DNA origami capture and tethering of intact viruses and label-free digital detection by photonic-crystal-enhanced scattering microscopy

DNA origami offers a powerful approach for creating flexible and bendable mechanical structures that can selectively capture nano-objects such as intact virions by incorporating target specific functions in precisely arranged spatial patterns. Recently, our group explored the design and implementation of designer DNA nets (DDNs) that display a 2-dimensional pattern of aptamers that bind with pattern matching to the outer spike proteins of intact virions. We took this concept further with implementation of a DNA NanoGripper, that contains a palm and four bendable fingers as inspired by naturally evolved human hands and animal claws. We demonstrate that the DDNs and NanoGrippers can be engineered to effectively interact with and capture gold nanoparticles, SARS-CoV-2, and HIV while tethering them to a photonic crystal (PC) surface. Using the PC surface in combination with a PC-enhanced microscope instrument, fluorescence, optical absorption, and scattering can be amplified to facilitate digital resolution detection with high signal-to-noise. Interestingly, when captured viruses are tethered to a PC surface and detected with label-free Photonic Resonator Interferometric Scattering Microscopy (PRISM) at high video frame rates, we observe persistent “wiggling” that differentiates selectively captured objects from those that contact the biosensor surface without being captured. Using DDNs and NanoGrippers to capture virions and PC-enhanced microscopy to detect them, we achieve detection limits in the ~100-1000 copies per milliliter concentration range, with a single-step, room temperature, 30-minute sample incubation time in saliva and serum. Our approach paves a path towards a rapid, simple, and highly sensitive alternative to conventional laboratory-based approaches.