Rapid,high-throughput,timely,multiplex diagnosis of respiratory-tract infections still relies on laboratory infrastructure,sequential assays,and trained personnel,thereby delaying targeted therapy and outbreak contain...Rapid,high-throughput,timely,multiplex diagnosis of respiratory-tract infections still relies on laboratory infrastructure,sequential assays,and trained personnel,thereby delaying targeted therapy and outbreak containment.In this study,a Fully Automated rotary microfluidic platform(FA-RMP)for high-throughput multiplex respiratory tract pathogens detection was presented.FA-RMP enables a true“sample-in,result-out”workflow through the integration of swab lysis,reagent partitioning,lyophilized reverse transcription loop-mediated isothermal amplification(RT-LAMP),and movingprobe fluorescence read-out,all encapsulated with a disposable microfluidic cartridge and paired with a 9 kg,fourchannel benchtop reader.The FA-RMP enables parallel processing of 16 independent reactions within 30 min,supporting simultaneous detection of up to 4 distinct clinical samples.Analytical validation using serially diluted Mycoplasma pneumoniae(MP)DNA established a limit of detection(LoD)of 50 copies μL^(-1) and a log-linear correlation between threshold time and template load(R^(2)=0.9528).Testing with eight non-target respiratory pathogens yielded no amplification,confirming high analytical specificity.FA-RMP successfully detected the clinical samples with influenza A,influenza B,and MP,further demonstrating its robust multiplex detection capability.By integrating automated sample preparation,multiplex isothermal amplification and quantitative detection into a portable,high-throughput system,the platform delivers laboratory-grade performance at the point of care,serving as a scalable tool for routine respiratory pathogens screening and rapid epidemic response.展开更多
文摘Rapid,high-throughput,timely,multiplex diagnosis of respiratory-tract infections still relies on laboratory infrastructure,sequential assays,and trained personnel,thereby delaying targeted therapy and outbreak containment.In this study,a Fully Automated rotary microfluidic platform(FA-RMP)for high-throughput multiplex respiratory tract pathogens detection was presented.FA-RMP enables a true“sample-in,result-out”workflow through the integration of swab lysis,reagent partitioning,lyophilized reverse transcription loop-mediated isothermal amplification(RT-LAMP),and movingprobe fluorescence read-out,all encapsulated with a disposable microfluidic cartridge and paired with a 9 kg,fourchannel benchtop reader.The FA-RMP enables parallel processing of 16 independent reactions within 30 min,supporting simultaneous detection of up to 4 distinct clinical samples.Analytical validation using serially diluted Mycoplasma pneumoniae(MP)DNA established a limit of detection(LoD)of 50 copies μL^(-1) and a log-linear correlation between threshold time and template load(R^(2)=0.9528).Testing with eight non-target respiratory pathogens yielded no amplification,confirming high analytical specificity.FA-RMP successfully detected the clinical samples with influenza A,influenza B,and MP,further demonstrating its robust multiplex detection capability.By integrating automated sample preparation,multiplex isothermal amplification and quantitative detection into a portable,high-throughput system,the platform delivers laboratory-grade performance at the point of care,serving as a scalable tool for routine respiratory pathogens screening and rapid epidemic response.
