Figure 2.

Schematic of nucleic acid detection by CARMEN–Cas13. (a) Identification of multiple circulating pathogens in human and animal species. (b) Depiction of the CARMEN–Cas13 workflow. ① After sample collection, RNA extraction, and amplification of the nucleic acid input, amplified targets are converted to RNA via T7 transcription and detected by Cas13 − crRNA RNP complexes. The resulting collateral cleavage activity of Cas13 produces a signal using a cleavage reporter RNA. In parallel, detection mixes are assembled, color-coded, and emulsified. ② Droplets from each emulsion are pooled into a single tube and mixed by pipetting. ③ The droplets are loaded onto a chip in a single pipetting step, during which the droplets self-organize into pairs. ④ Each microwell is imaged by fluorescence microscopy to identify the color code and to map the position of each droplet on the chip. ⑤ Droplets are merged, initiating the detection reaction. ⑥ After incubation, the detection reaction in each microwell can be monitored over time by fluorescence microscopy.