Integrated and finger-actuated microfluidic chip for point-of-care testing of multiple pathogens.

The integration of gel-based loop-mediated isothermal amplification (gLAMP) and finger-actuated microfluidic chip (μFAchip) was developed for the simultaneous detection of various different types of bacterial pathogens. The developed μFAchip consisted of three PDMS layers attached together by two adhesive tapes. Multiple chambers in the top PDMS layer were used for sample preparation, and the corresponding chambers in the bottom PDMS layer was used for long-term storage of LAMP reagents without DNA templates. The thin PDMS layer in the middle contained cross-shaped cuts as finger-actuated valves for fluid control. To reduce operation steps on the chip, such as pipetting and manipulation of samples, Whatman CloneSaver card was pre-embedded in the top chambers for on-chip DNA extraction and purification. Upon a simple press on the top layer, the finger-actuated valve was opened up, allowing DNA samples on the top layer flow into the bottom reaction chambers for gLAMP reaction. For POCT applications, on-chip LAMP reaction and imaging were conducted on a miniaturized peltier heater and a portable fluorescence imaging system respectively. Under the optimized condition, multiple pathogens were detected simultaneously with high selectivity and sensitivity (as low as 1.6 cells). The developed μFAchip provided a rapid and easy-to-operate platform for gLAMP-based pathogen detection, with the potential for in-field detection, especially in areas with limited resources.