Handheld RPA-based molecular POCT system for rapid, low-cost 8-plexed detection of respiratory pathogens at home

Area of Science:

• Biomedical Engineering
• Molecular Diagnostics
• Point-of-Care Testing

Background:

• Rapid diagnosis of respiratory pathogens is critical during outbreaks.
• Existing point-of-care testing (POCT) systems are often complex and costly for home use.
• Need for accessible, user-friendly home-based molecular diagnostic tools.

Purpose of the Study:

• To develop a novel microfluidic cartridge-based system for multiplexed home-based detection of respiratory pathogens.
• To overcome limitations of current POCT systems for home use.
• To enable rapid, accurate, and cost-effective diagnosis of respiratory infections in non-clinical settings.

Main Methods:

• A microfluidic cartridge with three parallel channels was designed, integrating lysis, RPA, and air storage chambers.
• Extraction-free, 3-plex recombinase polymerase amplification (RPA) assays were performed in each channel.
• A pneumatic pressure pumping strategy with hydraulic resistance matching ensured precise flow control and uniform reagent distribution.
• A dynamic mixing method facilitated homogeneous mixing of samples and reagents.

Main Results:

• The system achieved simultaneous identification of eight respiratory pathogens and one internal control in under 25 minutes.
• The microfluidic system is compact (<1 kg), lightweight, and has low power consumption (3 W).
• The cost per test is remarkably low ($1.4), enhancing accessibility.
• Validation with 356 nasopharyngeal swabs demonstrated high sensitivity (>97%) and specificity (>99%).

Conclusions:

• The developed microfluidic system offers a practical and accessible solution for home-based multiplexed molecular POCT.
• It enables reliable, rapid, and cost-effective diagnosis of respiratory co-infections.
• The system's user-friendliness and performance characteristics make it suitable for vulnerable populations and home environments.