Ultra-high-throughput screening of enzyme libraries with droplet-based microfluidics

Philip A. Romero, Tuan M. Tran, and Adam R. Abate

Department of Bioengineering and Therapeutic Sciences, University of California, San

Francisco, 1700 4th St., San Francisco, CA 94158

Directed evolution relies on the ability to screen large, diverse libraries of sequence variants. We have developed a microfluidic platform for ultra-high- throughput screening of enzyme libraries. Individual enzyme variants are assayed in a monodisperse water-in-oil microemulsion. Aqueous droplets compartmentalize the enzymatic reaction and maintain the genotype-phenotype linkage. A quantitative fluorescence detection system allows droplets to be analyzed and sorted at rates beyond 1 kHz. This screening platform provides a throughput that is orders of magnitude beyond traditional plate-based screens.