Positive dielectrophoresis-based Raman-activated droplet sorting for culture-free and label-free screening of enzyme function in vivo.

ABSTRACT: The potential of Raman-activated cell sorting (RACS) is inherently limited by conflicting demands for signal quality and sorting throughput. Here, we present positive dielectrophoresis-based Raman-activated droplet sorting (pDEP-RADS), where a periodical pDEP force was exerted to trap fast-moving cells, followed by simultaneous microdroplet encapsulation and sorting. Screening of yeasts for triacylglycerol (TAG) content demonstrated near-theoretical-limit accuracy, ~120 cells min^-1 throughput and full-vitality preservation, while sorting fatty acid degree of unsaturation (FA-DU) featured ~82% accuracy at ~40 cells min^-1. From a yeast library expressing algal diacylglycerol acyltransferases (DGATs), a pDEP-RADS run revealed all reported TAG-synthetic variants and distinguished FA-DUs of enzyme products. Furthermore, two previously unknown DGATs producing low levels of monounsaturated fatty acid-rich TAG were discovered. This first demonstration of RACS for enzyme discovery represents hundred-fold saving in time consumables and labor versus culture-based approaches. The ability to automatically flow-sort resonance Raman-independent phenotypes greatly expands RACS' application.

SUBMITTER: Wang X 

PROVIDER: S-EPMC7413728 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature