Development of a rapid, sensitive detection method for SARS‐CoV‐2 and influenza virus based on recombinase polymerase amplification combined with CRISPR‐Cas12a assay

Abstract Respiratory tract infections are associated with the most common diseases transmitted among people and remain a huge threat to global public health. Rapid and sensitive diagnosis of causative agents is critical for timely treatment and disease control. Here, we developed a novel method based on recombinase polymerase amplification (RPA) combined with CRISPR‐Cas12a to detect three viral pathogens, including SARS‐CoV‐2, influenza A, and influenza B, which cause similar symptom complexes of flu cold in the respiratory tract. The detection method can be completed within 1 h, which is faster than other standard detection methods, and the limit of detection is approximately 10 2 copies/μL. Additionally, this detection system is highly specific and there is no cross‐reactivity with other common respiratory tract pathogens. Based on this assay, we further developed a more simplified RPA/CRISPR‐Cas12a system combined with lateral flow assay on a manual microfluidic chip, which can simultaneously detect these three viruses. This low‐cost detection system is rapid and sensitive, which could be applied in the field and resource‐limited areas without bulky and expensive instruments, providing powerful tools for the point‐of‐care diagnostic.