Development of a high-throughput centrifugal microsystem for enzyme-linked immunosorbent assay to detect SARS-CoV-2

Enzyme-linked immunosorbent assay (ELISA) is a commonly used diagnostic and quantification tool for protein biomarkers in biomedical research and industrial quality control. However, this technique has prolonged assay durations, along with requiring expensive equipment and trained technicians, which limits its on-site applications. In this study, we report a novel centrifugal microfluidic device that can perform the conventional ELISA assay in a high-throughput (HTP) and automatic manner. The centrifugal microfluidic microsystem incorporates four innovative features: (1) a rapid and easy dispenser structure with aliquoting microchannels, which can accurately fill the solution in 40 reaction chambers with just one injection, replacing the labor-intensive aliquoting process, (2) ultra-fast reagent loading into and discharging from 40 reaction chambers simultaneously within 15 sec through the combination of a siphon valve and centrifugal force control, (3) the exact and automatic handling of a small volume (only 30 µL per reaction) of the designated solution such as a sample, a washing solution, an HRP-antibody solution, and a TMB solution, (4) a miniaturized ELISA diagnostic system, consisting of a solution injector, a simple spindle motor, and a UV–vis absorbance detector for the point-of-care testing (POCT). Using the HTP centrifugal microfluidic device with the in-house POCT ELISA analyzer, we could automatically detect 40 samples of SARS-CoV-2 per run in 75 min, significantly reducing the time, cost, and labor compared with the conventional ELISA assay. SARS-CoV-2 was diagnosed with a limit-of-detection (LOD) of 10.2 ng/mL. In addition, clinical samples as well as the SARS-CoV-2 variants were successfully confirmed on the POCT ELISA analyzer.