Publications
172 results found
Trantidou T, Friddin M, Salehi-Reyhani S, et al., 2018, Droplet microfluidics for the construction of compartmentalised model membranes, Lab on a Chip, Vol: 18, Pages: 2488-2509, ISSN: 1473-0189
The design of membrane-based constructs with multiple compartments is of increasing importance given their potential applications as microreactors, as artificial cells in synthetic-biology, as simplified cell models, and as drug delivery vehicles. The emergence of droplet microfluidics as a tool for their construction has allowed rapid scale-up in generation throughput, scale-down of size, and control over gross membrane architecture. This is true on several levels: size, level of compartmentalisation and connectivity of compartments can all be programmed to various degrees. This tutorial review explains and explores the reasons behind this. We discuss microfluidic strategies for the generation of a family of compartmentalised systems that have lipid membranes as the basic structural motifs, where droplets are either the fundamental building blocks, or are precursors to the membrane-bound compartments. We examine the key properties associated with these systems (including stability, yield, encapsulation efficiency), discuss relevant device fabrication technologies, and outline the technical challenges. In doing so, we critically review the state-of-play in this rapidly advancing field.
Chatzimichail S, Supramaniam P, Ces O, et al., 2018, Counting proteins in single cells with addressable droplet microarrays, Jove-Journal of Visualized Experiments, Vol: 137, ISSN: 1940-087X
Often cellular behaviour and cellular responses are analysed at the population level where the responses of many cells are pooled together as an average result masking the rich single cell behaviour within a complex population. Single cell protein detection and quantification technologies have made a remarkable impact in recent years. Here we describe a practical and flexible single cell analysis platform based on addressable droplet microarrays. This study describes how the absolute copy numbers of target proteins may be measured with single cell resolution. The tumour suppressor p53 is the most commonly mutated gene in human cancer, with more than 50% of total cancer cases exhibiting a non-healthy p53 expression pattern. The protocol describes steps to create 10nL droplets within which single human cancer cells are isolated and the copy number of p53 protein is measured with single molecule resolution to precisely determine the variability in expression. The method may be applied to any cell type including primary material to determine the absolute copy number of any target proteins of interest.
Bolognesi G, Friddin MS, Salehi-Reyhani S, et al., 2018, Sculpting and fusing biomimetic vesicle networks using optical tweezers, Nature Communications, Vol: 9, Pages: 1-11, ISSN: 2041-1723
Constructing higher-order vesicle assemblies has discipline-spanning potential from responsive soft-matter materials to artificial cell networks in synthetic biology. This potential is ultimately derived from the ability to compartmentalise and order chemical species in space. To unlock such applications, spatial organisation of vesicles in relation to one another must be controlled, and techniques to deliver cargo to compartments developed. Herein, we use optical tweezers to assemble, reconfigure and dismantle networks of cell-sized vesicles that, in different experimental scenarios, we engineer to exhibit several interesting properties. Vesicles are connected through double-bilayer junctions formed via electrostatically controlled adhesion. Chemically distinct vesicles are linked across length scales, from several nanometres to hundreds of micrometres, by axon-like tethers. In the former regime, patterning membranes with proteins and nanoparticles facilitates material exchange between compartments and enables laser-triggered vesicle merging. This allows us to mix and dilute content, and to initiate protein expression by delivering biomolecular reaction components.
Karamdad K, Hindley J, Friddin MS, et al., 2018, Engineering thermoresponsive phase separated vesicles formed via emulsion phase transfer as a content-release platform, Chemical Science, Vol: 9, Pages: 4851-4858, ISSN: 2041-6520
Giant unilamellar vesicles (GUVs) are a well-established tool for the study of membrane biophysics and are increasingly used as artificial cell models and functional units in biotechnology. This trend is driven by the development of emulsion-based generation methods such as Emulsion Phase Transfer (EPT), which facilitates the encapsulation of almost any water-soluble compounds (including biomolecules) regardless of size or charge, is compatible with droplet microfluidics, and allows GUVs with asymmetric bilayers to be assembled. However, the ability to control the composition of membranes formed via EPT remains an open question; this is key as composition gives rise to an array of biophysical phenomena which can be used to add functionality to membranes. Here, we evaluate the use of GUVs constructed via this method as a platform for phase behaviour studies and take advantage of composition-dependent features to engineer thermally-responsive GUVs. For the first time, we generate ternary GUVs (DOPC/DPPC/cholesterol) using EPT, and by compensating for the lower cholesterol incorporation efficiencies, show that these possess the full range of phase behaviour displayed by electroformed GUVs. As a demonstration of the fine control afforded by this approach, we demonstrate release of dye and peptide cargo when ternary GUVs are heated through the immiscibility transition temperature, and show that release temperature can be tuned by changing vesicle composition. We show that GUVs can be individually addressed and release triggered using a laser beam. Our findings validate EPT as a suitable method for generating phase separated vesicles and provide a valuable proof-of-concept for engineering content release functionality into individually addressable vesicles, which could have a host of applications in the development of smart synthetic biosystems.
