B-403 Validation of a high-resolution mass spectrometry-based comprehensive drug test

Abstract

Liquid chromatography tandem mass spectrometry (LC-MS/MS) is a powerful tool for toxicology testing, providing critical information about substances causing overdoses and statistics on drug use within the patient community. In this study, we transitioned our high-resolution mass spectrometry-based (HRMS) comprehensive drug test from a SCIEX TripleTOF®5600 to the SCIEX X500R QTOF. In addition to validating the assay, we expanded our library of drugs with additional substances seen in our patient population, including illicit drugs such as fentanyl analogs and designer benzodiazepines. Data was collected with high-resolution mass spectrometry (HRMS) on a SCIEX X500R QTOF using information dependent acquisition. The assay was validated for urine, serum, plasma, and whole blood samples. Scores were calculated for each compound based on mass error, retention time, isotope pattern, and library match; all data was manually reviewed for final calls. Limits of detection and matrix effects were quantified in urine and serum using 5-10 blank matrices into which mixes of drug standards were spiked. Over 90 (urine) and 50 (serum) patient comparisons were analyzed to assess equivalency of the instruments. A new spectral library was built on the X500R for over 220 compounds, including 18 compounds not part of the previously validated assay. Limits of detection were determined in the range of 5-5000 ng/mL and 1-1000 ng/mL in urine and serum, respectively. Carryover effects were assessed with spiking studies and determined to be negligible. Matrix effects showed ion suppression or enhancement depending on the compound without significantly affecting LODs. The comprehensive drug test was validated and implemented clinically. With the X500R we detect 704 of 787 (∼89%) compounds originally detected on the 5600; however, the X500R additionally detects 129 compounds not picked up by the 5600, yielding ∼6% improved sensitivity. Additionally, we detect at least one of the newly added compounds to our library in about one third of urine patient comparisons analyzed, with the most frequently observed compounds being 4-ANPP (fentanyl precursor) and beta-hydroxy fentanyl (fentanyl analog). We present a framework for validation and implementation of a HRMS-based comprehensive drug test in the clinical lab, including assessment of limits of detection, matrix effects, carryover, and evaluation of the assay using patient samples. Expansion of our menu to reflect drug use trends in our patient population provides valuable clinical information and furthers our bio-surveillance efforts.