Microfluidic droplet platform for ultrahigh-throughput single-cell screening of biodiversity

Stanislav S. Terekhov, Ivan V. Smirnov, Anastasiya V. Stepanova, Tatyana V. Bobik, Yuliana A. Mokrushina, Natalia A. Ponomarenko, Alexey A. Belogurov, Maria P. Rubtsova, Olga V. Kartseva, Marina O. Gomzikova, Alexey A. Moskovtsev, Anton S. Bukatin, Michael V. Dubina, Elena S. Kostryukova, Vladislav V. Babenko, Maria T. Vakhitova, Alexander I. Manolov, Maja V. Malakhova, Maria A. Kornienko, Alexander V. TyakhtAnna A. Vanyushkina, Elena N. Ilina, Patrick Masson, Alexander G. Gabibov, Sidney Altman

    Research output: Contribution to journalArticlepeer-review

    130 Citations (Scopus)

    Abstract

    Ultrahigh-throughput screening (uHTS) techniques can identify unique functionality from millions of variants. To mimic the natural selection mechanisms that occur by compartmentalization in vivo, we developed a technique based on single-cell encapsulation in droplets of a monodisperse microfluidic double water-in-oil-in-water emulsion (MDE). Biocompatible MDE enables in-droplet cultivation of different living species. The combination of droplet-generating machinery with FACS followed by next-generation sequencing and liquid chromatography-mass spectrometry analysis of the secretomes of encapsulated organisms yielded detailed genotype/phenotype descriptions. This platform was probed with uHTS for biocatalysts anchored to yeast with enrichment close to the theoretically calculated limit and cell-to-cell interactions. MDE-FACS allowed the identification of human butyrylcholinesterase mutants that undergo self-reactivation after inhibition by the organophosphorus agent paraoxon. The versatility of the platform allowed the identification of bacteria, including slow-growing oral microbiota species that suppress the growth of a common pathogen, Staphylococcus aureus, and predicted which genera were associated with inhibitory activity.

    Original languageEnglish
    Pages (from-to)2550-2555
    Number of pages6
    JournalProceedings of the National Academy of Sciences of the United States of America
    Volume114
    Issue number10
    DOIs
    Publication statusPublished - 7 Mar 2017

    Keywords

    • Butyrylcholinesterase
    • Cell-cell interactions
    • Microfluidic encapsulation
    • Staphylococcus aureus
    • Ultrahigh-throughput screening

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