The online screening technique for urinary benzodiazepines: comparison with EMIT, FPIA, and GC-MS

Three commercial immunoassay systems (EMIT, EPIA, Online) for the screening of benzodiazepines in urine were evaluated using authentic patient samples with gas chromatography-mass spectrometry (GC-MS) as the reference method. The Online system (kinetic interaction of microparticles in solution) gained in performance by applying a 100-ng/mL cutoff limit and by incorporating beta-glucuronidase treatment, which could be automated on the Cobas Mira Plus instrument. When using enzymatic hydrolysis, all three immunoassay systems had high levels of sensitivity, including samples containing only flunitrazepam and nitrazepam metabolites. A high degree of concordance was observed between the Online and FPIA (fluorescence polarization immunoassay) systems when analyzing 138 randomly selected patient samples. The EMIT II and EMIT d.a.u. (enzyme multiplied immuno technique) systems gave a higher number of positive results, but the presence of benzodiazepines could not be verified by GC-MS in a substantial number of these cases. The rate of unconfirmed positive results was increased when enzyme hydrolysis was incorporated in the EMIT II assay. Although differences in the performances of the investigated assay systems were observed, they all seem appropriate for clinical use in detecting benzodiazepine intake in drug abusers when enzymatic hydrolysis is included.