Clin Chem. 2026 Jul 1:hvag079. doi: 10.1093/clinchem/hvag079. Online ahead of print.
ABSTRACT
BACKGROUND: Comprehensive drug testing (CDT) by liquid chromatography-high-resolution mass spectrometry (LC-HRMS) is a valuable method for evaluating clinical samples for unknown toxicological agents. However, the sensitivity of CDT methods is generally lower than targeted approaches. A novel MS technology, linear ion trap (LIT)-pulsing, has demonstrated improved sensitivity in proteomics and metabolomics applications, but its utility and interactions with different HRMS acquisition types, such as information-dependent acquisition (IDA) and sequential window acquisition of all theoretical fragment ion spectra (SWATH), have not been explored in small molecule analysis.
METHODS: CDT methods utilizing both IDA and SWATH acquisition were developed on a quadrupole time-of-flight (QTOF) instrument featuring LIT pulsing. Limits of detection (LODs) and process efficiencies were estimated for 150 toxicological agents. Additionally, 101 patient urine specimens were used to compare both methods against an established clinical CDT without LIT-pulsing. Selectivity was assessed using a targeted LC-HRMS method.
RESULTS: LIT-pulsed SWATH acquisition improved detection of toxicological agents compared to LIT-pulsed IDA, with a median 5-fold reduction to measured LODs and an average 5.9% reduction to matrix suppression. In authentic patient samples, LIT-pulsed IDA and SWATH acquisition identified 889 and 1083 toxicological agents compared to 835 using IDA without LIT-pulsing. The estimated selectivity of LIT-pulsed IDA was improved compared to conventional IDA acquisition, but SWATH acquisition demonstrated significant nonselectivity.
CONCLUSION: LIT-pulsing improves the sensitivity and selectivity of CDT methods. The largest improvements to sensitivity are observed using SWATH acquisition; however, these gains are accompanied by reduced selectivity. This highlights the need for careful validation of SWATH CDT methods to ensure high clinical performance.
PMID:42384459 | DOI:10.1093/clinchem/hvag079