Poster Y5 - Biosensor for the Detection of Antiretroviral Drugs in Urine

Introduction: Testing has emerged as a critical gateway to prevention, treatment, and supportive care services in the management of antiretroviral (ARV) drugs. This study develops a novel urinalysis device tailored to detect ARVs in HIV/AIDS patients, particularly those in resource-poor areas and in low and middle-income countries (LMICs), to help healthcare providers ensure patients adhere to their prescriptions and promote long-term health.

Materials and

Methods: Metabolic by-products (MbP) stemming from ARV drugs were identified, and qualitative colorimetric assays were developed. Known amounts of MbPs were diluted in aqueous solutions and simulated urine and verified using colorimetric assays and spectroscopy as a control. Two distinct molecularly imprinted polymer (MIP) biosensors were tailored to recognize the MbPs. The effectiveness of each MIP procedure was carried out in artificial urine using cyclic voltammetry (CV). Known MbP amounts were diluted in urine to achieve a calibration curve through CV for each method. Method A utilized a crosslinked acrylamide polymer, and Method B utilized a crosslinked methyl methacrylate/acrylamide copolymer. The washing methods of these fabrication procedures varied as well. The washing is done to eliminate the excess reagents not bound to the biosensor. Once fabrication was completed for each method, a final wash step was included. Method A's washing procedure consisted of placing the sensor in DI water for 15 minutes and ethanol for 10 minutes under moderate stirring. Method B’s washing procedure consisted of placing the sensors in DI water for 30 minutes at 60℃ and then in 5% ethanol for 30 minutes under moderate stirring. The electrodes were washed between scans while creating the calibration curve.

Results, Conclusions, and Discussions: From preliminary testing, when the MbPs were diluted into an aqueous solution, the qualitative colorimetric assay could visually show the presence of the drug byproducts based on the concentration. However, when diluted in artificial urine and tested, samples appeared dark and could not be read under spectroscopy. Due to these limitations, we explored using MIP biosensors to detect ARV drugs in urine. Methods A & B’s preliminary testing has been shown to produce inconsistent linear calibration curves when performing CV testing seen in Figures 1 and 2. Primarily, the group is seeing difficulties in the chemical binding of the gold screen printed sensors and MIP. Our results also deviated from the expected values due to the washing technique of each method during CV testing. Although the washing technique was intended to release the MbPs from free binding sites, it inadvertently altered the binding sites, resulting in inefficient calibration curves.
Further work must be done in MIP biosensors to achieve consistent data to show it becoming a practical technology for detecting ARV medications in low and middle-income countries (LMICs). Establishing a uniform protocol for the washing technique step is essential to enhance data consistency. This standardized approach ensures measurement accuracy and reliability while consistently releasing MbPs from binding sites between CV testing. Testing regarding the life cycle, shelf life, and environmental conditions should be explored to further determine the practicality of MIP biosensors, especially in LMICs. Life cycle testing of MIPs will be beneficial in quantifying the degradation rate and identifying the reusability factor of MIPs. The shelf life is important to determine the performance longevity of MIPs post-fabrication. The shelf life and environmental testing will help establish the appropriate storage methods. From the literature reviews done by the group, the MpBs identified allow for more established electrochemical biosensor technologies to be tested and validated for this use case moving forward.

Acknowledgements (Optional): Acknowledgements: Clemson University Creative Inquiry for funding. REDDI Lab staff for MIP fabrication assistance. Multiscale Bioelectromechanics Lab Graduate Students for mentoring support- Arianna, Diego, Jeremiah, Ellie, Calvin, and Drew. Advisors - Dr. Delphine Dean, Dr. John DesJardins, Dr. Melinda Harman, and Dr. Melissa McCullough.