Microfluidic droplet platform for ultrahigh-throughput single-cell screening of biodiversity.
Proc Natl Acad Sci U S A
; 114(10): 2550-2555, 2017 03 07.
Article
em En
| MEDLINE
| ID: mdl-28202731
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.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Paraoxon
/
Butirilcolinesterase
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Técnicas Analíticas Microfluídicas
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Ensaios de Triagem em Larga Escala
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Análise de Célula Única
Tipo de estudo:
Diagnostic_studies
/
Screening_studies
Limite:
Humans
Idioma:
En
Ano de publicação:
2017
Tipo de documento:
Article