A PMMA microfluidic droplet platform for in vitro protein expression using crude E. coli S30 extract.

Droplet based microfluidics are promising new tools for biological and chemical assays. In this paper, a high throughput and high sensitivity microfluidic droplet platform is described for in vitro protein expression using crude Escherichia coli S30 extract. A flow-focusing polymethylmethacrylate (PMMA) microchip was designed and integrated with different functions involving droplet generation, storage, separation and detection. The material used for the chip is superior to the previously tested polydimethylsiloxane (PDMS) due to its mechanical and chemical properties. Droplet formation characteristics such as size and generation rate are investigated systematically. The effect of surfactants Abil EM90 and Span80 in the oil phase on droplet formation and optical detection is also studied. The performance of the system is demonstrated by the high throughput and stable droplet generation and ultralow detection limit. The robustness of the system is also demonstrated by the successful synthesis of a green fluorescent protein (GFP) using E. coli S30 extract as a source of RNA translation reagents.