T
authors.lastname: Dadd
authors.forename: Herman
authors.initials: H
authors.lastname: Adlercreutz
authors.forename: Geoff
authors.initials: G
authors.lastname: Barnard
authors.forename: Susanne
authors.initials: S
authors.lastname: Bugel
authors.forename: Fortune
authors.initials: F
authors.lastname: Kohen
authors.forename: Sandra
authors.initials: S
authors.lastname: Marlin
authors.forename: Jerry
authors.initials: J
authors.lastname: Piron
authors.forename: Aedin
authors.initials: A
authors.lastname: Cassidy
authors.forename: Jonathan
authors.initials: J
authors.lastname: Powell
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D001682:Biological Availability; D003327:Coronary Disease / Q000517:prevention & control*; D019587:Dietary Supplements; D060754:Equol; D005260:Female; D015200:Fluoroimmunoassay / Q000379:methods; D008401:Gas Chromatography-Mass Spectrometry; D019833:Genistein / Q000032:analysis* / Q000097:blood / Q000652:urine; D006801:Humans; D007529:Isoflavones / Q000032:analysis* / Q000097:blood / Q000652:urine; D051379:Mice; D008807:Mice, Inbred BALB C; D017698:Postmenopause; D015203:Reproducibility of Results
chemical_list: C007610:4',7-dihydroxy-3,4-dihydroisoflavone; D000911:Antibodies, Monoclonal; D007529:Isoflavones; D060754:Equol; C004742:daidzein; D019833:Genistein