Understanding Sensitivity and Cross-Reactivity of Xylazine Lateral Flow Immunoassay Test Strips for Drug Checking Applications

The continued prevalence of xylazine in the illicit drug market has necessitated the development of quick and simple methods for identification of adulterated materials. One method that could be employed is lateral flow immunoassays (also known as test strips), which are similar to those frequently used to determine the presence of fentanyl. In this work, we explored the use of xylazine test strips (XTS) for the detection of xylazine in drug residue samples and investigated their sensitivity and cross-reactivity. XTS were found to detect xylazine at concentrations as low as 0.5 µg/mL in solution and consistently produced a positive result at concentrations as low as 2.5 µg/mL. Cross-reactivity with other α2-agonists, α2-antagonists, commonly encountered drugs and cutting agents, and other structurally similar compounds was minimal, with lidocaine being the only compound of the 77 studied to illicit a positive response. When XTS were used to detect xylazine in real-world drug residue samples, xylazine was consistently detected in samples where the concentration was above 2 µg/mL and mixed results were obtained when the concentration ranged between 0.1 µg/mL and 2 µg/mL. When considering samples that were negative for xylazine or had xylazine present at a concentration above the 1 µg/mL cut-off, an overall accuracy of 98.6% was achieved.