Publications

2016
Lee, H. ; Choi, C. - H. ; Abbaspourrad, A. ; Wesner, C. ; Caggioni, M. ; Zhu, T. ; Weitz, D. A. Encapsulation and Enhanced Retention of Fragrance in Polymer Microcapsules. ACS Applied Materials & Interfaces 2016, 8 4007-4013. Publisher's VersionAbstract

Fragrances are amphiphilic and highly volatile, all of which makes them a challenging cargo to efficiently encapsulate and retain in microcapsules using traditional approaches. We address these limitations by introducing a new strategy that combines bulk and microfluidic emulsification: a stable fragrance-in-water (F/W) emulsion that is primarily prepared from bulk emulsification is incorporated within a polymer microcapsule via microfluidic emulsification. On the basis of the in-depth study of physicochemical properties of the microcapsules on fragrance leakage, we demonstrate that enhanced retention of fragrance can be achieved by using a polar polymeric shell and forming a hydrogel network within the microcapsule. We further extend the utility of these microcapsules by demonstrating the enhanced retention of encapsulated fragrance in powder state.

2016_acsappliedmaterialsinterface_lee.pdf
Verissimo, C. ; Abbaspourrad, A. ; Sunter, K. ; Capasso, F. ; Moshkalev, S. ; Weitz, D. Hydrothermal Synthesis and Properties of Hierarchical ZnO and Eu-doped ZnO 3D Structures. In une; 2016; Vol. 13, pp. 15. 2016_verissimo.pdf
Spencer, S. J. ; Tamminen, M. V. ; Preheim, S. P. ; Guo, M. T. ; Briggs, A. W. ; Brito, I. L. ; Weitz, D. A. ; Pitkanen, L. K. ; Vigneault, F. ; Virta, M. P. J. ; et al. Massively parallel sequencing of single cells by epicPCR links functional genes with phylogenetic markers. ISME JOURNAL 2016, 10, 427-436. Publisher's VersionAbstract

Many microbial communities are characterized by high genetic diversity. 16S ribosomal RNA sequencing can determine community members, and metagenomics can determine the functional diversity, but resolving the functional role of individual cells in high throughput remains an unsolved challenge. Here, we describe epicPCR (Emulsion, Paired Isolation and Concatenation PCR), a new technique that links functional genes and phylogenetic markers in uncultured single cells, providing a throughput of hundreds of thousands of cells with costs comparable to one genomic library preparation. We demonstrate the utility of our technique in a natural environment by profiling a sulfate-reducing community in a freshwater lake, revealing both known sulfate reducers and discovering new putative sulfate reducers. Our method is adaptable to any conserved genetic trait and translates genetic associations from diverse microbial samples into a sequencing library that answers targeted ecological questions. Potential applications include identifying functional community members, tracing horizontal gene transfer networks and mapping ecological interactions between microbial cells.

2016_isme_spencer.pdf
Wagner, O. ; Thiele, J. ; Weinhart, M. ; Mazutis, L. ; Weitz, D. A. ; Huck, W. T. S. ; Haag, R. Biocompatible fluorinated polyglycerols for droplet microfluidics as an alternative to PEG-based copolymer surfactants. Lab on a Chip 2016, 16, 65–69. Publisher's VersionAbstract

In droplet-based microfluidics, non-ionic, high-molecular weight surfactants are required to stabilize droplet interfaces. One of the most common structures that imparts stability as well as biocompatibility to water-in-oil droplets is a triblock copolymer surfactant composed of perfluoropolyether (PFPE) and polyethylene glycol (PEG) blocks. However, the fast growing applications of microdroplets in biology would benefit from a larger choice of specialized surfactants. PEG as a hydrophilic moiety, however, is a very limited tool in surfactant modification as one can only vary the molecular weight and chain-end functionalization. In contrast, linear polyglycerol offers further side-chain functionalization to create custom-tailored, biocompatible droplet interfaces. Herein, we describe the synthesis and characterization of polyglycerol-based triblock surfactants with tailored side-chain composition, and exemplify their application in cell encapsulation and in vitro gene expression studies in droplet-based microfluidics.

