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Correction for 'Sizing multimodal suspensions with differential dynamic microscopy' by Joe J. Bradley et al., Soft Matter, 2023, 19, 8179-8192, https://doi.org/10.1039/D3SM00593C.
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We characterize the full spatiotemporal gait of populations of swimming Escherichia coli using renewal processes to analyze the measurements of intermediate scattering functions. This allows us to demonstrate quantitatively how the persistence length of an engineered strain can be controlled by a chemical inducer and to report a controlled transition from perpetual tumbling to smooth swimming. For wild-type E. coli, we measure simultaneously the microscopic motility parameters and the large-scale effective diffusivity, hence quantitatively bridging for the first time small-scale directed swimming and macroscopic diffusion.
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Quimiotaxis , Escherichia coli , Natación , Difusión , MarchaRESUMEN
We introduce a numerical method to extract the parameters of run-and-tumble dynamics from experimental measurements of the intermediate scattering function. We show that proceeding in Laplace space is unpractical and employ instead renewal processes to work directly in real time. We first validate our approach against data produced using agent-based simulations. This allows us to identify the length and time scales required for an accurate measurement of the motility parameters, including tumbling frequency and swim speed. We compare different models for the run-and-tumble dynamics by accounting for speed variability at the single-cell and population level, respectively. Finally, we apply our approach to experimental data on wild-type Escherichia coli obtained using differential dynamic microscopy.
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Bacterias , Microscopía , Microscopía/métodos , Natación , Escherichia coli , Modelos BiológicosRESUMEN
Differential dynamic microscopy (DDM) can be used to extract the mean particle size from videos of suspensions. However, many suspensions have multimodal particle size distributions, for which a single 'mean' is not a sufficient description. After clarifying how different particle sizes contribute to the signal in DDM, we show that standard DDM analysis can extract the mean sizes of two populations in a bimodal suspension given prior knowledge of the sample's bimodality. Further, the use of the CONTIN algorithm obviates the need for such prior knowledge. Finally, we show that by selectively analysing portions of the DDM images, we can size a trimodal suspension where the large particles would otherwise dominate the signal, again without prior knowledge of the trimodality.
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Pseudomonas aeruginosa forms suspended multicellular aggregates when cultured in liquid media. These aggregates may be important in disease, and/or as a pathway to biofilm formation. The polysaccharide Psl and extracellular DNA (eDNA) have both been implicated in aggregation, but previous results depend strongly on the experimental conditions. Here we develop a quantitative microscopy-based method for assessing changes in the size distribution of suspended aggregates over time in growing cultures. For exponentially growing cultures of P. aeruginosa PAO1, we find that aggregation is mediated by cell-associated Psl, rather than by either eDNA or secreted Psl. These aggregates arise de novo within the culture via a growth process that involves both collisions and clonal growth, and Psl non-producing cells do not aggregate with producers. In contrast, we find that stationary phase (overnight) cultures contain a different type of multicellular aggregate, in which both eDNA and Psl mediate cohesion. Our findings suggest that the physical and biological properties of multicellular aggregates may be very different in early-stage vs late-stage bacterial cultures.
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Biopelículas , Pseudomonas aeruginosa , Polisacáridos Bacterianos/metabolismo , ADNRESUMEN
Drug nanocarriers (NCs) capable of crossing the vascular endothelium and deeply penetrating into dense tissues of the CNS could potentially transform the management of neurological diseases. In the present study, we investigated the interaction of bottle-brush (BB) polymers with different biological barriers in vitro and in vivo and compared it to nanospheres of similar composition. In vitro internalization and permeability assays revealed that BB polymers are not internalized by brain-associated cell lines and translocate much faster across a blood-brain barrier model compared to nanospheres of similar hydrodynamic diameter. These observations performed under static, no-flow conditions were complemented by dynamic assays performed in microvessel arrays on chip and confirmed that BB polymers can escape the vasculature compartment via a paracellular route. BB polymers injected in mice and zebrafish larvae exhibit higher penetration in brain tissues and faster extravasation of microvessels located in the brain compared to nanospheres of similar sizes. The superior diffusivity of BBs in extracellular matrix-like gels combined with their ability to efficiently cross endothelial barriers via a paracellular route position them as promising drug carriers to translocate across the blood-brain barrier and penetrate dense tissue such as the brain, two unmet challenges and ultimate frontiers in nanomedicine.
