title: Limitations of EMIT benzodiazepine immunoassay for monitoring compliance of patients with benzodiazepine therapy even after hydrolyzing glucuronide metabolites in urine to increase cross-reactivity: comparison of immunoassay results with LC-MS/MS values.
abstract: BACKGROUND
  Benzodiazepines are widely prescribed, and compliance of patients with benzodiazepine therapy is often monitored using urine specimens. Although various commercially available benzodiazepines immunoassays are widely used for compliance monitoring, such immunoassays usually have low cross-reactivity with glucuronide metabolites. We studied the effect of hydrolyzing such glucuronide before analysis to reevaluate suitability of Enzyme multiplied immunoassay technique benzodiazepine immunoassay for monitoring compliance with benzodiazepine therapy.
  
  
  METHODS
  In 31 urine specimens collected from patients taking benzodiazepines, the true analyte concentrations were determined (after hydrolyzing glucuronide metabolites using beta-glucuronidase) using liquid chromatography-tandem mass spectrometry. These urine specimens were reanalyzed using EMIT benzodiazepine assay (Flex Reagent Cartridge; Siemens Diagnostics) and Vista analyzer.
  
  
  RESULTS
  We observed false negative test results with EMIT in 11 of 31 specimens analyzed where liquid chromatography-tandem mass spectrometry values were above the 200 ng/mL cutoff concentration, but EMIT benzodiazepine assay showed a negative result, indicating that despite hydrolysis of the specimen to liberate parent drug (glucuronide metabolite often has poor cross-reactivity), the false negative rate using EMIT assay was 35.5%.
  
  
  CONCLUSIONS
  Patient compliance with benzodiazepine therapy must be monitored using a chromatographic method.
authors.affiliation: *Physicians Choice Laboratory Services, Rock Hill, SC; †Laboratory Services, Memorial-Hermann Hospital at Texas Medical Center; and ‡Department of Pathology and Laboratory Medicine, University of Texas-Houston Medical School.
authors.forename: R Brent
authors.initials: RB
authors.lastname: Dixon
authors.forename: Diana
authors.initials: D
authors.lastname: Floyd
authors.forename: Amitava
authors.initials: A
authors.lastname: Dasgupta
mesh_terms: D001569:Benzodiazepines / Q000032:analysis* / Q000627:therapeutic use* / Q000652:urine; D001711:Biotransformation; D002851:Chromatography, High Pressure Liquid; D003429:Cross Reactions; D016903:Drug Monitoring / Q000379:methods*; D017248:Enzyme Multiplied Immunoassay Technique; D005188:False Negative Reactions; D005966:Glucuronidase / Q000737:chemistry; D020719:Glucuronides / Q000652:urine; D006801:Humans; D010349:Patient Compliance; D053719:Tandem Mass Spectrometry
chemical_list: D020719:Glucuronides; D001569:Benzodiazepines; D005966:Glucuronidase