2016_labchip_wagner.pdf
Park, J. - S. ; Kim, D. ; Shin, J. H. ; Weitz, D. A. Efficient nematode swimming in a shear thinning colloidal suspension. Soft Matter 2016, 12, 1892-1897. Publisher's VersionAbstract

The swimming behavior of a nematode Caenorhabditis elegans (C. elegans) is investigated in a non-Newtonian shear thinning colloidal suspension. At the onset value ([curly or open phi] [similar] 8%), the suspension begins to exhibit shear thinning behavior, and the average swimming speed of worms jumps by approximately 12% more than that measured in a Newtonian solution exhibiting no shear dependent viscosity. In the shear thinning regime, we observe a gradual yet significant improvement in swimming efficiency with an increase in [curly or open phi] while the swimming speed remains nearly constant. We postulate that this enhanced swimming can be explained by the temporal change in the stroke form of the nematode that is uniquely observed in a shear thinning colloidal suspension: the nematode features a fast and large stroke in its head to overcome the temporally high drag imposed by the viscous medium, whose effective viscosity ([small eta]s) is shown to drop drastically, inversely proportional to the strength of its stroke. Our results suggest new insights into how nematodes efficiently maneuver through the complex fluid environment in their natural habitat.

2016_soft_matter_park.pdf
Henderson, S. J. ; Xia, J. ; Wu, H. ; Stafford, A. R. ; Leslie, B. A. ; Fredenburgh, J. C. ; Weitz, D. A. ; Weitz, J. I. ; others,. Zinc promotes clot stability by accelerating clot formation and modifying fibrin structure. Thrombosis and haemostasis 2016, 115, 533-542. Publisher's VersionAbstract

Zinc released from activated platelets binds fibrin(ogen) and attenuates fibrinolysis. Although zinc also affects clot formation, the mechanism and consequences are poorly understood. To address these gaps, the effect of zinc on clot formation and structure was examined in the absence or presence of factor (F) XIII. Zinc accelerated a) plasma clotting by 1.4-fold, b) fibrinogen clotting by 3.5- and 2.3-fold in the absence or presence of FXIII, respectively, c) fragment X clotting by 1.3-fold, and d) polymerisation of fibrin monomers generated with thrombin or batroxobin by 2.5– and 1.8-fold, respectively. Whereas absorbance increased up to 3.3-fold when fibrinogen was clotted in the presence of zinc, absorbance of fragment X clots was unaffected by zinc, consistent with reports that zinc binds to the αC-domain of fibrin(ogen). Scanning electron microscopic analysis revealed a twofold increase in fibre diameter in the presence of zinc and in permeability studies, zinc increased clot porosity by 30-fold with or without FXIII. Whereas FXIII increased clot stiffness from 128 ± 19 Pa to 415 ± 27 Pa in rheological analyses, zinc reduced clot stiffness by 10– and 8.5-fold in the absence and presence of FXIII, respectively. Clots formed in the presence of zinc were more stable and resisted rupture with or without FXIII. Therefore, zinc accelerates clotting and reduces fibrin clot stiffness in a FXIII-independent manner, suggesting that zinc may work in concert with FXIII to modulate clot strength and stability.

2016_thrombosis_haemostasis_henderson.pdf
2015
The Oxford Handbook of Soft Condensed Matter; Terentjev, E. M. ; Weitz, D. A., Ed. Oxford University Press, 2015; pp. 640. Publisher's VersionAbstract

This Handbook serves both as an introduction and an overview of the field of soft condensed matter. The discussion covers topics ranging from the fundamentals of colloid science to the principles and action of surfactants, modern directions of research in liquid crystals, and the key properties of foams. The book also explores the fundamental physics that controls the structure and mechanics of granular matter; how the unusual and often dramatic mechanical properties of concentrated polymer systems are determined by the physics of entanglements; the complex structures formed by block copolymers and the methods of structure analysis; rubber elasticity and new emerging classes of rubber-elastic materials; the physics of polyelectrolytes; the solvent dynamics in polymer gels, in equilibrium and under mechanical stress; the hierarchical structure and characteristics of an extracellular matrix; and the hierarchical structure and resulting physical properties of the cell cytoskeleton. The book concludes with an analysis of the properties of interfaces and membranes.

