Loop-Mediated Isothermal Amplification-Coupled Glass Nanopore Counting Toward Sensitive and Specific Nucleic Acid Testing

Solid-state nanopores have shown great promise and achieved tremendous success in label-free single-molecule analysis. However, there are three common challenges in solid-state nanopore sensors, including the nanopore size variations from batch to batch that makes the interpretation of the sensing results difficult, the incorporation of sensor specificity, and the impractical analysis time at low analyte concentration due to diffusion-limited mass transport. Here, we demonstrate a novel loop-mediated isothermal amplification (LAMP)-coupled glass nanopore counting strategy that could effectively address these challenges. By using the glass nanopore in the counting mode (versus the sizing mode), the device fabrication challenge is considerably eased since it allows a certain degree of pore size variations and no surface functionalization is needed. The specific molecule replication effectively breaks the diffusion-limited mass transport thanks to the exponential growth of the target molecules. We show the LAMP-coupled glass nanopore counting has the potential to be used in a qualitative test as well as in a quantitative nucleic acid test. This approach lends itself to most amplification strategies as long as the target template is specifically replicated in numbers. The highly sensitive and specific sensing strategy would open a new avenue for solid-state nanopore sensors toward a new form of compact, rapid, low-cost nucleic acid testing at the point of care.