title: A bulk sub-femtoliter in vitro compartmentalization system using super-fine electrosprays.
abstract: 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 "digitizing" 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 μm and 1.5 μm, 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.
authors.affiliation: Department of Bioscience and Biotechnology, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan.
authors.forename: Bineet
authors.initials: B
authors.lastname: Sharma
authors.affiliation: Department of Bioscience and Biotechnology, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan.|Center for Single Nanoscale Innovative Devices, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan.
authors.forename: Yuzuru
authors.initials: Y
authors.lastname: Takamura
authors.affiliation: Department of Bioscience and Biotechnology, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan.|Center for Single Nanoscale Innovative Devices, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan.
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Shimoda
authors.affiliation: Department of Bioscience and Biotechnology, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan.|Center for Single Nanoscale Innovative Devices, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa 923-1292, Japan.|Biyani BioSolutions Pvt. Ltd., Biyani Research Group, R-4, Sector 3, Vidhyadhar Nagar, Jaipur 302023, India.
authors.forename: Manish
authors.initials: M
authors.lastname: Biyani