@article{canoura_alkhamis_venzke_ly_xiao_2024, title={Developing Aptamer-Based Colorimetric Opioid Tests}, volume={3}, ISSN={["2691-3704"]}, DOI={10.1021/jacsau.3c00801}, abstractNote={Opioids collectively cause over 80,000 deaths in the United States annually. The ability to rapidly identify these compounds in seized drug samples on-site will be essential for curtailing trafficking and distribution. Chemical reagent-based tests are fast and simple but also notorious for giving false results due to poor specificity, whereas portable Raman spectrometers have excellent selectivity but often face interference challenges with impure drug samples. In this work, we develop on-site sensors for morphine and structurally related opioid compounds based on in vitro-selected oligonucleotide affinity reagents known as aptamers. We employ a parallel-and-serial selection strategy to isolate aptamers that recognize heroin, morphine, codeine, hydrocodone, and hydromorphone, along with a toggle-selection approach to isolate aptamers that bind oxycodone and oxymorphone. We then utilize a new high-throughput sequencing-based approach to examine aptamer growth patterns over the course of selection and a high-throughput exonuclease-based screening assay to identify optimal aptamer candidates. Finally, we use two high-performance aptamers with KD of ∼1 μM to develop colorimetric dye-displacement assays that can specifically detect opioids like heroin and oxycodone at concentrations as low as 0.5 μM with a linear range of 0–16 μM. Importantly, our assays can detect opioids in complex chemical matrices, including pharmaceutical tablets and drug mixtures; in contrast, the conventional Marquis test completely fails in this context. These aptamer-based colorimetric assays enable the naked-eye identification of specific opioids within seconds and will play an important role in combatting opioid abuse.}, journal={JACS AU}, author={Canoura, Juan and Alkhamis, Obtin and Venzke, Matthew and Ly, Phuong T. and Xiao, Yi}, year={2024}, month={Mar} }
 @article{wang_canoura_byrd_nguyen_alkhamis_ly_xiao_2024, title={Examining the Relationship between Aptamer Complexity and Molecular Discrimination of a Low-Epitope Target}, volume={11}, ISSN={["2374-7951"]}, DOI={10.1021/acscentsci.4c01377}, abstractNote={Aptamers are oligonucleotide-based affinity reagents that are increasingly being used in various applications. Systematic evolution of ligands by exponential enrichment (SELEX) has been widely used to isolate aptamers for small-molecule targets, but it remains challenging to generate aptamers with high affinity and specificity for targets with few functional groups. To address this challenge, we have systematically evaluated strategies for optimizing the isolation of aptamers for (+)-methamphetamine, a target for which previously reported aptamers have weak or no binding affinity. We perform four trials of library-immobilized SELEX against (+)-methamphetamine and demonstrate that N30 libraries do not yield high-quality aptamers. However, by using a more complex N40 library design, stringent counter-SELEX, and fine-tuned selection conditions, we identify aptamers with high affinity for (+)-methamphetamine and better selectivity relative to existing antibodies. Bioinformatic analysis from our selections reveals that high-quality aptamers contain long conserved motifs and are more informationally dense. Finally, we demonstrate that our best aptamer can rapidly detect (+)-methamphetamine at toxicologically relevant concentrations in saliva in a colorimetric dye-displacement assay. The insights provided here demonstrate the challenges in generating high-quality aptamers for low complexity small-molecule targets and will help guide the design of more efficient future selection efforts.}, journal={ACS CENTRAL SCIENCE}, author={Wang, Linlin and Canoura, Juan and Byrd, Caleb and Nguyen, Thinh and Alkhamis, Obtin and Ly, Phuong and Xiao, Yi}, year={2024}, month={Nov} }