AbstractThe rise in antibiotic-resistant pathogens, highly infectious viruses, and chronic diseases has prompted the search for rapid and versatile medical tests that can be performed by the patient. An electronic biosensing platform based on field-effect transistors (FETs) is particularly attractive due to sensitivity, fast turn-around, and compatibility with semiconductor manufacturing. However, the lack of methods for pathogen-specific functionalization of individual FETs prevents parallel detection of multiple pathogens. Indeed, so far functionalization of FET based biosensors is achieved by drop casting without any spatial selectivity. Here, we propose a paradigm shift in FET’s biofunctionalization. Specifically, we use thermal scanning probe lithography (tSPL) with a thermochemically sensitive polymer that can be spin-coated on any FET material. We demonstrate that this scalable, CMOS compatible methodology can be used to functionalize individual FETs with different bioreceptors on the same chip, at sub-20 nm resolution, paving the way for massively parallel FET detection of multiple pathogens. Antibody- and aptamer-modified FET sensors are then realized, achieving an ultra-sensitive detection of 5 aM of SARS-CoV-2 spike proteins and 10 human SARS-CoV-2 infectious live virus particles/ml, and selectivity against human influenza A (H1N1) live virus.