Sharma, Y. ; Vargas, D. A. ; Pegoraro, A. F. ; Lepzelter, D. ; Weitz, D. A. ; Zaman, M. H. Collective motion of mammalian cell cohorts in 3D. Integrative Biology 2015, 7 1526–1533. Publisher's VersionAbstract

Collective cell migration is ubiquitous in biology, from development to cancer; it occurs in complex systems comprised of heterogeneous cell types, signals and matrices, and requires large scale regulation in space and time. Understanding how cells achieve organized collective motility is crucial to addressing cellular and tissue function and disease progression. While current two-dimensional model systems recapitulate the dynamic properties of collective cell migration, quantitative three-dimensional equivalent model systems have proved elusive. To establish such a model system, we study cell collectives by tracking individuals within cell cohorts embedded in three dimensional collagen scaffolding. We develop a custom algorithm to quantify the temporal and spatial heterogeneity of motion in cell cohorts during motility events. In the absence of external driving agents, we show that these cohorts rotate in short bursts, <2 hours, and translate for up to 6 hours. We observe, track, and analyze three dimensional motion of cell cohorts composed of 3–31 cells, and pave a path toward understanding cell collectives in 3D as a complex emergent system.

2015_ib_sharma.pdf
Amstad, E. ; Spaepen, F. ; Weitz, D. A. Crystallization of undercooled liquid fenofibrate. Physical Chemistry Chemical Physics 2015, 17, 30158–30161. Publisher's VersionAbstract

Formulation of hydrophobic drugs as amorphous materials is highly advantageous as this increases their solubility in water and therefore their bioavailability. However, many drugs have a high propensity to crystallize during production and storage, limiting the usefulness of amorphous drugs. We study the crystallization of undercooled liquid fenofibrate, a model hydrophobic drug. Nucleation is the rate-limiting step; once seeded with a fenofibrate crystal, the crystal rapidly grows by consuming the undercooled liquid fenofibrate. Crystal growth is limited by the incorporation of molecules into its surface. As nucleation and growth both entail incorporation of molecules into the surface, this process likely also limits the formation of nuclei and thus the crystallization of undercooled liquid fenofibrate, contributing to the good stability of undercooled liquid fenofibrate against crystallization.

2015_pccp_amstad.pdf
Rotem, A. ; Ram, O. ; Shoresh, N. ; Sperling, R. A. ; Goren, A. ; Weitz, D. A. ; Bernstein, B. E. Single-cell ChIP-seq reveals cell subpopulations defined by chromatin state. Nature biotechnology 2015, 33, 1165–1172. Publisher's VersionAbstract

Chromatin profiling provides a versatile means to investigate functional genomic elements and their regulation. However, current methods yield ensemble profiles that are insensitive to cell-to-cell variation. Here we combine microfluidics, DNA barcoding and sequencing to collect chromatin data at single-cell resolution. We demonstrate the utility of the technology by assaying thousands of individual cells and using the data to deconvolute a mixture of ES cells, fibroblasts and hematopoietic progenitors into high-quality chromatin state maps for each cell type. The data from each single cell are sparse, comprising on the order of 1,000 unique reads. However, by assaying thousands of ES cells, we identify a spectrum of subpopulations defined by differences in chromatin signatures of pluripotency and differentiation priming. We corroborate these findings by comparison to orthogonal single-cell gene expression data. Our method for single-cell analysis reveals aspects of epigenetic heterogeneity not captured by transcriptional analysis alone.

