Validation of the Only Commercially Available Immunoassay for Synthetic Cathinones in Urine: Randox Drugs of Abuse V Biochip Array Technology

摘要

Deterrence of synthetic cathinone abuse is hampered by the lack of a high-throughput immunoassay screen. The Randox Drugs of Abuse V biochip immunoassay (DOA-V) contains two synthetic cathinones antibodies: Bath Salt I (BSI) targets mephedrone/methcathinone and Bath Salt II (BSII) targets 3’,4’-methylenedioxypyrovalerone (MDPV)/3’,4’-methylenedioxy-α-pyrrolidinobutiophenone (MDPBP). We evaluated DOA-V synthetic cathinones performance and conducted a full validation on the original assay with calibrators reconstituted in water, and the new assay with calibrators prepared in lyophilized urine; both utilized the same antibodies and were run on the fully automated Evidence^® Analyzer. 20,017 authentic military urine specimens were screened and confirmed by LC-MS/MS for 28 synthetic cathinones. Limits of detection (LOD) for the original and new assays were 0.35 and 0.18 (BSI), and 8.5 and 9.2µg/L (BSII), respectively. Linearity was acceptable (R²>0.98); however, a large negative bias was observed with in-house prepared calibrators. Intra-assay imprecision was <20% BSI-II, while inter-assay imprecision was 18–42% BSI and <22% BSII. Precision was acceptable for Randox controls. Cross-reactivities of many additional synthetic cathinones were determined. Authentic drug-free negative urine pH <4 produced false positive results for BSI (6.3µg/L) and BSII (473µg/L). Oxidizing agents reduced BSI and increased BSII results. Sensitivity, specificity, and efficiency of 100%, 52.1%, and 53.0% were obtained at manufacturer’s proposed cutoffs (BSI 5µg/L, BSII 30µg/L). Performance improved if cutoff concentrations increased (BSI 7.5µg/L, BSII 40µg/L); however, there were limited confirmed positive specimens. Currently, this is the first and only fully validated immunoassay for preliminary detection of synthetic cathinones in urine.