6 Fentanyl Comparison Study, evaluating second-generation Immunoassay and Point of Care Test

Background Fentanyl is a μ-opioid receptor agonist that has become a significant concern in the opioid crisis. Detection and medical intervention are crucial components in addressing this issue. Within clinical laboratories, we rely on immunoassays to presumptively screen samples for the presence of fentanyl. In this study, we compare the Lin Zhi II and ARK 2, both FDA-cleared, open-channel immunoassays for fentanyl detection in urine. Additionally, three point-of-care (POCT) CLIA-waived tests: InstaStrip, AllSource, and AllTest were evaluated. In the point-of-care setting, rapid results are essential for patient management and triage. All methods were compared to a quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS). Objective This study aims to compare three POCT methods against each other and assess the performance of central laboratory methods. Methods A total of 150 urine samples were collected over a period of six months and stored at -80 °C until analysis. Samples were selected based on the ARK2 immunoassay, which is currently used for clinical service. The samples were then analyzed using an LC-MS/MS method to detect fentanyl, norfentanyl, and xylazine. Subsequently, the samples were tested using three-POCT and the Lin Zhi II immunoassay on a Roche Cobas automated instrument. Discordant samples were further investigated using a high-resolution mass spectrometer to identify any interfering compounds. Results Of the 150 urine samples collected, the population distribution was 59% male and 41% female, with samples obtained from three different departments: the Emergency Department, Primary Care, and Special Clinic. LC-MS/MS analysis identified 52 true negatives and 98 true positives among the samples. Fentanyl concentrations ranged from 0.5 ng/mL to 4970 ng/mL, while norfentanyl concentrations ranged from 2 ng/mL to 27850 ng/mL. The ARK 2 method demonstrated a sensitivity of 100% and a specificity of 96%, whereas the Lin Zhi II assay exhibited a sensitivity of 96% and a specificity of 98%. The POCT results were as follows: InstaStrip had a sensitivity and specificity of 96%, AllTest had a sensitivity of 98% and specificity of 82%, and AllSource had a sensitivity of 100% and specificity of 62%. Overall, there were a total of 26 false positives across all methods. These 26 samples were further analyzed using a high-resolution mass spectrometer in an untargeted approach to evaluate for interfering substances. Conclusions The ARK 2 and Lin Zhi II immunoassays both demonstrated high sensitivity and specificity, with the ARK 2 assay showing slightly better overall performance. Among the POCT, InstaStrip exhibited the most balanced sensitivity and specificity. However, AllTest and AllSource, despite having high sensitivity, showed lower specificity, emphasizing the need for confirmatory testing. The wide dynamic range of fentanyl and norfentanyl concentrations observed in patient samples underscores the variability in patient exposure and the necessity for robust screening methods.