Trantidou T, Regoutz A, Voon X, et al., 2018, A “cleanroom-free” and scalable manufacturing technology for the microfluidic generation of lipid-stabilized droplets and cell-sized multisomes, Sensors and Actuators B: Chemical, Vol: 267, Pages: 34-41, ISSN: 0925-4005
There is a growing demand to construct artificial biomimetic structures from the bottom-up using simple chemical components in a controlled and high-throughput way. These cell mimics are encapsulated by lipid membranes and can reconstitute biological machinery within them. To date, such synthetic cells based upon droplet microfluidics are fabricated using non-scalable, expensive and time-consuming strategies, and are thus restricted to small-scale in-house manufacturing. Here, we report a “cleanroom-free” and highly scalable microfluidic manufacturing technology based on dry film resists and multilayer lamination. The technology facilitates the controlled and high-throughput generation of stable and monodisperse droplets using anionic surfactants and more biologically relevant phospholipids. We demonstrate the versatility of this approach by selectively patterning the surface chemistry of the device, enabling the production of compartmentalized lipid structures based on droplet interface bilayers (multisomes). This technology has the potential to simultaneously unlock the widespread exploitation of microfluidics to chemists and synthetic biologists not having access to controlled production environments and facilitate low-cost (< £1) high-volume fabrication of self-contained disposable devices with minimum feature sizes of 30 μm. The associated material and equipment costs approach those of other deskilled prototyping technologies, such as 3D printing that have made the transition into the mainstream.
Hindley JW, Elani Y, McGilvery CM, et al., 2018, Light-triggered enzymatic reactions in nested vesicle reactors, Nature Communications, Vol: 9, Pages: 1-6, ISSN: 2041-1723
Cell-sized vesicles have tremendous potential both as miniaturised pL reaction vessels and in bottom-up synthetic biology as chassis for artificial cells. In both these areas the introduction of light-responsive modules affords increased functionality, for example, to initiate enzymatic reactions in the vesicle interior with spatiotemporal control. Here we report a system composed of nested vesicles where the inner compartments act as phototransducers, responding to ultraviolet irradiation through diacetylene polymerisation-induced pore formation to initiate enzymatic reactions. The controlled release and hydrolysis of a fluorogenic β-galactosidase substrate in the external compartment is demonstrated, where the rate of reaction can be modulated by varying ultraviolet exposure time. Such cell-like nested microreactor structures could be utilised in fields from biocatalysis through to drug delivery.
Elani Y, Trantidou T, Wylie D, et al., 2018, Constructing vesicle-based artificial cells with embedded living cells as organelle-like modules, Scientific Reports, Vol: 8, Pages: 1-8, ISSN: 2045-2322
There is increasing interest in constructing artificial cells by functionalising lipid vesicles with biological and synthetic machinery. Due to their reduced complexity and lack of evolved biochemical pathways, the capabilities of artificial cells are limited in comparison to their biological counterparts. We show that encapsulating living cells in vesicles provides a means for artificial cells to leverage cellular biochemistry, with the encapsulated cells serving organelle-like functions as living modules inside a larger synthetic cell assembly. Using microfluidic technologies to construct such hybrid cellular bionic systems, we demonstrate that the vesicle host and the encapsulated cell operate in concert. The external architecture of the vesicle shields the cell from toxic surroundings, while the cell acts as a bioreactor module that processes encapsulated feedstock which is further processed by a synthetic enzymatic metabolism co-encapsulated in the vesicle.
