Clinical and forensic diagnosis of very recent Heroin intake by 6-acetylmorphine immunoassay test and LC-MS/MS analysis in Urine and Blood

Aim. The study evaluates the suitability of a specific immunoassay screening test for 6-acetylmorphine (6-AM) in the setting of suspected very recent heroin consumption for forensic and clinical purposes. Material and method. The EMIT® II Plus 6-AM immunoassay was applied in 65 cases that had already tested positive for morphine in urine or blood. Biological samples (n.65 urine and n.53 blood) were obtained from workplace drug tests (WDT n. 5), tests for driving under the influence of drugs (DUID n. 30), vehicle accidents (n. 10), overdoses (n. 12) and heroin-related deaths (n. 8) cases. The 6-AM screening assay results were confirmed with the LC-MS/MS analysis in relation to the cut-off set at 10 ng/mL for both urine and blood. Results. Among the 65 urine samples (all morphine-positive), 38 samples were 6-AM-positive and 27 were 6-AM-negative with 100% agreement between the positive/negative results of the two assays. Among the 53 blood samples (34 positive and 19 negative for the morphine), 16 were 6-AM positive and 37 were negative. Only one of the blood samples, positive for 6-AM by LC-MS/MS at 10.3 ng/mL, was negative by the immunoassay test. Based on the concordance between the results of the 6-AM immunoassay versus the LC-MS/MS, the sensitivity of the 6-AM assay was calculated as 100% and 95% for urine and blood respectively, with a specificity and accuracy of 100% for both biological samples. In addition, the study demonstrated that the 6-AM assay test, originally developed for urine, is also sufficiently sensitive to identify 6-AM in blood. Therefore, it could be applied in cases of vehicle accidents or overdose to distinguish rapidly between very recent heroin use and the intake of other opiates for therapeutic purposes.