Digital Polymerase Chain Reaction (dPCR) is a novel method for the absolute quantification of target nucleic acids. Quantification by dPCR hinges on the fact that the random distribution of molecules in many partitions follows a Poisson distribution. Each partition acts as an individual PCR microreactor and partitions containing amplified target sequences are detected by fluorescence. The proportion of PCR-positive partitions suffices to determine the concentration of the target sequence without a need for calibration. Advances in microfluidics enabled the current revolution of digital quantification by providing efficient partitioning methods. In this review, we compare the fundamental concepts behind the quantification of nucleic acids by dPCR and quantitative real-time PCR (qPCR). We detail the underlying statistics of dPCR and explain how it defines its precision and performance metrics. We review the different microfluidic digital PCR formats, present their underlying physical principles, and analyze the technological evolution of dPCR platforms. We present the novel multiplexing strategies enabled by dPCR and examine how isothermal amplification could be an alternative to PCR in digital assays. Finally, we determine whether the theoretical advantages of dPCR over qPCR hold true by perusing studies that directly compare assays implemented with both methods.

The evolution of carbon dioxide bubbles dissolving in water is experimentally examined using long microchannels. We study the coupling between bubble hydrodynamics and dissolution in confined geometries. The gas impregnation process in liquid produces significant flow rearrangements. Depending on the initial volumetric liquid fraction, three operating regimes are identified, namely saturating, coalescing, and dissolving. The morphological and dynamical transition from segmented to dilute bubbly flows is investigated. Tracking individual bubbles along the flow direction is used to calculate the temporal evolution of the liquid volumetric fraction and the average flow velocity near reference bubbles over long distances. This method allows us to empirically establish the functional relationship between bubble size and velocity. Finally, we examine the implication of this relationship during the coalescing flow regime, which limits the efficiency of the dissolution process.

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We present a novel method for conducting true single-cell encapsulation at very high efficiency for the manipulation of precious samples. Our unique strategy is based on the sequential capture and original encapsulation of single-cells into a series of hydrodynamic traps. We identified two distinct modes of encapsulation and we established their associated design rules. We improved the trapping scheme to reach a near perfect capture efficiency and make it compatible with the encapsulation process. Finally, we developed the complete device operation that permits highly efficient single-cell encapsulation and droplet retrieval. This platform provides the foundation to a fully integrated multiparameter platform that will impact the analysis of tissues at single-cell resolution.