Heide C, Ces O, Polizzi K, et al., 2018, Creating cell-free protein synthesis factories, Pharmaceutical Bioprocessing, Vol: 6, Pages: 3-6, ISSN: 2048-9145
Barlow NE, Bolognesi G, Haylock S, et al., 2017, Rheological Droplet Interface Bilayers (rheo-DIBs): Probing the Unstirred Water Layer Effect on Membrane Permeability via Spinning Disk Induced Shear Stress, Scientific Reports, Vol: 7, ISSN: 2045-2322
A new rheological droplet interface bilayer (rheo-DIB) device is presented as a tool to apply shear stress on biological lipid membranes. Despite their exciting potential for affecting high-throughput membrane translocation studies, permeability assays conducted using DIBs have neglected the effect of the unstirred water layer (UWL). However as demonstrated in this study, neglecting this phenomenon can cause significant underestimates in membrane permeability measurements which in turn limits their ability to predict key processes such as drug translocation rates across lipid membranes. With the use of the rheo-DIB chip, the effective bilayer permeability can be modulated by applying shear stress to the droplet interfaces, inducing flow parallel to the DIB membranes. By analysing the relation between the effective membrane permeability and the applied stress, both the intrinsic membrane permeability and UWL thickness can be determined for the first time using this model membrane approach, thereby unlocking the potential of DIBs for undertaking diffusion assays. The results are also validated with numerical simulations.
Thomas JM, Friddin MS, Ces O, et al., 2017, Programming membrane permeability using integrated membrane pores and blockers as molecular regulators, Chemical Communications, Vol: 53, Pages: 12282-12285, ISSN: 1359-7345
We report a bottom-up synthetic biology approach to engineering vesicles with programmable permeabilities. Exploiting the concentration-dependent relationship between constitutively active pores (alpha-hemolysin) and blockers allows blockers to behave as molecular regulators for tuning permeability, enabling us to systematically modulate cargo release kinetics without changing the lipid fabric of the system.
de Bruin A, Friddin MS, Elani Y, et al., 2017, A transparent 3D printed device for assembling droplet hydrogel bilayers (DHBs), RSC Advances, Vol: 7, Pages: 47796-47800, ISSN: 2046-2069
We report a new approach for assembling droplet hydrogel bilayers (DHBs) using a transparent 3D printed device. We characterise the transparency of our platform, confirm bilayer formation using electrical measurements and show that single-channel recordings can be obtained using our reusable rapid prototyped device. This method significantly reduces the cost and infrastructure required to develop devices for DHB assembly and downstream study.
Richens JL, Tyler AII, Barriga HMG, et al., 2017, Spontaneous charged lipid transfer between lipid vesicles, Scientific Reports, Vol: 7, ISSN: 2045-2322
An assay to study the spontaneous charged lipid transfer between lipid vesicles is described. A donor/acceptor vesicle system is employed, where neutrally charged acceptor vesicles are uorescentlylabelled with the electrostatic membrane probe Fluoresceinphosphatidylethanolamine (FPE).Upon addition of charged donor vesicles, transfer of negatively charged lipid occurs, resulting ina uorescently detectable change in the membrane potential of the acceptor vesicles. Using this approach we have studied the transfer properties of a range of lipids, varying both the headgroup and the chain length. At the low vesicle concentrations chosen, the transfer follows a rst-order process where lipid monomers are transferred presumably through the aqueous solution phase from donor to acceptor vesicle. The rate of transfer decreases with increasing chain length which is consistent with energy models previously reported for lipid monomer vesicle interactions. Our assay improves on existing methods allowing the study of a range of unmodi ed lipids, continuous monitoring of transfer and simpli ed experimental procedures.
