Fentanyl Immunoassays Effective at Detecting Designer Variants of the Drug, Study Finds

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Fentanyl lollipops

NEW YORK (360Dx) – Researchers at Sweden's Karolinska Institute have established that existing fentanyl immunoassays can effectively detect designer versions of the drug in user's urine.

Their findings, presented in a study published this month in the journal Drug Testing and Analysis, indicate that commercial fentanyl tests can be used to screen for designer fentanyl products, which, said Olof Beck, an adjunct professor at Karolinska and senior author on the paper, present a challenge for drug testing in Sweden and other countries.

According to Beck and his coauthors, the last decade has seen a sharp rise in the availability of what they term new psychoactive substances (NPS), with more than 530 such substances identified in Europe between 2009 and 2016. Once identified, these substances are often registered and made illegal. However, Beck said, this process can take as long as a year, during which time they can be sold legally, typically over the internet. Additionally, once an NPS is made illegal, sellers will often begin offering an NPS featuring a structural variant of the drug that has not yet been outlawed. This makes for a growing and shifting collection of substances for which labs need to test.

Initially, NPS products were mainly synthetic cannabinoids and stimulants, but more recently opioids, and designer fentanyls especially, have become more prominent. Commercial tests for fentanyl exist, but Beck said researchers had not yet determined how effectively those assays could detect the various fentanyl variants being used in NPS. These variants can be detected using mass spectrometry, but an effective immunoassay would allow for more rapid, high-throughput screening of urine samples, with positives then being passed on to mass spec for confirmation.

The Karolinska researchers tested three different fentanyl immunoassays: the DRI Fentanyl Enzyme Immunoassay from Thermo Fisher Scientific, the ARK Fentanyl Assay homogeneous enzyme immunoassay from ARK Diagnostics, and the Immunalysis Fentanyl Urine SEFRIA Drug Screening Kit from Immunalysis. They then prepared test samples by spiking amounts of fentanyl and 12 fentanyl variants in urine.

All three assays were accurate enough in detecting the different fentanyls to be used as initial screening tests, Beck said, though, he noted that there were some differences in performance. Most notably, the DRI assay struggled with some of the blank urine samples, Beck said, noting that in some cases it returned negative concentration measurements in the samples, indicating that the "low-cutoff calibrators are a little bit variable in different urine samples." This raises the possible risk of false negatives with this test.

The coefficients of variation for each of the assays varied, with CVs for the DRI test somewhat higher than those for the ARK or SEFRIA test. However, the authors noted, the CVs for all three tests "were acceptable for routine use."

The authors added that the screening cut-offs used could be lowered from those recommended by the test manufacturers to improve sensitivity, with positive results then confirmed via mass spec.

The researchers also looked at 58 urine samples that had been confirmed positive for fentanyl or fentanyl analogs by mass spec. The DRI test returned a positive result in 49 of the 58 samples; the ARK test returned a positive result in 56 of 58, and the SEFRIA test returned a positive result in 53 of 58.

Beck said he and his colleagues now plan to implement the immunoassays in their drug testing routines. They have been relying on mass spec-based assays for testing for fentanyl variants, he said.

"In practice you have maybe 10 or 20 substances you want to measure, but there's no immunoassay available, and so to do it you need to use mass spectrometry," he said. "But now, I think that for fentanyl, these assays are working quite well."

The fentanyl work is part of a larger effort Beck and his colleagues have undertaken in collaboration with Swedish authorities on identifying new designer drugs.

"We've published maybe 30 papers on new narcotic substances [appearing among Swedish drug users]," he said. "Fentanyl is just one of them."