A fully automated rotary microfluidic platform for high-throughput multiplex detection of respiratory tract pathogens

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 moving-probe fluorescence read-out, all encapsulated with a disposable microfluidic cartridge and paired with a 9 kg, four-channel 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 (R² = 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. (Figure presented.)