(238g) A Click Chemistry Amplified Nanopore (CAN) Assay for Ultrasensitive Quantification of Infectious Diseases Related Biomarkers in Clinical Samples (Invited Speaker)

Despite major advances in HIV testing, ultrasensitive detection of early infection remains a key challenge, especially for the viral capsid protein p24, which is recognized as an alternative early virological biomarker of HIV-1 infection. To improve p24 detection in patients missed by immunological tests that dominate the diagnostics market, we have engineered new sandwich structures through biotin-labeled copper oxide nanoparticles and p24 antigen to develop a Click chemistry Amplified Nanopore (CAN) assay for ultrasensitive detection of HIV infection. This strategy achieved a limit of detection of 0.5 pg/ml for HIV-1 p24 antigen in human serum, demonstrating 20~100-fold higher analytical sensitivity than latest copper nanocluster-based immunoassays and clinically used benchmark enzyme-linked immunosorbent assays (ELISA), respectively. The excellent linear correlation between the biomarker concentration and signal events frequency further enables quantitative detection. Clinical validation in a pilot HIV cohort demonstrated superiority of the CAN assay for quantification at ultra-low concentration range and strong correlation with viral load. We believe that this sensitive, cost-effective, and robust strategy could greatly improve the utility of p24 antigen in detecting early infection and monitoring HIV progression/treatment efficacy, and also have demonstrated it for detecting circulating tuberculosis antigens and COVID-19 antibodies.