High-Throughput and Integrated CRISPR/Cas12a-Based Molecular Diagnosis Using a Deep Learning Enabled Microfluidic System.

CRISPR/Cas-based molecular diagnosis demonstrates potent potential for sensitive and rapid pathogen detection, notably in SARS-CoV-2 diagnosis and mutation tracking. Yet, a major hurdle hindering widespread practical use is its restricted throughput, limited integration, and complex reagent preparation. Here, a system, microfluidic multiplate-based ultrahigh throughput analysis of SARS-CoV-2 variants of concern using CRISPR/Cas12a and nonextraction RT-LAMP (mutaSCAN), is proposed for rapid detection of SARS-CoV-2 and its variants with limited resource requirements. With the aid of the self-developed reagents and deep-learning enabled prototype device, our mutaSCAN system can detect SARS-CoV-2 in mock swab samples below 30 min as low as 250 copies/mL with the throughput up to 96 per round. Clinical specimens were tested with this system, the accuracy for routine and mutation testing (22 wildtype samples, 26 mutational samples) was 98% and 100%, respectively. No false-positive results were found for negative (n = 24) samples.