Evaluation of Four Immunoassay Screening Kits for the Detection of Benzodiazepines in Urine*

Abstract

The identification of benzodiazepines (BZD) in biological fluids can be a challenging process. The large number of various BZD in pharmaceutical distribution, with similar core structures, and individual metabolic profiles all contribute to a complicated and time-consuming analysis. The purpose of the current study was to evaluate the performance of four commercially available immunoassay urine screening kits for use in a forensic urine analysis testing program. The four kits included the Roche Benzodiazepine Plus KIMS assay, Microgenics CEDIA Benzodiazepine assay, Microgenics CEDIA high sensitivity assay with beta-glucuronidase, and Microgenics DRI reagent ready Benzodiazepine assay. Each kit was evaluated for linearity, precision, accuracy, carryover, reagent specificity, and confirmation rates. BZD reagent specificity was compared by analysis of 55 structurally dissimilar compounds to BZD. Negative responses to all 55 compounds were elicited by all four reagent assays. Cross-reactivity for the assays was demonstrated by detecting 27 structurally similar BZD. In addition, greater than 10,000 randomly collected urine samples were screened at a 200 ng/mL cutoff for each assay. Positive samples were confirmed by gas chromatography-mass spectrometry using a panel of 13 BZD confirmation standards. The Microgenics CEDIA high sensitivity assay demonstrated the highest positive screening rate as well as the highest confirmation rate of the four assays.