Xiong, MengqiuWang, YaoLu, ShuhanLubanga, NasifuLi, TaoLi, ZhihaoHe, BangshunLi, Ying2025-03-142025-03-142025-02-26Xiong, M., Wang, Y., Lu, S., Lubanga, N., Li, T., Li, Z., ... & Li, Y. (2025). Space-coded microchip for multiplexed respiratory virus detection via CRISPR-Cas12a and RPA. Talanta, 127815. https://doi.org/10.1016/j.talanta.2025.1278151873-3573https://dir.muni.ac.ug/handle/20.500.12260/735Multiple infections of respiratory viruses are common in patients with clinical respiratory diseases, but current detection methods still have problems such as complex equipment and long detection time. Rapid, low-cost, and on-site detection of human respiratory viruses is crucial for both clinical diagnosis and population screening. In this research, we created a space-coded microfluidic chip (SC-Chip) for the recognition of nine respiratory viruses: influenza A virus, influenza B virus, severe acute respiratory syndrome coronavirus 2, human coronavirus OC43, human coronavirus NL63, human coronavirus HKU1, human respiratory syncytial virus, human parainfluenza virus, and human metapneumovirus. For the first time, a comprehensive sequence comparison among these viruses was performed to design the recombinase polymerase amplification (RPA) primers and Cas12a-crRNAs. The SC-Chip partitions samples amplified by RPA into spatially coded wells preloaded with CRISPR-Cas12a detection reagents, enabling the identification of all nine viral targets in a single test using a single fluorescence probe. The chip-based assay displays 9 respiratory viruses in less than 40 min with a minimum detection limit at a concentration of 10−18 M (∼1 copy/reaction). Additionally, the efficacy of the method was assessed through its application to 35 clinical patient samples identified as being at risk for respiratory virus infection, yielding a sensitivity of 90 % and a specificity of 100 %. In summary, this space-coded microfluidic CRISPR system offers several advantages, including ease of operation, cost-effectiveness, and rapid data acquisition, thereby holding great potential for multiplexed detection of nucleic acid targets in a clinical setting.enCRISPR-Cas12aMicrofluidicsRespiratory virusesNucleic acid detectionRPA (Recombinase Polymerase Amplification)Article