Versatile digital polymerase chain reaction chip design, fabrication, and image processing

Abstract

We present a family of digital polymerase chain reaction (dPCR) chips made of silicon that are (9 × 9) mm2 in size, have a unique layout design, and have between 26,448 and 1,656,000 reaction wells. The chip layout was split into six fields sized 4200 × 2785 μm2 that contain reaction wells with a gap of a few μm separating the fields from each other. We also developed a universal fluorescence imaging method capable of processing images from those chips. It comprised automatic image acquisition, image stitching, alignment mark registration, rotation correction, determination of reaction wells’ positions, and result processing. The chip design, with fully automated image processing, provided a fast and reliable system for the absolute quantitative analysis of dPCR. Combined with ease-of-use, this family of dPCR chips offers a pathway to enable fast and affordable digital microfluidic diagnostic applications.