2015_natbiotech_rotem.pdf
Vogel, N. ; Utech, S. ; England, G. T. ; Shirman, T. ; Phillips, K. R. ; Koay, N. ; Burgess, I. B. ; Kolle, M. ; Weitz, D. A. ; Aizenberg, J. Color from hierarchy: Diverse optical properties of micron-sized spherical colloidal assemblies. Proceedings of the National Academy of Sciences 2015, 112, 10845–10850. Publisher's VersionAbstract

Materials in nature are characterized by structural order over multiple length scales have evolved for maximum performance and multifunctionality, and are often produced by self-assembly processes. A striking example of this design principle is structural coloration, where interference, diffraction, and absorption effects result in vivid colors. Mimicking this emergence of complex effects from simple building blocks is a key challenge for man-made materials. Here, we show that a simple confined self-assembly process leads to a complex hierarchical geometry that displays a variety of optical effects. Colloidal crystallization in an emulsion droplet creates micron-sized superstructures, termed photonic balls. The curvature imposed by the emulsion droplet leads to frustrated crystallization. We observe spherical colloidal crystals with ordered, crystalline layers and a disordered core. This geometry produces multiple optical effects. The ordered layers give rise to structural color from Bragg diffraction with limited angular dependence and unusual transmission due to the curved nature of the individual crystals. The disordered core contributes nonresonant scattering that induces a macroscopically whitish appearance, which we mitigate by incorporating absorbing gold nanoparticles that suppress scattering and macroscopically purify the color. With increasing size of the constituent colloidal particles, grating diffraction effects dominate, which result from order along the crystal’s curved surface and induce a vivid polychromatic appearance. The control of multiple optical effects induced by the hierarchical morphology in photonic balls paves the way to use them as building blocks for complex optical assemblies—potentially as more efficient mimics of structural color as it occurs in nature.

2015_pnas_vogel.pdf
Zhang, H. ; Cockrell, S. K. ; Kolawole, A. O. ; Rotem, A. ; Serohijos, A. W. R. ; Chang, C. B. ; Tao, Y. ; Mehoke, T. S. ; Han, Y. ; Lin, J. S. ; et al. Isolation and analysis of rare norovirus recombinants from co-infected mice using drop-based microfluidics. Journal of virology 2015, JVI–01137. Publisher's VersionAbstract

Human noroviruses (HuNoVs) are positive-sense RNA viruses that can cause severe, highly infectious gastroenteritis. HuNoV outbreaks are frequently associated with recombination between circulating strains. Strain genotyping and phylogenetic analyses show that noroviruses often recombine in a highly conserved region near the junction of the viral polyprotein (open reading frame 1 [ORF1]) and capsid (ORF2) genes and occasionally within the RNA-dependent RNA polymerase (RdRP) gene. Although genotyping methods are useful for tracking changes in circulating viral populations, they report only the dominant recombinant strains and do not elucidate the frequency or range of recombination events. Furthermore, the relatively low frequency of recombination in RNA viruses has limited studies to cell culture or in vitro systems, which do not reflect the complexities and selective pressures present in an infected organism. Using two murine norovirus (MNV) strains to model coinfection, we developed a microfluidic platform to amplify, detect, and recover individual recombinants following in vitro and in vivo coinfection. One-step reverse transcriptase PCR (RT-PCR) was performed in picoliter drops with primers that identified the wild-type and recombinant progenies and scanned for recombination breakpoints at ∼1-kb intervals. We detected recombination between MNV strains at multiple loci spanning the viral protease, RdRP, and capsid ORFs and isolated individual recombinant RNA genomes that were present at a frequency of 1/300,000 or higher. This study is the first to examine norovirus recombination following coinfection of an animal and suggests that the exchange of RNA among viral genomes in an infected host occurs in multiple locations and is an important driver of genetic diversity.

2015_jvi_zhang.pdf
Abbaspourrad, A. ; Zhang, H. ; Tao, Y. ; Cui, N. ; Asahara, H. ; Zhou, Y. ; Yue, D. ; Koehler, S. A. ; Ung, W. L. ; Heyman, J. ; et al. Label-free single-cell protein quantification using a drop-based mix-and-read system. Scientific reports 2015, 5 12756. Publisher's VersionAbstract

Quantitative protein analysis of single cells is rarely achieved due to technical difficulties of detecting minute amounts of proteins present in one cell. We develop a mix-and-read assay for drop-based label-free protein analysis of single cells. This high-throughput method quantifies absolute, rather than relative, amounts of proteins and does not involve antibody labeling or mass spectrometry.