Trantidou T, Friddin M, Elani Y, et al., 2017, Engineering compartmentalized biomimetic micro- and nanocontainers, ACS Nano, Vol: 11, Pages: 6549-6565, ISSN: 1936-086X
Compartmentalization of biological content and function is a key architectural feature in biology, where membrane bound micro- and nanocompartments are used for performing a host of highly specialized and tightly regulated biological functions. The benefit of compartmentalization as a design principle is behind its ubiquity in cells and has led to it being a central engineering theme in construction of artificial cell-like systems. In this review, we discuss the attractions of designing compartmentalized membrane-bound constructs and review a range of biomimetic membrane architectures that span length scales, focusing on lipid-based structures but also addressing polymer-based and hybrid approaches. These include nested vesicles, multicompartment vesicles, large-scale vesicle networks, as well as droplet interface bilayers, and double-emulsion multiphase systems (multisomes). We outline key examples of how such structures have been functionalized with biological and synthetic machinery, for example, to manufacture and deliver drugs and metabolic compounds, to replicate intracellular signaling cascades, and to demonstrate collective behaviors as minimal tissue constructs. Particular emphasis is placed on the applications of these architectures and the state-of-the-art microfluidic engineering required to fabricate, functionalize, and precisely assemble them. Finally, we outline the future directions of these technologies and highlight how they could be applied to engineer the next generation of cell models, therapeutic agents, and microreactors, together with the diverse applications in the emerging field of bottom-up synthetic biology.
Salehi-Reyhani A, Ces O, Elani Y, 2017, Artificial cell mimics as simplified models for the study of cell biology, Experimental Biology and Medicine, Vol: 242, Pages: 1309-1317, ISSN: 1535-3702
Living cells are hugely complex chemical systems composed of a milieu of distinct chemical species (including DNA, proteins, lipids, and metabolites) interconnected with one another through a vast web of interactions: this complexity renders the study of cell biology in a quantitative and systematic manner a difficult task. There has been an increasing drive towards the utilization of artificial cells as cell mimics to alleviate this, a development that has been aided by recent advances in artificial cell construction. Cell mimics are simplified cell-like structures, composed from the bottom-up with precisely defined and tunable compositions. They allow specific facets of cell biology to be studied in isolation, in a simplified environment where control of variables can be achieved without interference from a living and responsive cell. This mini-review outlines the core principles of this approach and surveys recent key investigations that use cell mimics to address a wide range of biological questions. It will also place the field in the context of emerging trends, discuss the associated limitations, and outline future directions of the field.
Trantidou T, Elani Y, Parsons E, et al., 2017, Hydrophilic surface modification of PDMS for droplet microfluidics using a simple, quick, and robust method via PVA deposition, Microsystems and Nanoengineering, Vol: 3, ISSN: 2055-7434
Polydimethylsiloxane (PDMS) is a dominant material in the fabrication of microfluidic devices to generate water-in-oil droplets, particularly lipid-stabilized droplets, because of its highly hydrophobic nature. However, its key property of hydrophobicity has hindered its use in the microfluidic generation of oil-in-water droplets, which requires channels to have hydrophilic surface properties. In this article, we developed, optimized, and characterized a method to produce PDMS with a hydrophilic surface via the deposition of polyvinyl alcohol following plasma treatment and demonstrated its suitability for droplet generation. The proposed method is simple, quick, effective, and low cost and is versatile with respect to surfactants, with droplets being successfully generated using both anionic surfactants and more biologically relevant phospholipids. This method also allows the device to be selectively patterned with both hydrophilic and hydrophobic regions, leading to the generation of double emulsions and inverted double emulsions.
Armstrong A, Ces O, Compte RV, 2017, All aboard for chemistry, CHEMISTRY & INDUSTRY, Vol: 81, Pages: 14-15, ISSN: 0009-3068
Miller RM, Ces O, Brooks NJ, et al., 2017, Crystallization of Sodium Dodecyl Sulfate-Water Micellar Solutions under Linear Cooling, CRYSTAL GROWTH & DESIGN, Vol: 17, Pages: 2428-2437, ISSN: 1528-7483
The crystallization kinetics of sodium dodecyl sulfate (SDS)-water micellar solutions, under linear cooling conditions, were experimentally investigated using optical microscopy, differential scanning calorimetry, and infrared spectroscopy. Cooling rates were systematically varied, from 0.1 to 50 °C min–1, encompassing environmental to near-“isothermal” temperature changes, between 22 and −5 °C, for a reference concentration of 20% SDS-H2O. The cooling rate was shown to determine the dominant crystal morphologies, with platelets and needles predominating at the lowest and highest rates, respectively. The results were rationalized in terms of isothermal crystallization data and the time–temperature cooling profile. Rates 0.1, 5.0, and 10 °C min–1 yield morphologies and kinetics analogous to those of isothermal quenches at the corresponding crystallization temperature window. Nontrivial deviations were observed for intermediate rates (0.5, 1.0 °C min–1), due to commensurate changes in temperature and crystallization mechanism, accompanied by solute depletion. The polythermal metastable zone width was estimated, and the non-isothermal nucleation described by the Nývlt equation, while the Avrami and Kissinger models described overall crystallization kinetics. Our measurements quantify the impact of temperature gradients in the crystallization of ubiquitous SDS micellar solutions, for a range of practically relevant profiles incurred during manufacturing and storage.
