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

Abstract

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.

Xylazine Test Strips for Drug Checking

“…20 Early results from the John Hopkins Bloomberg School of Public Health regarding the use of the xylazine test strips for drug checking show the overall sensitivity of the test strips to be good and cross-reactivity to be limited to a single compound -lidocaine. 21 The xylazine test strips were found to detect xylazine at concentrations as low as 0.5 mcg/mL in solution and consistently produced a positive result at concentrations as low as 2.5 mcg/mL. 21 When the xylazine test strips were used to detect xylazine in realworld drug residue samples, xylazine was consistently detected in samples with a concentration greater than 2 mcg/mL; there were mixed results when the concentration ranged between 0.1 mcg/mL and 2 mcg/ mL.…”

“…21 The xylazine test strips were found to detect xylazine at concentrations as low as 0.5 mcg/mL in solution and consistently produced a positive result at concentrations as low as 2.5 mcg/mL. 21 When the xylazine test strips were used to detect xylazine in realworld drug residue samples, xylazine was consistently detected in samples with a concentration greater than 2 mcg/mL; there were mixed results when the concentration ranged between 0.1 mcg/mL and 2 mcg/ mL. 21 These findings about the limitations of these test strips in the smallest concentrations are pertinent to the overall effectiveness of the strips to detect xylazine in the unregulated drug supply.…”

“…21 When the xylazine test strips were used to detect xylazine in realworld drug residue samples, xylazine was consistently detected in samples with a concentration greater than 2 mcg/mL; there were mixed results when the concentration ranged between 0.1 mcg/mL and 2 mcg/ mL. 21 These findings about the limitations of these test strips in the smallest concentrations are pertinent to the overall effectiveness of the strips to detect xylazine in the unregulated drug supply. These results plus the findings from the study in Philadelphia suggest the BTNX test strips are a good tool for drug checking programs.…”

“…One potential safety concern outlined by the studies was the inconsistent detection of lower concentrations of xylazine (i.e., amounts lower than 2.5 mcg/mL). 21 It is possible that a substance that tests negative for xylazine using 1 of these strips might in fact contain a small amount of the substance. Because there is no safe blood concentration of xylazine for humans, more research is needed to determine the level of risk posed by missing smaller concentrations of xylazine for people who use drugs.…”