2015_srep_abbaspourrad.pdf
Utech, S. ; Prodanovic, R. ; Mao, A. S. ; Ostafe, R. ; Mooney, D. J. ; Weitz, D. A. Microfluidic Generation of Monodisperse, Structurally Homogeneous Alginate Microgels for Cell Encapsulation and 3D Cell Culture. Advanced healthcare materials 2015, 4 1628–1633. Publisher's VersionAbstract

Recent studies have shown that basic cellular behavior varies significantly between two- and three-dimensional culture systems. To identify the origins of these fundamental differences the design of reliable and precisely controlled environments is essential. While 2D cell culture is a well-established technique, the fabrication of defined three-dimensional culture models is still challenging. We present a new method for the microfluidic generation of a micron-sized three-dimensional cell culture system. We use a triggered ionic crosslink formation to generate highly monodisperse and structurally homogeneous alginate microbeads. Aqueous droplets containing a mixture of alginate and a water-soluble calcium-EDTA complex are formed by droplet-based microfluidics. In their complexed form, the calcium ions are homogenously distributed inside the droplet but not accessible for the crosslinking process. Acid addition is used to trigger the degradation of the complex, releasing calcium ions on demand that can physically crosslink the alginate chains. A homogeneous hydrogel network is thus generated which can be transferred into an aqueous environment without losing its structural integrity. Single cells can be encapsulated into these controlled microenvironments which provide structural support while allowing for continuous nutrient supply. We encapsulate individual mesenchymal stem cells (MSCs) into the microbeads which show the aspired cell growth and proliferation.

2015_advancedhealthcarematerials_utech.pdf
Mazutis, L. ; Vasiliauskas, R. ; Weitz, D. A. Microfluidic Production of Alginate Hydrogel Particles for Antibody Encapsulation and Release. Macromolecular Bioscience 2015, 15, 1641-1646. Publisher's VersionAbstract

Owing to their biocompatibility and reduced side effects, natural polymers represent an attractive choice for producing drug delivery systems. Despite few successful examples, however, the production of monodisperse biopolymer-based particles is often hindered by high viscosity of polymer fluids. In this work, we present a microfluidic approach for production of alginate-based particles carrying encapsulated antibodies. We use a triple-flow micro-device to induce hydrogel formation inside droplets before their collection off-chip. The fast mixing and gelation process produced alginate particles with a unique biconcave shape and dimensions of the mammalian cells. We show slow and fast dissolution of particles in different buffers and evaluate antibody release over time.

2015_macromolecularbioscience_mazutis.pdf
Chang, C. B. ; Wilking, J. N. ; Kim, S. - H. ; Shum, H. C. ; Weitz, D. A. Monodisperse Emulsion Drop Microenvironments for Bacterial Biofilm Growth. Small 2015, 11, 3954–3961. Publisher's VersionAbstract

In this work, microfluidic technology is used to rapidly create hundreds of thousands of monodisperse double and triple emulsion drops that serve as 3D microenvironments for the containment and growth of bacterial biofilms. The size of these drops, with diameters from tens to hundreds of micrometers, makes them amenable to rapid manipulation and analysis. This is demonstrated by using microscopy to visualize cellular differentiation of Bacillus subtilis biofilm communities within each drop and the bacterial biofilm microstructure. Biofilm growth is explored upon specific interfaces in double and triple emulsions and upon negative and positive radii of curvature. Biofilm attachment of matrix and flagella mutants is studied as well as biofilms of Pseudomonas aeruginosa. This is the first demonstration of biofilms grown in microscale emulsion drops, which serve as both templates and containers for biofilm growth and attachment. These microenvironments have the potential to transform existing high-throughput screening methods for bacterial biofilms.

2015_small_chang.pdf
Wagner, O. ; Zieringer, M. ; Duncanson, W. J. ; Weitz, D. A. ; Haag, R. Perfluoroalkyl-Functionalized Hyperbranched Polyglycerol as Pore Forming Agents and Supramolecular Hosts in Polymer Microspheres. International journal of molecular sciences 2015, 16, 20183–20194. Publisher's VersionAbstract

Perfluoroalkyl-functionalized, hyperbranched polyglycerols that produce stable microbubbles are integrated into a microfluidic emulsion to create porous microspheres. In a previously-presented work a dendrimer with a perfluorinated shell was used. By replacing this dendrimer core with a hyperbranched core and evaluating different core sizes and degrees of fluorinated shell functionalization, we optimized the process to a more convenient synthesis and higher porosities. The new hyperbranched polyglycerol porogens produced more pores and can be used to prepare microspheres with porosity up to 12% (v/v). The presented preparation forms pores with a perfluoroalkyl-functionalized surface that enables the resulting microspheres to act as supramolecular host systems. The microspheres can incorporate gases into the pores and actives in the polymer matrix, while the perfluoroalkylated pore surface can be used to immobilize perfluoro-tagged molecules onto the pores by fluorous-fluorous interaction.

