SIGNIFY™ ER tricyclic antidepressant false positives in diphenhydramine overdose cases

The SIGNIFY™ ER test for diphenhydramine (DPH) overdose yields false-positive results for tricyclic antidepressants (TCA) in multiple cases, complicating the identification of the causative substance of poisoning. We investigated the causes of TCA false positives in DPH overdose cases. From March 2021 to September 2023, 11 cases of DPH overdose with no concomitant TCA use were identified and categorized into two groups: four with false-positive TCA results (FPG) and seven with negative results (NG) in SIGNIFY™ ER. The blood and urinary DPH concentrations and the urinary concentrations of its three major metabolites (diphenhydramine N-oxide, DPH-NO; N-desmethyl diphenhydramine, DM-DPH; and diphenhydramine N-glucuronide, DPHG) were measured using liquid chromatography quadrupole time-of-flight mass spectrometry, and differences between the groups were examined. Standard substances of DPH, DPH-NO, DM-DPH, DPHG, and mixtures of DPH-NO and DPHG at ratios of 50:50, 25:75, and 75:25 were added to blank urine samples, and TCA was measured using the SIGNIFY™ ER. The concentrations of the substances in the FPG and NG, respectively, were: blood DPH, 0.9-9 and 0.33-49 µg/mL; urinary DPH, 10-110 and 2.3-36 µg/mL; urinary DPH-NO, 110-170 and 0.05-10 µg/mL; urinary DM-DPH, 5.3-47 and 0.13-3.4 µg/mL; and urinary DPHG, 28-370 and 0.24-67 µg/mL. When using spiked urine samples, false positives were obtained for DPH, DPH-NO, DM-DPH, and DPHG at 500, 110, 200, and 70 µg/mL, respectively. In the mixtures of DPH-NO and DPHG, false positives were obtained at all three ratios. TCA false positives in the SIGNIFY™ ER test for DPH overdose cases are suggested to be yielded by DPH-NO and DPHG. For a positive test result without information on TCA use, DPH overdose should be considered.