Droplet Encoding‐Pairing Enabled Multiplexed Digital Loop‐Mediated Isothermal Amplification for Simultaneous Quantitative Detection of Multiple Pathogens

• DOI: 10.1002/advs.202205863
• Publication Date: 2023-01-17T02:39:47Z
• Authors: Dongyang Cai; Yu Wang; Jingjing Zou; Zhujun Li; Enqi Huang; Xiuyun Ouyang; Zhiquan Que; Yanzhang Luo; Zhenhua Chen; Yanqing Jiang; Guohao Zhang; Hongkai Wu; Dayu Liu
• Keywords: Isothermal process

Abstract Despite the advantages of digital nucleic acid analysis (DNAA) in terms sensitivity, precision, and resolution, current DNAA methods commonly suffer a limitation multiplexing capacity. To address this issue, droplet encoding‐pairing enabled strategy is developed, wherein unique tricolor combinations are deployed to index individual primer droplets. The template droplets sequentially introduced into microfluidic chip with calabash‐shaped microwell array pairwise trapped merged microwells. Pre‐merging post‐amplification image machine learning algorithm used identify, enumerate, By incorporating amplification signals encoding information, simultaneous quantitative detection multiple targets achieved. This allows for establishment flexible multiplexed by simply adjusting library. Its flexibility demonstrated establishing two (8‐plex) loop‐mediated isothermal (mddLAMP) assays individually detecting lower respiratory tract infection urinary causative pathogens. Clinical sample shows that microbial outcomes mddLAMP consistent those conventional assay. can achieve high‐order on demand, making it desirable tool high‐content pathogen detection.