Barlow NE, Smpokou E, Friddin MS, et al., 2017, Engineering plant membranes using droplet interface bilayers, Biomicrofluidics, Vol: 11, ISSN: 1932-1058
Droplet interface bilayers (DIBs) have become widely recognised as a robust platform for constructing model membranes and are emerging as a key technology for the bottom-up assembly of synthetic cell-like and tissue-like structures. DIBs are formed when lipid-monolayer coated water droplets are brought together inside a well of oil, which is excluded from the interface as the DIB forms. The unique features of the system, compared to traditional approaches (e.g., supported lipid bilayers, black lipid membranes, and liposomes), is the ability to engineer multi-layered bilayer networks by connecting multiple droplets together in 3D, and the capability to impart bilayer asymmetry freely within these droplet architectures by supplying droplets with different lipids. Yet despite these achievements, one potential limitation of the technology is that DIBs formed from biologically relevant components have not been well studied. This could limit the reach of the platform to biological systems where bilayer composition and asymmetry are understood to play a key role. Herein, we address this issue by reporting the assembly of asymmetric DIBs designed to replicate the plasma membrane compositions of three different plant species; Arabidopsis thaliana, tobacco, and oats, by engineering vesicles with different amounts of plant phospholipids, sterols and cerebrosides for the first time. We show that vesicles made from our plant lipid formulations are stable and can be used to assemble asymmetric plant DIBs. We verify this using a bilayer permeation assay, from which we extract values for absolute effective bilayer permeation and bilayer stability. Our results confirm that stable DIBs can be assembled from our plant membrane mimics and could lead to new approaches for assembling model systems to study membrane translocation and to screen new agrochemicals in plants.
Barlow NE, Bolognesi G, Flemming AJ, et al., 2016, Multiplexed droplet Interface bilayer formation, Lab on a Chip, Vol: 16, Pages: 4653-4657, ISSN: 1473-0197
We present a simple method for the multiplexed formation ofdroplet interface bilayers (DIBs) using a mechanically operatedlinear acrylic chamber array. To demonstrate the functionality ofthe chip design, a lipid membrane permeability assay is performed.We show that multiple, symmetric DIBs can be created andseparated using this robust low-cost approach.
Boyd C, Parsons ES, Smith RAG, et al., 2016, Disentangling the roles of cholesterol and CD59 in intermedilysin pore formation, Scientific Reports, Vol: 6, ISSN: 2045-2322
The plasma membrane provides an essential barrier, shielding a cell from the pressures of its external environment. Pore-forming proteins, deployed by both hosts and pathogens alike, breach this barrier to lyse target cells. Intermedilysin is a cholesterol-dependent cytolysin that requires the human immune receptor CD59, in addition to cholesterol, to form giant β-barrel pores in host membranes. Here we integrate biochemical assays with electron microscopy and atomic force microscopy to distinguish the roles of these two receptors in mediating structural transitions of pore formation. CD59 is required for the specific coordination of intermedilysin (ILY) monomers and for triggering collapse of an oligomeric prepore. Movement of Domain 2 with respect to Domain 3 of ILY is essential for forming a late prepore intermediate that releases CD59, while the role of cholesterol may be limited to insertion of the transmembrane segments. Together these data define a structural timeline for ILY pore formation and suggest a mechanism that is relevant to understanding other pore-forming toxins that also require CD59.
