Flow cytometry-based ultra-high-throughput screening assay for cellulase activity

We have developed a novel, ultra-high-throughput screening assay for the detection of cellulase activity based on fluorescence-activated cell sorting (FACS) and double emulsion technology. Cellulase activity is detected using a series of coupled enzymes, including hexose oxidase (HOx), which generates hydrogen peroxide from the reducing sugars released by cellulases in the presence of any natural or artificial sub- strate. The assay can be adapted to suit a microtiter plate format, but the highest throughput is achieved by using FACS to screen high-complexity cellulase clone libraries. Using this approach, we achieved a 12- fold enrichment of positive (cellulase-expressing) cells in cellulase reference libraries after just one sort- ing round.