Droplet-based microsystem for multi-step bioreactions.

A droplet-based microfluidic platform was used to perform on-chip droplet generation, merging and mixing for applications in multi-step reactions and assays. Submicroliter-sized droplets can be produced separately from three identical droplet-generation channels and merged together in a single chamber. Three different mixing strategies were used for mixing the merged droplet. For pure diffusion, the reagents were mixed in approximately 10 min. Using flow around the stationary droplet to induce circulatory flow within the droplet, the mixing time was decreased to approximately one minute. The shortest mixing time (10 s) was obtained with bidirectional droplet motion between the chamber and channel, and optimization could result in a total time of less than 1 s. We also tested this on-chip droplet generation and manipulation platform using a two-step thermal cycled bioreaction: nested TaqMan PCR. With the same concentration of template DNA, the two-step reaction in a well-mixed merged droplet shows a cycle threshold of approximately 6 cycles earlier than that in the diffusively mixed droplet, and approximately 40 cycles earlier than the droplet-based regular (single-step) TaqMan PCR.