Lab-In-Fiber Optofluidic Device for Droplet Digital Polymerase Chain Reaction (DdPCR) with Real-Time Monitoring.

ABSTRACT

Droplet microfluidic systems have emerged as indispensable and advanced tools in contemporary biological science. A prominent example is the droplet digital polymerase chain reaction (ddPCR), which plays a pivotal role in next-generation sequencing and the detection of rare nucleic acids or mutations. However, existing optical detection configurations are bulky, intricate, and costly, and require meticulous optical alignment to optimize fluorescence sensing. Herein, we propose a lab-in-fiber optofluidic system (LiFO), which provides a stable and compact footprint, self-alignment, and enhanced optical coupling for high-accuracy ddPCR. Moreover, LiFO could expand its capabilities for multiangle-scattering light collection in which we collect focused forward-scattering light (fFSL) to enable real-time droplet counting and size monitoring. To accomplish these attributes, LiFO incorporates optical fibers, along with fabricated PDMS grooves, for a self-aligned optical setup to implement simultaneous fluorescence and scattering detection. Furthermore, LiFO harnesses the concept of flowing droplets functioning as microlenses, which allows us to collect and translate fFSL signals into droplet size information. We have demonstrated the effectiveness of LiFO in ddPCR applications, illustrating its capacity to enhance the accuracy and precision of DNA quantification. Notably, LiFO exhibits improved linearity in the measurement of serial DNA dilutions, reflected by an increase in R2 from 0.956 to 0.997. These results demonstrate the potential of LiFO to serve as a valuable tool across a wide spectrum of droplet microfluidic platforms, offering opportunities for advancement in practical applications.