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Abstract Background The opioid epidemic has caused an increase in overdose deaths which can be attributed to fentanyl combined with various illicit substances. Drug checking programs have been started by many harm reduction groups to provide tools for users to determine the composition of their street drugs. Immunoassay fentanyl test strips (FTS) allow users to test drugs for fentanyl by either filling a baggie or cooker with water to dissolve the sample and test. The antibody used in FTS is very selective for fentanyl at high dilutions, a characteristic of the traditional use of urine testing. These street sample preparation methods can lead to mg/mL concentrations of several potential interferents. We tested whether these concentrated samples could cause false positive results on a FTS. Methods 20 ng/mL Rapid Response FTS were obtained from BTNX Inc. and tested against 4 different pharmaceuticals (diphenhydramine, alprazolam, gabapentin, and naloxone buprenorphine) and 3 illicit stimulants [cocaine HCl, methamphetamine, and 3,4-methylenedioxymethamphetamine (MDMA)] in concentrations from 20 to 0.2 mg/mL. The FTS testing pad is divided into 2 sections: the control area and the test area. Control and test area signal intensities were quantified by ImageJ from photographs of the test strips and compared to a threshold set by fentanyl at the FTS limit of detection. Results False positive results indicating the presence of fentanyl were obtained from samples of methamphetamine, MDMA, and diphenhydramine at concentrations at or above 1 mg/mL. Diphenhydramine is a common cutting agent in heroin. The street sample preparation protocols for FTS use suggested by many online resources would produce such concentrations of these materials. Street samples need to be diluted more significantly to avoid interference from potential cutting agents and stimulants. Conclusions Fentanyl test strips are commercially available, successful at detecting fentanyl to the specified limit of detection and can be a valuable tool for harm reduction efforts. Users should be aware that when drugs and adulterants are in high concentrations, FTS can give a false positive result.