Fig. 2.

Assay cartridge and integrated portable DM device for POC-CRISPR. (A) The assay cartridge, which is fabricated via simple assembly of inexpensive thermoplastic materials, has 3 independent wells for holding the mixture of sample and magnetic bead buffer (pH = 5), a wash buffer (pH = 7), and the CRISPR-Cas12a-assisted RT-RPA reaction mixture. An immiscible oil overlays these aqueous reagents to prevent evaporation and reagent mixing. After injection of sample into the assay cartridge, the pH-responsive magnetic beads can bind to SARS-CoV-2 RNA and transport the RNA to the wash buffer – where the RNA remains beads-bound due to the neutral pH – and finally to the CRISPR-Cas12a-assisted RT-RPA reaction mixture. (B) The integrated portable DM device (represented here with a CAD schematic) houses a motorized magnetic arm for manipulating magnetic beads and executing sample preparation procedures, a miniature heating module for controlling the CRISPR-Cas12a-assisted RT-RPA reaction temperature, a compact fluorescence detector for detecting the fluorescence emitted from the reaction in real-time, and a microcontroller for controlling these components and hence automating the assay in the cartridge.