One drop at a time: toward droplet microfluidics as a versatile tool for single-cell analysis

Rakszewska, Agata; Tel, Jurjen; Chokkalingam, Venkatachalam; Huck, Wilhelm T. S.

doi:10.1038/am.2014.86

Cited by 103 publications

(91 citation statements)

References 99 publications

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“…[2][3][4] While there has been considerable progress in transferring MTP-based cell screening assays to droplet based protocols, 5,6 implementation of appropriate droplet incubation methods has been mostly neglected. Although stable incubation of microuidic emulsions for up to several weeks was enabled by engineering novel surfactants, 7,8 oxygen supply of droplets has not been a research focus so far.…”

Section: Introductionmentioning

confidence: 99%

Enhanced and homogeneous oxygen availability during incubation of microfluidic droplets

et al. 2015

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Droplet microfluidic-based cell screening has the potential to surpass time-and cost efficiency of established screening platforms by several orders of magnitude, but so far lacks sufficient and homogeneous oxygen supply for large droplet numbers (>10 6 ), which is a key parameter affecting metabolism and growth of encapsulated cells. Here, we describe and validate an approach based on continuous carrier oil recirculation that ensures enhanced and homogeneous oxygen availability during mid and long-term incubation of picoliter droplets retained in a 3D-printed storage device. Using biotechnologically relevant microorganisms, we demonstrate that improved oxygen transfer results in three to eleven-fold increased biomass and highly elevated protein production with minimal interdroplet variation. In fact, obtained yields are comparable to those achieved in conventional cultivation devices, so that screening strategies commonly applied in microtiter plates or shaking flasks can now be scaled down to pL-droplets, which offer highly enhanced throughput. In contrast to mere single-cell screening, this approach allows monoclonal cell and metabolite accumulation inside droplets resulting in elevated read-out signals and reduced variability associated to stochasticity in gene expression.Additionally, the range of screening strategies is broadened, since screening for increased biomass yields or mining for microbial natural products from complex environmental samples can now be targeted with pL-droplets. This development substantially improves the robustness and versatility of droplet-based cell assays, further consolidating pL-droplets as a powerful ultrahigh-throughput experimentation platform.

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Section: Introductionmentioning

confidence: 99%

Enhanced and homogeneous oxygen availability during incubation of microfluidic droplets

et al. 2015

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“…Microfluidic techniques provide a flexible way to manipulate single-cells and to perform various single-cell analyses17181920212223 because they can handle from nanoLiter to femtoLiter amounts of liquid precisely. Droplet microfluidics especially provides an emerging tool for single-cell analysis to isolate single-cells in their individual environment for manipulation212425.…”

mentioning

confidence: 99%

“…Droplet microfluidics especially provides an emerging tool for single-cell analysis to isolate single-cells in their individual environment for manipulation212425. The microfluidic platform developed here is based on the general principle of existing droplet microfluidics systems.…”

mentioning

confidence: 99%

Synchronizing stochastic circadian oscillators in single cells of Neurospora crassa

Deng

Arsenault

Caranica

et al. 2016

Sci Rep

113

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The synchronization of stochastic coupled oscillators is a central problem in physics and an emerging problem in biology, particularly in the context of circadian rhythms. Most measurements on the biological clock are made at the macroscopic level of millions of cells. Here measurements are made on the oscillators in single cells of the model fungal system, Neurospora crassa, with droplet microfluidics and the use of a fluorescent recorder hooked up to a promoter on a clock controlled gene-2 (ccg-2). The oscillators of individual cells are stochastic with a period near 21 hours (h), and using a stochastic clock network ensemble fitted by Markov Chain Monte Carlo implemented on general-purpose graphical processing units (or GPGPUs) we estimated that >94% of the variation in ccg-2 expression was stochastic (as opposed to experimental error). To overcome this stochasticity at the macroscopic level, cells must synchronize their oscillators. Using a classic measure of similarity in cell trajectories within droplets, the intraclass correlation (ICC), the synchronization surface ICC is measured on >25,000 cells as a function of the number of neighboring cells within a droplet and of time. The synchronization surface provides evidence that cells communicate, and synchronization varies with genotype.

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“…This feature makes the microdroplet technology extremely useful for biological applications including the high-throughput investigation of single cells and droplet digital polymerase chain reactions (ddPCR) (Agresti et al, 2010;Baker, 2012;Duncombe et al, 2015;Hindson et al, 2011;Rakszewska et al, 2014;Zinchenko et al, 2014). Droplet microfluidics is also useful for drug research (Sackmann et al, 2014;Vladisavljević et al, 2013), food industry (Neethirajan et al, 2011;Ushikubo et al, 2014;Zanatta et al, 2017), and material sciences for the synthesis and investigation of micro-and nanomaterials such as, e.g.…”

Section: Applications Of Droplet Microfluidicsmentioning

confidence: 99%

Droplet Microfluidics as a Tool for the Generation of Granular Matters and Functional Emulsions

Opalski

Kamiński

Garstecki

2019

KONA

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Emulsion-a liquid dispersed in another liquid-is in many respects very similar to granular matter. In the early 2000s a new technology-droplet microfluidics-began emerging from the wider field of microfluidics. Droplet microfluidics was quickly established as a discipline of science and engineering and has been used for the generation of highly uniform emulsions. The last few years have brought significant advances to the field, directed towards a wide range of applications in material sciences-from synthesis of nanoparticles in droplets to assembling complex droplets and using droplets as templates for ordered materials, with applications in food, cosmetic and diagnostic industries. Droplet microfluidic platforms are also successfully used as analytical tools in molecular biology and biochemistry, in e.g. high-throughput screening, digital assays, encapsulation of single cells, sequencing technologies, and in point-of-care diagnostic applications. This article provides an accessible overview of the physical phenomena observed in multiphase flow at the microscale and the techniques in droplet microfluidics systems. We also present the most interesting applications and potential further directions of research in this fascinating young field of science and engineering.

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One drop at a time: toward droplet microfluidics as a versatile tool for single-cell analysis

Cited by 103 publications

References 99 publications

Enhanced and homogeneous oxygen availability during incubation of microfluidic droplets

Enhanced and homogeneous oxygen availability during incubation of microfluidic droplets

Synchronizing stochastic circadian oscillators in single cells of Neurospora crassa

Droplet Microfluidics as a Tool for the Generation of Granular Matters and Functional Emulsions

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