SOFT - TIAFT 1998 Poster Session 2 Wednesday October 7, 1998

Kathryn S. Schwenzer, Ryan Pearlman, Michael Tsilimidos, Salvatore J. Salamone, Rudolf Brenneisen*, Mahmoud A. ElSohly** and Shizia Feng**

Roche Diagnostics, Somerville, NJ, USA
* University of Bern, Bern, Switzerland
** ElSohly Laboratories, Oxford, MS, USA

A new reagent, cassette COBAS® INTEGRA Serum Benzodiazepines (SBENZ) for analysis of benzodiazepines in serum and/or plasma on the COBAS INTEGRA 700 analyzer using fluorescence polarization (FPIA) technology, was further evaluated for analysis of benzodiazepines in urine. Urine from 10 different donors was spiked with nordiazepam; recovery was within 95.2% at 130.6 ng/mL and within 91.7% at 41.2 ng/mL. Precision in urine was evaluated according to NCCLS Guideline EP5-T2 where the total precision at 7.4 ng/mL nordiazepam was 19.4%, 10 ng/mL was 11.1%, 19.6 ng/mL was 6.34%, 27.24 ng/mL was 4.98%, 41.22 ng/mL was 3.24%, 51.66 ng/mL was 2.71%, 83.72 ng/mL was 1.74%, and 104.6 ng/mL was 1.8%.

Clinical samples, N = 66, were evaluated by GC/MS for nordiazepam and oxazepam, LOD 1 ng/mL, a commercially available FPIA, LOD 40 ng/mL and the SBENZ urine application, LOD 7 ng/mL. The diagnostic sensitivity of the two immunoassays vs GC/MS was 100% and the diagnostic selectivity was 87.5%.

A clinical study was conducted to assess the ability to detect flunitrazepam (FNP) by the different methods. The clinical study consisted of four individuals (2 male, 2 female) who had taken a single 2 mg dose of FNP. Urine was collected over a 72-hour period. The urine samples were evaluated by GC/MS with a 1 ng/mL LOD for 7-aminoflunitrazepam, a commercially available FPIA and COBAS INTEGRA SBENZ. The GC/MS, SBENZ, and a commercial FPIA picked up 45, 41, and 35 positive FNP urines respectively. Three samples were negative by all methods.

The improved detection of FNP use by the SBENZ assay as compared to the other commercially available FPIA may be explained by increased cross-reactivity of the major metabolite 7-aminoFNP which cross-reacts 60.8% compared to 15% (Clin. Chem., Vol. 38, No. 2, 271-275 (1992)) for the other commercially available FPIA. SBENZ cross-reactivity to the other metabolites of FNP, were 0.47% to 7-acetamido-FNP, non detectable to 7-acetamido-3-OH-desmethyl-FNP, 12% to 7-amino3-OHFNP, 47.6% to desmethyl-FNP, and 13.8% to 3-hydroxyFNP. The cassette COBAS INTEGRA SBENZ assay may be used for the analysis of serum, plasma and urine.

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