Miller DM, Findlay HE, Ces O, et al., 2016, Light-activated control of protein channel assembly mediated by membrane mechanics, Nanotechnology, Vol: 27, Pages: 1-10, ISSN: 1361-6528
Photochemical processes provide versatile triggers of chemical reactions. Here, we use a photoactivated lipid switch to modulate the folding and assembly of a protein channel within a model biological membrane. In contrast to the information rich field of water-soluble protein folding, there is only a limited understanding of the assembly of proteins that are integral to biological membranes. It is however possible to exploit the foreboding hydrophobic lipid environment and control membrane protein folding via lipid bilayer mechanics. Mechanical properties such as lipid chain lateral pressure influence the insertion and folding of proteins in membranes, with different stages of folding having contrasting sensitivities to the bilayer properties. Studies to date have relied on altering bilayer properties through lipid compositional changes made at equilibrium, and thus can only be made before or after folding. We show that light-activation of photoisomerisable di-(5-[[4-(4-butylphenyl)azo]phenoxy]pentyl)phosphate (4-Azo-5P) lipids influences the folding and assembly of the pentameric bacterial mechanosensitive channel MscL. The use of a photochemical reaction enables the bilayer properties to be altered during folding, which is unprecedented. This mechanical manipulation during folding, allows for optimisation of different stages of the component insertion, folding and assembly steps within the same lipid system. The photochemical approach offers the potential to control channel assembly when generating synthetic devices that exploit the mechanosensitive protein as a nanovalve.
Buck M, Mcdonald C, Jovanovic G, et al., 2016, Structure and function of PspA and Vipp1 N-terminal peptides:Insights into the membrane stress sensing and mitigation, BBA Biomembranes, Vol: 1859, Pages: 28-39, ISSN: 0005-2736
The phage shock protein (Psp) response maintains integrity ofthe innermembrane (IM) in responsetoextracytoplasmic stress conditionsand is widely distributed amongstenterobacteria. Its central componentPspA, a member of the IM30 peripheral membrane protein family, acts asa major effector of the systemthrough its direct association with the IM. Under non-stress conditions PspA also negatively regulates its own expression via direct interaction with the AAA+ ATPase PspF. PspA hasa counterpart in cyanobacteria calledVipp1, which is implicated in protection of the thylakoid membranes. PspA’s and Vipp1’s conservedN-terminal regions contain a putative amphipathic helix a (AHa) required for membranebinding.Anadjacent amphipathic helix b (AHb) in PspAis required for imposing negative control uponPspF.Here, purified peptides derived from the putative AH regions of PspA and Vipp1were used to directly probe their effectorand regulatory functions.We observed direct membrane-binding of AHaderived peptides and an accompanying change in secondary structure from unstructuredto alpha-helical establishing them as bonafidemembrane-sensing AH’s. The peptide-binding specificitiesand theireffects on membrane stability depend onmembrane anionic lipid content and stored curvature elastic stress,in agreement withfull length PspA and Vipp1 proteinfunctionalities. AHbof PspA inhibited the ATPase activity of PspF demonstratingits direct regulatory role. These findings provide new insight into the membrane binding and function of PspA and Vipp1and establish that synthetic peptides can be used to probe the structure-function of the IM30 protein family.
Chan CL, Bolognesi G, Bhandarkar A, et al., 2016, DROPLAY: laser writing of functional patterns within biological microdroplet displays, Lab on a Chip, Vol: 16, Pages: 4621-4627, ISSN: 1473-0197
In this study, we introduce an optofluidic method for the rapid construction of large-area cell-sized droplet assemblieswith user-defined re-writable two-dimensional patterns of functional droplets. Light responsive water-in-oil dropletscapable of releasing fluorescent dye molecules upon exposure were generated and self-assembled into arrays in amicrofluidic device. This biological architecture was exploited by the scanning laser of a confocal microscope to ‘write’ userdefined patterns of differentiated (fluorescent) droplets in a network of originally undifferentiated (non-fluorescent)droplets. As a result, long lasting images were produced on a droplet fabric with droplets acting as pixels of a biologicalmonitor, which can be erased and re-written on-demand. Regio-specific light-induced droplet differentiation within a largepopulation of droplets provides a new paradigm for the rapid construction of bio-synthetic systems with potential as tissuemimics and biological display materials.
Friddin MS, Bolognesi G, Elani Y, et al., 2016, Optically assembled droplet interface bilayer (OptiDIB) networks from cell-sized microdroplets, Soft Matter, Vol: 12, Pages: 7731-7734, ISSN: 1744-6848
We report a new platform technology to systematically assemble droplet interface bilayer (DIB) networks in user-defined 3D architectures from cell-sized droplets using optical tweezers. Our OptiDIB platform is the first demonstration of optical trapping to precisely construct 3D DIB networks, paving the way for the development of a new generation of modular bio-systems.
