High-throughput multiplexed fluorescence-activated droplet sorting

Fluorescence-activated droplet sorting (FADS) is one of the most important features provided by droplet-based microfluidics. However, to date, it does not allow to compete with the high-throughput multiplexed sorting capabilities offered by flow cytometery. Here, we demonstrate the use of a dielectrophoretic-based FADS, allowing to sort up to five different droplet populations simultaneously. Our system provides means to select droplets of different phenotypes in a single experimental run to separate initially heterogeneous populations. Our experimental results are rationalized with the help of a numerical model of the actuation of droplets in electric fields providing guidelines for the prediction of sorting designs for upscaled or downscaled microsystems.Bioanalysis: microfluidic-sorters to make better choicesDroplet-based microfluidics provides new technologies for cell screening by steering cell-containing microdroplets of interest into different channels at the kilohertz frequency. Until now, in conventional microfluidic sorting, droplets are sorted using electric pulses to move them into one of the two channels. Jean–Christophe Baret from the CNRS and University of Bordeaux in France, Valérie Taly from the CNRS and University of Paris Descartes and Tobias Schneider from EPFL in Switzerland have now boosted dielectrophoretic droplet manipulation to allow for multiple selections in a single device. The teams show how emulsified dye-loaded microdroplets are sorted in five different channels depending on fluorescence intensity: a computer algorithm controls the application of electric pulses to direct the droplets into one of the five channels depending on the fluorescence signals. Numerical models unravel the effect of the dielectrophoretic actuation on the droplet trajectories and are now used to predict the sorting conditions for droplet volumes compatible with single-cell encapsulation.

[1]  Dan Luo,et al.  Multiplexed detection of pathogen DNA with DNA-based fluorescence nanobarcodes , 2005, Nature Biotechnology.

[2]  Fabienne Courtois,et al.  Picoliter cell lysate assays in microfluidic droplet compartments for directed enzyme evolution. , 2012, Chemistry & biology.

[3]  R. Ismagilov,et al.  Detecting bacteria and determining their susceptibility to antibiotics by stochastic confinement in nanoliter droplets using plug-based microfluidics. , 2008, Lab on a chip.

[4]  A. Abate,et al.  Ultrahigh-throughput screening in drop-based microfluidics for directed evolution , 2010, Proceedings of the National Academy of Sciences.

[5]  Weihua Li,et al.  Active droplet sorting in microfluidics: a review. , 2017, Lab on a chip.

[6]  Kevin L Holmes,et al.  International Society for the Advancement of Cytometry cell sorter biosafety standards , 2014, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[7]  Adam Sciambi,et al.  Accurate microfluidic sorting of droplets at 30 kHz. , 2015, Lab on a chip.

[8]  Alexander K. Price,et al.  hνSABR: Photochemical Dose–Response Bead Screening in Droplets , 2016, Analytical chemistry.

[9]  J. S. Johnson,et al.  Biocompatible surfactants for water-in-fluorocarbon emulsions. , 2008, Lab on a chip.

[10]  D. Moras,et al.  Quantitative cell-based reporter gene assays using droplet-based microfluidics. , 2010, Chemistry & biology.

[11]  Christoph A. Merten,et al.  High-throughput screening of enzymes by retroviral display using droplet-based microfluidics. , 2010, Chemistry & biology.

[12]  D. Weitz,et al.  Single-cell analysis and sorting using droplet-based microfluidics , 2013, Nature Protocols.

[13]  D. Weitz,et al.  Fluorescence-activated droplet sorting (FADS): efficient microfluidic cell sorting based on enzymatic activity. , 2009, Lab on a chip.

[14]  S. Yao,et al.  Electrostatic charging and control of droplets in microfluidic devices. , 2013, Lab on a chip.

[15]  A. Abate,et al.  Identification and genetic analysis of cancer cells with PCR-activated cell sorting , 2014, Nucleic acids research.

[16]  S. Quake,et al.  A microfabricated fluorescence-activated cell sorter , 1999, Nature Biotechnology.

[17]  Lianbo Yu,et al.  Detection of microRNA Expression in Human Peripheral Blood Microvesicles , 2008, PloS one.

[18]  J. Nielsen,et al.  High-throughput screening for industrial enzyme production hosts by droplet microfluidics. , 2014, Lab on a chip.

[19]  Andrew D Griffiths,et al.  CotA laccase: high-throughput manipulation and analysis of recombinant enzyme libraries expressed in E. coli using droplet-based microfluidics. , 2014, The Analyst.

[20]  J. Sherwood,et al.  Breakup of fluid droplets in electric and magnetic fields , 1988, Journal of Fluid Mechanics.

[21]  M. Konrad,et al.  Parallelized ultra-high throughput microfluidic emulsifier for multiplex kinetic assays. , 2015, Biomicrofluidics.

[22]  Garry P. Nolan,et al.  Simultaneous measurement of multiple active kinase states using polychromatic flow cytometry , 2002, Nature Biotechnology.

[23]  N. Perrimon,et al.  Droplet microfluidic technology for single-cell high-throughput screening , 2009, Proceedings of the National Academy of Sciences.

