Publications by Year: 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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

Han, H. - S. ; Cantalupo, P. G. ; Rotem, A. ; Cockrell, S. K. ; Carbonnaux, M. ; Pipas, J. M. ; Weitz, D. A. Whole-Genome Sequencing of a Single Viral Species from a Highly Heterogeneous Sample. Angewandte Chemie 2015, 54, 13985-13988. Publisher's VersionAbstract

Metagenomic studies suggest that only a small fraction of the viruses that exist in nature have been identified and studied. Characterization of unknown viral genomes is hindered by the many genomes populating any virus sample. A new method is reported that integrates drop‐based microfluidics and computational analysis to enable the purification of any single viral species from a complex mixed virus sample and the retrieval of complete genome sequences. By using this platform, the genome sequence of a 5243 bp dsDNA virus that was spiked into wastewater was retrieved with greater than 96 % sequence coverage and more than 99.8 % sequence identity. This method holds great potential for virus discovery since it allows enrichment and sequencing of previously undescribed viruses as well as known viruses.

Tao, Y. ; Rotem, A. ; Zhang, H. ; Cockrell, S. K. ; Koehler, S. A. ; Chang, C. B. ; Ung, W. L. ; Cantalupo, P. G. ; Ren, Y. ; Lin, J. S. ; et al. Artifact-Free Quantification and Sequencing of Rare Recombinant Viruses by Using Drop-Based Microfluidics. ChemBioChem 2015, 16, 2167-2171. Publisher's VersionAbstract

Recombination is an important driver in the evolution of viruses and thus is key to understanding viral epidemics and improving strategies to prevent future outbreaks. Characterization of rare recombinant subpopulations remains technically challenging because of artifacts such as artificial recombinants, known as chimeras, and amplification bias. To overcome this, we have developed a high‐throughput microfluidic technique with a second verification step in order to amplify and sequence single recombinant viruses with high fidelity in picoliter drops. We obtained the first artifact‐free estimate of in vitro recombination rate between murine norovirus strains MNV‐1 and WU20 co‐infecting a cell (Prec=3.3×10−4±2×10−5) for a 1205 nt region. Our approach represents a time‐ and cost‐effective improvement over current methods, and can be adapted for genomic studies requiring artifact‐ and bias‐free selective amplification, such as microbial pathogens, or rare cancer cells.

Licup, A. J. ; Münster, S. ; Sharma, A. ; Sheinman, M. ; Jawerth, L. M. ; Fabry, B. ; Weitz, D. A. ; Mackintosh, F. C. Stress controls the mechanics of collagen networks. Proceedings of the National Academy of Sciences 2015, 112, 9573-9578. Publisher's VersionAbstract

Collagen is the main structural and load-bearing element of various connective tissues, where it forms the extracellular matrix that supports cells. It has long been known that collagenous tissues exhibit a highly nonlinear stress–strain relationship, although the origins of this nonlinearity remain unknown. Here, we show that the nonlinear stiffening of reconstituted type I collagen networks is controlled by the applied stress and that the network stiffness becomes surprisingly insensitive to network concentration. We demonstrate how a simple model for networks of elastic fibers can quantitatively account for the mechanics of reconstituted collagen networks. Our model points to the important role of normal stresses in determining the nonlinear shear elastic response, which can explain the approximate exponential relationship between stress and strain reported for collagenous tissues. This further suggests principles for the design of synthetic fiber networks with collagen-like properties, as well as a mechanism for the control of the mechanics of such networks.

Amstad, E. ; Gopinadhan, M. ; Holtze, C. ; Osuji, C. O. ; Brenner, M. P. ; Spaepen, F. ; Weitz, D. A. Production of amorphous nanoparticles by supersonic spray-drying with a microfluidic nebulator. Science 2015, 349, 956-960. Publisher's VersionAbstract

Amorphous nanoparticles (a-NPs) have physicochemical properties distinctly different from those of the corresponding bulk crystals; for example, their solubility is much higher. However, many materials have a high propensity to crystallize and are difficult to formulate in an amorphous structure without stabilizers. We fabricated a microfluidic nebulator that can produce amorphous NPs from a wide range of materials, even including pure table salt (NaCl). By using supersonic air flow, the nebulator produces drops that are so small that they dry before crystal nuclei can form. The small size of the resulting spray-dried a-NPs limits the probability of crystal nucleation in any given particle during storage. The kinetic stability of the a-NPs—on the order of months—is advantageous for hydrophobic drug molecules.

Soft Poly(dimethylsiloxane) Elastomers from Architecture-Driven Entanglement Free Design
Cai, L. - H. ; Kodger, T. E. ; Guerra, R. E. ; Pegoraro, A. F. ; Rubinstein, M. ; Weitz, D. A. Soft Poly(dimethylsiloxane) Elastomers from Architecture-Driven Entanglement Free Design. Advanced Materials 2015, 27, 5132-5140. Publisher's VersionAbstract

Soft, solvent‐free poly(dimethylsiloxane) elastomers are fabricated by a one‐step process via crosslinking bottlebrush polymers. The bottlebrush architecture prevents the formation of entanglements, resulting in elastomers with precisely controllable low moduli from 1 to 100 kPa, below the lower limit of traditional elastomers; moreover, the solvent‐free nature enables their negligible adhesiveness compared to commercially available silicone products of similar stiffness.

Duncanson, W. J. ; Kodger, T. E. ; Babaee, S. ; Gonzalez, G. ; Weitz, D. A. ; Bertoldi, K. Microfluidic Fabrication and Micromechanics of Permeable and Impermeable Elastomeric Microbubbles. Langmuir 2015, 31, 3489-3493. Publisher's VersionAbstract

We use droplet microfluidics to produce monodisperse elastomeric microbubbles consisting of gas encapsulated in a polydimethylsiloxane shell. These microbubbles withstand large, repeated deformations without rupture. We perform μN-scale compression tests on individual microbubbles and find their response to be highly dependent on the shell permeability; during deformation, the pressure inside impermeable microbubbles increases, resulting in an exponential increase in the applied force. Finite element models are used to interpret and extend these experimental results enabling the design and development of deformable microbubbles with a predictable mechanical response. Such microbubbles can be designed to repeatedly transit through the narrow constrictions found in a porous medium functioning as probes of the local pressure.