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The extreme miniaturization of biological and chemical assays in aqueous-droplet compartments enables spatiotemporal control for large-scale parallel experimentation and can thus permit new capabilities for â € digitizingâ € directed molecular evolution methodologies. We report a remarkably facile bulk method to generate mega-scale monodisperse sub-femtoliter aqueous droplets by electrospray, using a prototype head with super-fine inkjet technology. Moreover, the electrostatic inkjet nozzle that injects the aqueous phase when immersed within an immiscible phase (an optimized oil/surfactant mixture) has the advantage of generating cell-like sub-femtoliter compartments for biomolecule encapsulation and successive biological and chemical reactions. Sub-femtoliter droplets of both liquid (water-in-oil, volumes ranging from 0.2 to 6.4 fL) and gel bead (agarose-in-oil, volume ranging from 0.3 to 15.6 fL) compartments with average sizes of 1.3 1/4m and 1.5 1/4m, respectively, were successfully generated using an inkjet nozzle at a speed of more than 10 5 droplets per second. We demonstrated the applicability of this system by synthesizing fluorescent proteins using a cell-free expression system inside electrosprayed sub-femtoliter droplets at an accelerated rate, thereby extending the utility of in vitro compartmentalization with improved analytical performance for a top-down artificial cellular system.
Sharma, B., Takamura, Y., Shimoda, T., & Biyani, M. (2016). A bulk sub-femtoliter in vitro compartmentalization system using super-fine electrosprays. Scientific Reports, 6. https://doi.org/10.1038/srep26257