[25]  John B. Shoven,et al.  I , Edinburgh Medical and Surgical Journal.

[26]  Jie-Long He,et al.  Digital Microfluidics for Manipulation and Analysis of a Single Cell , 2015, International journal of molecular sciences.

[27]  F. Rosenbauer,et al.  Microglia emerge from erythromyeloid precursors via Pu.1- and Irf8-dependent pathways , 2013, Nature Neuroscience.

[28]  H. Keh,et al.  Slow motion of a droplet between two parallel plane walls , 2001 .

[29]  M. Figge,et al.  Real-time image processing for label-free enrichment of Actinobacteria cultivated in picolitre droplets. , 2013, Lab on a chip.

[30]  P. Tabeling,et al.  Droplet-based microfluidics at the femtolitre scale. , 2015, Lab on a chip.

[31]  G. Stephanopoulos,et al.  Microfluidic high-throughput culturing of single cells for selection based on extracellular metabolite production or consumption , 2014, Nature Biotechnology.

[32]  J. Baret,et al.  High throughput single cell counting in droplet-based microfluidics , 2017, Scientific Reports.

[33]  A. deMello,et al.  3D Droplet Microfluidic Systems for High-Throughput Biological Experimentation. , 2015, Analytical chemistry.

[34]  Philip A. Romero,et al.  Dissecting enzyme function with microfluidic-based deep mutational scanning , 2015, Proceedings of the National Academy of Sciences.

[35]  H. Shapiro Practical Flow Cytometry: Shapiro/Flow Cytometry 4e , 2005 .

[36]  Martin Fischlechner,et al.  Ultrahigh-throughput–directed enzyme evolution by absorbance-activated droplet sorting (AADS) , 2016, Proceedings of the National Academy of Sciences.

[37]  Rong Fan,et al.  Protein signaling networks from single cell fluctuations and information theory profiling. , 2011, Biophysical journal.

[38]  Andrew D Griffiths,et al.  A completely in vitro ultrahigh-throughput droplet-based microfluidic screening system for protein engineering and directed evolution. , 2012, Lab on a chip.

[39]  Garry P Nolan,et al.  Fluorescent cell barcoding in flow cytometry allows high-throughput drug screening and signaling profiling , 2006, Nature Methods.

[40]  Christoph A Merten,et al.  Microfluidic train station: highly robust and multiplexable sorting of droplets on electric rails , 2017, Lab on a chip.

[41]  Valerie L. Ng,et al.  Practical Flow Cytometry, 4th Edition , 2004 .

[42]  A. Abate,et al.  Microfluidic droplet enrichment for targeted sequencing , 2015, Nucleic acids research.

[43]  Elinore M Mercer,et al.  Microfluidic sorting of mammalian cells by optical force switching , 2005, Nature Biotechnology.

[44]  Howard M. Shapiro,et al.  Practical Flow Cytometry , 1985 .

[45]  K. Sachs,et al.  Causal Protein-Signaling Networks Derived from Multiparameter Single-Cell Data , 2005, Science.

[46]  Christoph A. Merten,et al.  Droplet-based microfluidic platforms for the encapsulation and screening of Mammalian cells and multicellular organisms. , 2008, Chemistry & biology.

[47]  J. Herskowitz,et al.  Proceedings of the National Academy of Sciences, USA , 1996, Current Biology.

[48]  A. Griffiths,et al.  Droplet-based microfluidics platform for ultra-high-throughput bioprospecting of cellulolytic microorganisms. , 2014, Chemistry & biology.

[49]  Hubert W. Schreier,et al.  Image Correlation for Shape, Motion and Deformation Measurements: Basic Concepts,Theory and Applications , 2009 .

[50]  R. Lathe Phd by thesis , 1988, Nature.

[51]  S. Quake,et al.  An Integrated Microfabricated Cell Sorter , 2022 .

[52]  D A Weitz,et al.  Surface acoustic wave actuated cell sorting (SAWACS). , 2010, Lab on a chip.

[53]  A. Griffiths,et al.  High-throughput screening of filamentous fungi using nanoliter-range droplet-based microfluidics , 2016, Scientific Reports.

[54]  G. G. Stokes "J." , 1890, The New Yale Book of Quotations.

[55]  Kenji Yasuda,et al.  An on-chip imaging droplet-sorting system: a real-time shape recognition method to screen target cells in droplets with single cell resolution , 2017, Scientific Reports.

[56]  Howard A. Stone,et al.  Relaxation and breakup of an initially extended drop in an otherwise quiescent fluid , 1989, Journal of Fluid Mechanics.

[57]  Jonathan M Irish,et al.  Single Cell Profiling of Potentiated Phospho-Protein Networks in Cancer Cells , 2004, Cell.

[58]  W. Bonner,et al.  Cell Sorting: Automated Separation of Mammalian Cells as a Function of Intracellular Fluorescence , 1969, Science.

[59]  Christoph A. Merten,et al.  Functional single-cell hybridoma screening using droplet-based microfluidics , 2012, Proceedings of the National Academy of Sciences.

[60]  Cataldo Guaragnella,et al.  A new algorithm for ball recognition using circle Hough transform and neural classifier , 2004, Pattern Recognit..