Project Summary/Abstract The United States is experiencing an epidemic of unprecedented scope with heroin use resurging and a troubling pervasive increase in abuse of prescription opioids such as fentanyl, fentanyl derivatives, methadone, oxycodone and hydrocodone. More than 6 out of 10 drug overdoses are now opioid-related, resulting in ~90 deaths/day and a more than $75B/year economic impact. Additionally, there is a rapidly expanding need to detect numerous other small molecules such as therapeutic agents, in the home or potential WMDs like sarin and toxic pesticides in the remote or field setting. To develop a general purpose tool to address such detection, we propose to refine our compensated interferometric reader (CIR) and further develop a unique and efficient, matrix insensitive, assay methodology to address the need for high sensitivity, rapid testing of opioids, therapeutics and environmental contaminants. Our efforts will produce a first of its kind bench- top (and field-compatible) CIR and user-friendly (turn-key) assay system for the detection and/or quantification at ng/mL-pg/mL sensitivity of a wide range of targets including opiates, small molecule drugs, and organophosphorus pesticides. The assay based on a near-surface assay (NSA) embodiment, consists of three-steps; 1) collect sample (dilute if needed), 2) fill the pre- coated capillary and 3) read the ‘sample’ with CIR. Results for drug or metabolite detection in urine by CIR is predicted to be <300 pg/mL sensitivity for the fentanyl family, a >10-1000-fold improvement over the best lateral flow tests (LFT)1 and the current cutoff for standard drug tests. The non-invasive, nanoliter volume, high specificity urine or buffer-diluent assays will facilitate rapid field-based detection of environmental samples, seized illicit narcotics, counterfeit drugs and organophosphorus nerve agents (OPNA). Similar minimally invasive assays are feasible with minute amounts of finger-stick blood. The novel methodology is enabled by the marriage of three patented technologies, a multiplex-SELEX approach for aptamer selection, an aptamer-probe assay, and CIR to quantify the target. Phase I and Phase II Aims proposed here are designed to enable rapid translation/commercialization of the proposed technology.