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Polímeros , Pez Cebra , Ratones , Animales , Polímeros/metabolismo , Pez Cebra/metabolismo , Barrera Hematoencefálica/metabolismo , Encéfalo/metabolismo , Transporte BiológicoRESUMEN
We study a synthetic system of motile Escherichia coli bacteria encapsulated inside giant lipid vesicles. Forces exerted by the bacteria on the inner side of the membrane are sufficient to extrude membrane tubes filled with one or several bacteria. We show that a physical coupling between the membrane tube and the flagella of the enclosed cells transforms the tube into an effective helical flagellum propelling the vesicle. We develop a simple theoretical model to estimate the propulsive force from the speed of the vesicles and demonstrate the good efficiency of this coupling mechanism. Together, these results point to design principles for conferring motility to synthetic cells.
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Células Artificiales/microbiología , Escherichia coli/fisiología , Vesículas Citoplasmáticas/microbiología , Escherichia coli/citología , Flagelos/fisiología , Lípidos , Membranas ArtificialesRESUMEN
A new theoretical framework that enables the use of differential dynamic microscopy (DDM) in fluorescence imaging mode to quantify in situ protein adsorption onto nanoparticles (NP) while simultaneously monitoring for NP aggregation is proposed. This methodology is used to elucidate the thermodynamic and kinetic properties of the protein corona (PC) in vitro and in vivo. The results show that protein adsorption triggers particle aggregation over a wide concentration range and that the formed aggregate structures can be quantified using the proposed methodology. Protein affinity for polystyrene (PS) NPs is observed to be dependent on particle concentration. For complex protein mixtures, this methodology identifies that the PC composition changes with the dilution of serum proteins, demonstrating a Vroman effect never quantitatively assessed in situ on NPs. Finally, DDM allows monitoring of the evolution of the PC in vivo. This results show that the PC composition evolves significantly over time in zebrafish larvae, confirming the inherently dynamic nature of the PC. The performance of the developed methodology allows to obtain quantitative insights into nano-bio interactions in a vast array of physiologically relevant conditions that will serve to further improve the design of nanomedicine.
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Nanopartículas , Corona de Proteínas , Animales , Proteínas Sanguíneas , Nanopartículas/química , Poliestirenos/química , Corona de Proteínas/química , Pez CebraRESUMEN
Motile bacteria are known to accumulate at surfaces, eventually leading to changes in bacterial motility and biofilm formation. We use a novel two-color, three-dimensional Lagrangian tracking technique to follow simultaneously the body and the flagella of a wild-type Escherichia coli. We observe long surface residence times and surface escape corresponding mostly to immediately antecedent tumbling. A motility model accounting for a large behavioral variability in run-time duration reproduces all experimental findings and gives new insights into surface trapping efficiency.
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Escherichia coli , Flagelos , BacteriasRESUMEN
Few techniques can reliably measure the dynamics of colloidal suspensions or other soft materials over a wide range of turbidities. Here we systematically investigate the capability of Differential Dynamic Microscopy (DDM) to characterise particle dynamics in turbid colloidal suspensions based on brightfield optical microscopy. We measure the Intermediate Scattering Function (ISF) of polystyrene microspheres suspended in water over a range of concentrations, turbidities, and up to 4 orders of magnitude in time-scales. These DDM results are compared to data obtained from both Dynamic Light Scattering (DLS) and Two-colour Dynamic Light Scattering (TCDLS). The latter allows for suppression of multiple scattering for moderately turbid suspensions. We find that DDM can obtain reliable diffusion coefficients at up to 10 and 1000 times higher particle concentrations than TCDLS and standard DLS, respectively. Additionally, we investigate the roles of the four length-scales relevant when imaging a suspension: the sample thickness L, the imaging depth z, the imaging depth of field DoF, and the photon mean free path î . More detailed experiments and analysis reveal the appearance of a short-time process as turbidity is increased, which we associate with multiple scattering events within the imaging depth of the field. The long-time process corresponds to the particle dynamics from which particle-size can be estimated in the case of non-interacting particles. Finally, we provide a simple theoretical framework, ms-DDM, for turbid samples, which accounts for multiple scattering.
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Microscopía , Fotones , Dispersión Dinámica de Luz , Microscopía/métodos , Tamaño de la Partícula , SuspensionesRESUMEN
This Review aims to provide a systematic analysis of the literature regarding ongoing debates in protein corona research. Our goal is to portray the current understanding of two fundamental and debated characteristics of the protein corona, namely, the formation of mono- or multilayers of proteins and their binding (ir)reversibility. The statistical analysis we perform reveals that these characterisitics are strongly correlated to some physicochemical factors of the NP-protein system (particle size, bulk material, protein type), whereas the technique of investigation or the type of measurement (in situ or ex situ) do not impact the results, unlike commonly assumed. Regarding the binding reversibility, the experimental design (either dilution or competition experiments) is also shown to be a key factor, probably due to nontrivial protein binding mechanisms, which could explain the paradoxical phenomena reported in the literature. Overall, we suggest that to truly predict and control the protein corona, future efforts should be directed toward the mechanistic aspects of protein adsorption.
