Monolithically integrated microchannel plate functionalized with ZnO nanorods for fluorescence-enhanced digital polymerase chain reaction

Abstract

In this paper, we report on an integrated microfluidic digital PCR system for rapid and high-performance absolute quantification of DNA at a single-molecular level. Microchannel plate (MCP), a highly porous glass membrane conventionally used as a particle multiplier in detectors, is demonstrated here as an ideal platform for the sample partition and high-fidelity DNA detection. The density of the microreactors reaches up to 1563 mm−2, with a total number of 25,000 chambers each in 100 pL volumes embedded in 4 × 4 mm2 MCP. This glass membrane consisting of highly-regular, densely-packed and high-aspect ratio microchannels is batch-fabricated through fiber-draw technology that requires no cumbersome lithography or etching process. The sidewalls, having a slight tilting angle with respect to the surface normal, can be functionalized with ZnO nanorods through one-step shadowing-based deposition without resorting to complicated solution-process or lithography. In this way, the required thermal cycling time for end-point detection has been reduced from 40 min to 25 min through plasmonic fluorescence enhancement compared to bare MCP without nanostructures. The digital PCR performance of our system has been validated by using bacteriophage λDNA and PLAU genes. The dynamic range achieved is noted as 105 ranging from 1 × 103 to 1 × 107 copies/μL and the measurement deviation is less than 5%. The detection limit is determined to be 1.4 copies/μL. The density of microchambers could be easily scaled for extensive applications and detection ranges by fabricating various MCP matrix structures. Given this high performance and a straightforward fabrication process of MCP, the device is expected to replace conventional PCR equipment for high fidelity and wide dynamic range single-molecule counting.