Friddin MS, Bolognesi G, Elani Y, et al., 2016, The optical assembly of bilayer networks from cell-sized droplets for synthetic biology, Systems and Synthetic Biology
Friddin MS, Bolognesi G, Elani Y, et al., 2016, Optical tweezers to assemble 2D and 3D droplet interface bilayer networks from cell-sized droplets, EMBL Microfluidics
Poulos AS, Nania M, Lapham P, et al., 2016, Microfluidic SAXS Study of Lamellar and Multilamellar Vesicle Phases of Linear Sodium Alkylbenzenesulfonate Surfactant with Intrinsic Isomeric Distribution, Langmuir, Vol: 32, Pages: 5852-5861, ISSN: 1520-5827
The structure and flow behavior of a concentrated aqueous solution (45 wt %) of the ubiquitous linear sodium alkylbenzenesulfonate (NaLAS) surfactant is investigated by microfluidic small-angle X-ray scattering (SAXS) at 70 °C. NaLAS is an intrinsically complex mixture of over 20 surfactant molecules, presenting coexisting micellar (L1) and lamellar (Lα) phases. Novel microfluidic devices were fabricated to ensure pressure and thermal resistance, ability to handle viscous fluids, and low SAXS background. Polarized light optical microscopy showed that the NaLAS solution exhibits wall slip in microchannels, with velocity profiles approaching plug flow. Microfluidic SAXS demonstrated the structural spatial heterogeneity of the system with a characteristic length scale of 50 nL. Using a statistical flow–SAXS analysis, we identified the micellar phase and multiple coexisting lamellar phases with a continuous distribution of d spacings between 37.5 and 39.5 Å. Additionally, we showed that the orientation of NaLAS lamellar phases is strongly affected by a single microfluidic constriction. The bilayers align parallel to the velocity field upon entering a constriction and perpendicular to it upon exiting. On the other hand, multilamellar vesicle phases are not affected under the same flow conditions. Our results demonstrate that despite the compositional complexity inherent to NaLAS, microfluidic SAXS can rigorously elucidate its structure and flow response.
Zhang N, Jovanovic G, McDonald C, et al., 2016, Transcription regulation and membrane stress management in enterobacterial pathogens, Advances in Experimental Medicine and Biology, Vol: 915, Pages: 207-230, ISSN: 0065-2598
Transcription regulation in a temporal and conditional manner underpins the lifecycle of enterobacterial pathogens. Upon exposure to a wide array of environmental cues, these pathogens modulate their gene expression via the RNA polymerase and associated sigma factors. Different sigma factors, either involved in general 'house-keeping' or specific responses, guide the RNA polymerase to their cognate promoter DNAs. The major alternative sigma54 factor when activated helps pathogens manage stresses and proliferate in their ecological niches. In this chapter, we review the function and regulation of the sigma54-dependent Phage shock protein (Psp) system-a major stress response when Gram-negative pathogens encounter damages to their inner membranes. We discuss the recent development on mechanisms of gene regulation, signal transduction and stress mitigation in light of different biophysical and biochemical approaches.
Elani Y, Solvas XC, Edel JB, et al., 2016, Microfluidic generation of encapsulated droplet interface bilayer networks (multisomes) and their use as cell-like reactors., Chemical Communications, Vol: 52, Pages: 5961-5964, ISSN: 1364-548X
Compartmentalised structures based on droplet interface bilayers (DIBs), including multisomes and compartmentalised vesicles, are seen by many as the next generation of biomimetic soft matter devices. Herein, we outline a microfluidic approach for the construction of miniaturised multisomes of pL volumes in high-throughput and demonstrate their potential as vehicles for in situ chemical synthesis.
Karamdad K, Law R, Seddon J, et al., 2016, Studying the effects of asymmetry on the bending rigidity of lipid membranes formed by microfluidics, Chemical Communications (London), Vol: 52, Pages: 5277-5280, ISSN: 0009-241X
In this article we detail a robust high-throughput microfluidic platform capable of fabricating either symmetric or asymmetric giant unilamellar vesicles (GUVs) and characterise the mechanical properties of their membranes.
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