Optimization and validation of the fast blue BB (FBBB) screening test for the detection of delta-9-tetrahydrocannabinol (Δ9-THC) in oral fluid

The fast blue BB test (FBBB) colorimetric test has been previously validated for the differentiation between marijuana-type and hemp-type cannabis plants. Individuals under the influence of delta-9-tetrahydrocannabinol (Δ9-THC) may be impaired to perform activities such as driving and the rapid, on-site detection of recent use of Δ9-THC could help to prevent traffic accidents. The current study describes, for the first time, the optimization and validation of the FBBB color test from oral fluid (OF) for field use. The FBBB reaction with Δ9-THC forms a red chromophore and a bright red fluorophore that is visualized with a portable fluorescence reader under UV light. The test was optimized for a miniaturized reaction on a 6.3 mm diameter glass fiber filter spot. The limit of detection (LOD) was established as 0.5 µg/mL or 500 ng/mL of Δ9-THC in OF (5 ng of Δ9-THC on the spot) by fluorescence detection. Other figures of merit include linearity in the 0.5–10 µg/mL range, acceptable precision (9.6–28% RSD) and an accuracy of ± 23.2–56.2%. Results from interference studies using different OF collection devices and substances are reported. Interferences might also occur when minor cannabinoids (CBD, CBN, CBG, and Δ8-THC) and Δ9-THC metabolites (THC-COOH and THC-OH) are also present in the matrix. Pre-loaded FBBB reagent is stable on the substrate when stored below 4 °C for 15 days and the color and fluorescence persist on the spot for at least 30 days post reaction. Future studies include development of an OF extraction procedure coupled to a prototype device for field application.