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Nanopartículas , Corona de Proteínas , Adsorción , Nanopartículas/metabolismo , Tamaño de la Partícula , Unión Proteica , Corona de Proteínas/metabolismoRESUMEN
Microscopic dynamics reveal the origin of the bulk rheological response in complex fluids. In model systems particle motion can be tracked, but for industrially relevant samples this is often impossible. Here we adapt differential dynamic microscopy (DDM) to study flowing highly-concentrated samples without particle resolution. By combining an investigation of oscillatory flow, using a novel "echo-DDM" analysis, and steady shear, through flow-DDM, we characterise the yielding of a silicone oil emulsion on both the microscopic and bulk level. Through measuring the rate of shear-induced droplet rearrangements and the flow velocity, the transition from a solid-like to liquid-like state is shown to occur in two steps: with droplet mobilisation marking the limit of linear visco-elasticity, followed by the development of shear localisation and macroscopic yielding. Using this suite of techniques, such insight could be developed for a wide variety of challenging complex fluids.
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Particle size is a key variable in understanding the behaviour of the particulate products that underpin much of our modern lives. Typically obtained from suspensions at rest, measuring the particle size under flowing conditions would enable advances for in-line testing during manufacture and high-throughput testing during development. However, samples are often turbid, multiply scattering light and preventing the direct use of common sizing techniques. Differential dynamic microscopy (DDM) is a powerful technique for analysing video microscopy of such samples, measuring diffusion and hence particle size without the need to resolve individual particles while free of substantial user input. However, when applying DDM to a flowing sample, diffusive dynamics are rapidly dominated by flow effects, preventing particle sizing. Here, we develop "flow-DDM", a novel analysis scheme that combines optimised imaging conditions, a drift-velocity correction and modelling of the impact of flow. Flow-DDM allows a decoupling of flow from diffusive motion that facilitates successful particle size measurements at flow speeds an order of magnitude higher than for DDM. We demonstrate the generality of the technique by applying flow-DDM to two separate microscopy methods and flow geometries.
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Anisotropic dynamics on the colloidal length scale is ubiquitous in nature. Of particular interest is the dynamics of systems approaching a kinetically arrested state. The failure of classical techniques for investigating the dynamics of highly turbid suspensions has contributed toward the limited experimental information available up until now. Exploiting the recent developments in the technique of differential dynamic microscopy (DDM), we report the first experimental study of the anisotropic collective dynamics of colloidal ellipsoids with a magnetic hematite core over a wide concentration range approaching kinetic arrest. In addition, we have investigated the effect of an external magnetic field on the resulting anisotropic collective diffusion. We combine DDM with small-angle x-ray scattering and rheological measurements to locate the glass transition and to relate the collective short- and long-time diffusion coefficients to the structural correlations and the evolution of the zero shear viscosity as the system approaches an arrested state.
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Suspending self-propelled "pushers" in a liquid lowers its viscosity. We study how this phenomenon depends on system size in bacterial suspensions using bulk rheometry and particle-tracking rheoimaging. Above the critical bacterial volume fraction needed to decrease the viscosity to zero, [Formula: see text], large-scale collective motion emerges in the quiescent state, and the flow becomes nonlinear. We confirm a theoretical prediction that such instability should be suppressed by confinement. Our results also show that a recent application of active liquid-crystal theory to such systems is untenable.
