Jump to main content
Jump to site search

Issue 12, 2020
Previous Article Next Article

Double emulsion flow cytometry with high-throughput single droplet isolation and nucleic acid recovery

Author affiliations

Abstract

Droplet microfluidics has made large impacts in diverse areas such as enzyme evolution, chemical product screening, polymer engineering, and single-cell analysis. However, while droplet reactions have become increasingly sophisticated, phenotyping droplets by a fluorescent signal and sorting them to isolate individual variants-of-interest at high-throughput remains challenging. Here, we present sdDE-FACS ([s with combining low line]ingle [d with combining low line]roplet [D with combining low line]ouble [E with combining low line]mulsion-FACS), a new method that uses a standard flow cytometer to phenotype, select, and isolate individual double emulsion droplets of interest. Using a 130 μm nozzle at high sort frequency (12–14 kHz), we demonstrate detection of droplet fluorescence signals with a dynamic range spanning 5 orders of magnitude and robust post-sort recovery of intact double emulsion (DE) droplets using 2 commercially-available FACS instruments. We report the first demonstration of single double emulsion droplet isolation with post-sort recovery efficiencies >70%, equivalent to the capabilities of single-cell FACS. Finally, we establish complete downstream recovery of nucleic acids from single, sorted double emulsion droplets via qPCR with little to no cross-contamination. sdDE-FACS marries the full power of droplet microfluidics with flow cytometry to enable a variety of new droplet assays, including rare variant isolation and multiparameter single-cell analysis.

Graphical abstract: Double emulsion flow cytometry with high-throughput single droplet isolation and nucleic acid recovery

Back to tab navigation

Supplementary files

Article information


Submitted
12 Mar 2020
Accepted
20 Mar 2020
First published
17 May 2020

This article is Open Access

Lab Chip, 2020,20, 2062-2074
Article type
Paper

Double emulsion flow cytometry with high-throughput single droplet isolation and nucleic acid recovery

K. K. Brower, C. Carswell-Crumpton, S. Klemm, B. Cruz, G. Kim, S. G. K. Calhoun, L. Nichols and P. M. Fordyce, Lab Chip, 2020, 20, 2062
DOI: 10.1039/D0LC00261E

This article is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported Licence. Material from this article can be used in other publications provided that the correct acknowledgement is given with the reproduced material and it is not used for commercial purposes.

Reproduced material should be attributed as follows:

  • For reproduction of material from NJC:
    [Original citation] - Published by The Royal Society of Chemistry (RSC) on behalf of the Centre National de la Recherche Scientifique (CNRS) and the RSC.
  • For reproduction of material from PCCP:
    [Original citation] - Published by the PCCP Owner Societies.
  • For reproduction of material from PPS:
    [Original citation] - Published by The Royal Society of Chemistry (RSC) on behalf of the European Society for Photobiology, the European Photochemistry Association, and RSC.
  • For reproduction of material from all other RSC journals:
    [Original citation] - Published by The Royal Society of Chemistry.

Information about reproducing material from RSC articles with different licences is available on our Permission Requests page.


Social activity

Search articles by author

Spotlight

Advertisements