Multiplexed target detection using DNA-binding dye chemistry in droplet digital PCR.

Two years ago, we described the first droplet digital PCR (ddPCR) system aimed at empowering all researchers with a tool that removes the substantial uncertainties associated with using the analogue standard, quantitative real-time PCR (qPCR). This system enabled TaqMan hydrolysis probe-based assays for the absolute quantification of nucleic acids. Due to significant advancements in droplet chemistry and buoyed by the multiple benefits associated with dye-based target detection, we have created a "second generation" ddPCR system compatible with both TaqMan-probe and DNA-binding dye detection chemistries. Herein, we describe the operating characteristics of DNA-binding dye based ddPCR and offer a side-by-side comparison to TaqMan probe detection. By partitioning each sample prior to thermal cycling, we demonstrate that it is now possible to use a DNA-binding dye for the quantification of multiple target species from a single reaction. The increased resolution associated with partitioning also made it possible to visualize and account for signals arising from nonspecific amplification products. We expect that the ability to combine the precision of ddPCR with both DNA-binding dye and TaqMan probe detection chemistries will further enable the research community to answer complex and diverse genetic questions.

Simultaneous patterning of nanoparticles and polymers using an evaporation driven flow in a vapor permeable template.

Nanoparticles and polymers have great potential for lowering cost and increasing functionality of printed sensors and electronics. However, creation of practical devices requires that many of these materials be patterned on a single substrate, and many current patterning processes can only handle a single material at a time, necessitating alignment of serial processing steps. Higher throughput and lower cost can be achieved by patterning multiple materials simultaneously. To this end, the microfluidic molding process is adapted to pattern various nanoparticle and polymer inks simultaneously, in a completely additive manner, with three-dimensional control and high relative positional accuracy between the different materials. A differential template distortion observed in channels containing different inks is analyzed and found to result from pressure force in the template due to flow of a highly viscous and highly concentrated ink in small channels. The resulting optimization between patterning speed and dimensional fidelity is discussed.