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Fenómenos Fisiológicos Bacterianos , Suspensiones/química , Bacterias/citología , Rastreo Celular , Escherichia coli/citología , Escherichia coli/fisiología , Locomoción , Reología , Resistencia al Corte , ViscosidadRESUMEN
We present chitosan hydrogel microfluidic devices with self-assembled complex microcapillary patterns, conveniently formed by a diffusion-reaction process. These patterns in chitosan hydrogels are formed by a single-step procedure involving diffusion of a gelation agent into the polymer solution inside a microfluidic channel. By changing the channel geometry, it is demonstrated how to control capillary length, trajectory and branching. Diffusion of nanoparticles (NPs) in the capillary network is used as a model to effectively mimic the transport of nano-objects in vascularized tissues. Gold NPs diffusion is measured locally in the hydrogel chips, and during their two-step transport through the capillaries to the gel matrix and eventually to embedded cell clusters in the gel. In addition, the quantitative analyses reported in this study provide novel opportunities for theoretical investigation of capillary formation and propagation during diffusive gelation of biopolymers. STATEMENT OF SIGNIFICANCE: Hydrogel micropatterning is a challenging task, which is of interest in several biomedical applications. Creating the patterns through self assembly is highly beneficial, because of the accessible and practical preparation procedure. In this study, we introduced complex self-assembled capillary patterns in chitosan hydrogels using a microfluidic approach. To demonstrate the potential application of these capillary patterns, a vascularized hydrogel with microwells occupied by cells was produced, and the diffusion of gold nanoparticles travelling in the capillaries and diffusing in the gel were evaluated. This model mimics a simplified biological tissue, where nanomedicine has to travel through the vasculature, extravasate into and diffuse through the extracellular matrix and eventually reach targeted cells.
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Quitosano/química , Hidrogeles/química , Microcirculación/efectos de los fármacos , Nanopartículas/química , Animales , Biopolímeros/química , Capilares , Bovinos , Difusión , Dimetilpolisiloxanos/química , Sistemas de Liberación de Medicamentos , Fibroblastos/citología , Oro/química , Ensayo de Materiales , Nanopartículas del Metal/química , Microfluídica , Microscopía Confocal , Hidróxido de Sodio/química , Ingeniería de Tejidos/métodos , Andamios del TejidoRESUMEN
Improving nanoparticles (NPs) transport across biological barriers is a significant challenge that could be addressed through understanding NPs diffusion in dense and confined media. Here, we report the ability of soft NPs to shrink in confined environments, therefore boosting their diffusion compared to hard, non-deformable particles. We demonstrate this behavior by embedding microgel NPs in agarose gels. The origin of the shrinking appears to be related to the overlap of the electrostatic double layers (EDL) surrounding the NPs and the agarose fibres. Indeed, it is shown that screening the EDL interactions, by increasing the ionic strength of the medium, prevents the soft particle shrinkage. The shrunken NPs diffuse up to 2 orders of magnitude faster in agarose gel than their hard NP counterparts. These findings provide valuable insights on the role of long range interactions on soft NPs dynamics in crowded environments, and help rationalize the design of more efficient NP-based transport systems.
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We use moving light patterns to control the motion of Escherichia coli bacteria whose motility is photo-activated. Varying the pattern speed controls the magnitude and direction of the bacterial flux, and therefore the accumulation of cells in up- and down-stream reservoirs. We validate our results with two-dimensional simulations and a 1-dimensional analytic model, and use these to explore parameter space. We find that cell accumulation is controlled by a competition between directed flux and undirected, stochastic transport. Our results point to a number of design principles for using moving light patterns and light-activated micro-swimmers in a range of practical applications.
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Self-propelled colloids constitute an important class of intrinsically non-equilibrium matter. Typically, such a particle moves ballistically at short times, but eventually changes its orientation, and displays random-walk behaviour in the long-time limit. Theory predicts that if the velocity of non-interacting swimmers varies spatially in 1D, v(x), then their density ρ(x) satisfies ρ(x) = ρ(0)v(0)/v(x), where x = 0 is an arbitrary reference point. Such a dependence of steady-state ρ(x) on the particle dynamics, which was the qualitative basis of recent work demonstrating how to 'paint' with bacteria, is forbidden in thermal equilibrium. Here we verify this prediction quantitatively by constructing bacteria that swim with an intensity-dependent speed when illuminated and implementing spatially-resolved differential dynamic microscopy (sDDM) for quantitative analysis over millimeter length scales. Applying a spatial light pattern therefore creates a speed profile, along which we find that, indeed, ρ(x)v(x) = constant, provided that steady state is reached.
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We report a high-throughput technique for characterising the motility of spermatozoa using differential dynamic microscopy. A movie with large field of view (â¼10mm2) records thousands of cells (e.g. ≈ 5000 cells even at a low cell density of 20 × 106 cells/ml) at once and yields averaged measurements of the mean ([Formula: see text]) and standard deviation (σ) of the swimming speed, head oscillation amplitude (A0) and frequency (f0), and the fraction of motile spermatozoa (α). Interestingly, we found that the measurement of α is facilitated because the swimming spermatozoa enhance the motion of the non-swimming population. We demonstrate the ease and rapidity of our method by performing on-farm characterisation of bull spermatozoa motility, and validate the technique by comparing laboratory measurements with tracking. Our results confirm the long-standing theoretical prediction that [Formula: see text] for swimming spermatozoa.