Evaporation-Enhanced Redox Cycling for Rapid Detection of Attomolar SARS-CoV-2 Virions Using Nanolithography-Free Electrochemical Devices

In fighting against infectious diseases such as COVID-19, simple-to-use, sensitive, scalable, and rapid diagnostics are crucial for early disease diagnosis. In this regard, electrochemical biosensors are particularly attractive in developing point-of-need diagnostics. Importantly, by being compatible with nano- and microfabrication methods, they are amenable to miniaturization, which reduces background noise and the required sample volume. However, miniaturization also reduces the signal level, making it challenging to detect low virus counts. In this work, microfabricated electrochemical sensors with a dual signal amplification scheme based on evaporation-enhanced redox cycling (E2RC) in a generator–collector configuration are developed. A scalable, nanolithography-free fabrication method is proposed to achieve a controllable sub-micrometer gap between three dimensional (3D) interdigitated microelectrodes by combining photolithography with template-driven electrodeposition. Using the optimized electrodes, the sensors achieve rapid detection with a limit of quantification of ≈1.2 × 10³ particles mL⁻¹ through continuous measurement in evaporating droplets containing SARS-CoV-2 virion mimics. Investigating particle charge and size reveals the role of electrophoretic enrichment in the overall response. The sensor performance is also validated using heat-inactivated SARS-CoV-2 virions, with selective response to SARS-CoV-2 against HCoV-299E, SARS-CoV S1, and MERS-CoV S1 (captured using antibody-functionalized magnetic nanoparticles). The proposed sensing method is sensitive, rapid, scalable, and can be extended to broader applications, including detection of bacteria, extracellular vesicles, and other viruses.