Using Capture-SELEX, three DNA aptamers were identified that bind to benzodiazepines in synthetic urine. Binding constants for the three aptamers with 14 unique benzodiazepines were measured and possible binding interactions between the aptamers and the benzodiazepine molecule were conceptualized. Using a strand-displacement fluorescence polarization assay, a biosensor was created for the detection of benzodiazepines in synthetic urine.
Benzodiazepines are a class of psychoactive compounds that are commonly prescribed for conditions such as anxiety, depression, and chronic pain. The use and abuse of benzodiazepines is slowly rising and requires reliable detection methods to monitor patient use and compliance. Most current screening methods are antibody-based and have inherent issues, including limited stability and shelf life. The use of non-antibody-based detection methods that overcome these challenges would be useful for testing purposes. Using in vitro selection, we have identified three DNA aptamers for benzodiazepines with Kd values between 0.2–0.6 μM for diazepam and alprazolam and their metabolites but little to no binding for lorazepam or clonazepam. One aptamer, 20DF42, was used to develop a strand-displacement fluorescence polarization biosensor for detection of benzodiazepines in synthetic urine. The resulting sensor had a limit of detection of 231 ng/mL with temazepam as the calibrant.Zum Volltext