2015_ijms_wagner.pdf
Tao, Y. ; Rotem, A. ; Zhang, H. ; Chang, C. B. ; Basu, A. ; Kolawole, A. O. ; Koehler, S. A. ; Ren, Y. ; Lin, J. S. ; Pipas, J. M. ; et al. Rapid, targeted and culture-free viral infectivity assay in drop-based microfluidics. Lab on a Chip 2015, 15, 3934–3940. Publisher's VersionAbstract

A key viral property is infectivity, and its accurate measurement is crucial for the understanding of viral evolution, disease and treatment. Currently viral infectivity is measured using plaque assays, which involve prolonged culturing of host cells, and whose measurement is unable to differentiate between specific strains and is prone to low number fluctuation. We developed a rapid, targeted and culture-free infectivity assay using high-throughput drop-based microfluidics. Single infectious viruses are incubated in a large number of picoliter drops with host cells for one viral replication cycle followed by in-drop gene-specific amplification to detect infection events. Using murine noroviruses (MNV) as a model system, we measure their infectivity and determine the efficacy of a neutralizing antibody for different variants of MNV. Our results are comparable to traditional plaque-based assays and plaque reduction neutralization tests. However, the fast, low-cost, highly accurate genomic-based assay promises to be a superior method for drug screening and isolation of resistant viral strains. Moreover our technique can be adapted to measuring the infectivity of other pathogens, such as bacteria and fungi.

2015_labchip_tao.pdf
Arriaga, L. R. ; Amstad, E. ; Weitz, D. A. Scalable single-step microfluidic production of single-core double emulsions with ultra-thin shells. Lab on a Chip 2015, 15, 3335–3340. Publisher's VersionAbstract

We report a versatile and robust device for the continuous production of double emulsion drops with very thin shell thicknesses, of about 5% of the radius: for emulsions 50 μm in radius the shells can be as thin as a few micrometers. Importantly, the viscosity of the oil shell can be varied from that of water up to 70 times that of water without compromising device operation. Furthermore, this device can be easily scaled-up as it is made through soft lithography; this may enable the production of industrial quantities of double emulsion drops with ultra-thin shells, which may serve as templates to form capsules with homogeneous shell thicknesses, useful beyond scientific applications.

2015_labchip_arriage.pdf
Park, J. - A. ; Kim, J. H. ; Bi, D. ; Mitchel, J. A. ; Qazvini, N. T. ; Tantisira, K. ; Park, C. Y. ; McGill, M. ; Kim, S. - H. ; Gweon, B. ; et al. Unjamming and cell shape in the asthmatic airway epithelium. Nature materials 2015, 14, 1040–1048. Publisher's VersionAbstract

From coffee beans flowing in a chute to cells remodelling in a living tissue, a wide variety of close-packed collective systems—both inert and living—have the potential to jam. The collective can sometimes flow like a fluid or jam and rigidify like a solid. The unjammed-to-jammed transition remains poorly understood, however, and structural properties characterizing these phases remain unknown. Using primary human bronchial epithelial cells, we show that the jamming transition in asthma is linked to cell shape, thus establishing in that system a structural criterion for cell jamming. Surprisingly, the collapse of critical scaling predicts a counter-intuitive relationship between jamming, cell shape and cell–cell adhesive stresses that is borne out by direct experimental observations. Cell shape thus provides a rigorous structural signature for classification and investigation of bronchial epithelial layer jamming in asthma, and potentially in any process in disease or development in which epithelial dynamics play a prominent role.

2015_natmater_park_ja.pdf

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