Despite the advantages of digital nucleic acid analysis (DNAA) in terms of sensitivity, precision, and resolution, current DNAA methods commonly suffer a limitation in multiplexing capacity. To address this issue, a droplet encoding‐pairing enabled DNAA multiplexing strategy is developed, wherein unique tricolor combinations are deployed to index individual primer droplets. The template droplets and primer droplets are sequentially introduced into a microfluidic chip with a calabash‐shaped microwell array and are pairwise trapped and merged in the microwells. Pre‐merging and post‐amplification image analysis with a machine learning algorithm is used to identify, enumerate, and address the droplets. By incorporating the amplification signals with droplet encoding information, simultaneous quantitative detection of multiple targets is achieved. This strategy allows for the establishment of flexible multiplexed DNAA by simply adjusting the primer droplet library. Its flexibility is demonstrated by establishing two multiplexed (8‐plex) droplet digital loop‐mediated isothermal amplification (mddLAMP) assays for individually detecting lower respiratory tract infection and urinary tract infection causative pathogens. Clinical sample analysis shows that the microbial detection outcomes of the mddLAMP assays are consistent with those of the conventional assay. This DNAA multiplexing strategy can achieve flexible high‐order multiplexing on demand, making it a desirable tool for high‐content pathogen detection.
The authors would like to call the reader's attention to the fact that, unfortunately, there was a mistake in one of the authors' names, which is 'Zou' instead of 'Zhou'.
Early screening of cancer can effectively prolong survival time and reduce cancer mortality. However, the existing health-monitoring devices can only be carried out in professional laboratories, so large-scale early cancer screening in resource-limited settings is hardly achieved. To embrace the challenge, we developed a novel chemiluminescence immunoassay (CLIA) analyzer that does not require a professional operation. Then, it was applied to detect carbohydrate antigen 50 (CA50), a non–organ-specific tumor marker for screening various cancers. As a result, the analyzer exhibited excellent performance that the total assay time was only 15 min, and the detection limit reached 0.057 U ml−1. A coefficient of variance (CV) less than 15% was well-controlled for both intra- and inter-assay precision, and the linear range was 0–500 U ml−1. More importantly, this analyzer can continuously detect 60 samples per hour without any professional paramedic. Finally, this analyzer has been applied to evaluate clinical samples and the detected results showed a good correlation with the clinical test results (correlation coefficient, 0.9958). These characteristics exactly meet large-scale and high-throughput early screening of cancer. Thus, this miniaturized analyzer for CA50 detection is promising to achieve early large-scale screening of cancer in the resource-limited grassroots community.
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