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This article introduces an analysis-aware microscopy video compression method designed for microscopy videos that are consumed by analysis algorithms rather than by the human visual system. We define the quality of a microscopy video based on the level of preservation of analysis results. We evaluated our method with a bead tracking analysis program. For the same error level in the analysis result, our method can achieve 1,000× compression on certain test microscopy videos. Compared with a previous technique that yields exactly the exact same results by analysis algorithms, our method gives more flexibility for a user to control the quality. A modification to the new method also provides faster compression speed., (© 2018 Wiley Periodicals, Inc.)

The Virtual Pediatric Airways Workbench (VPAW) is a patient-centered surgical planning software system targeted to pediatric patients with airway obstruction. VPAW provides an intuitive surgical planning interface for clinicians and supports quantitative analysis regarding prospective surgeries to aid clinicians deciding on potential surgical intervention. VPAW enables a full surgical planning pipeline, including importing DICOM images, segmenting the airway, interactive 3D editing of airway geometries to express potential surgical treatment planning options, and creating input files for offline geometric analysis and computational fluid dynamics simulations for evaluation of surgical outcomes. In this paper, we describe the VPAW system and its use in one case study with a clinician to successfully describe an intended surgery outcome.

ChromoShake is a three-dimensional simulator designed to find the thermodynamically favored states for given chromosome geometries. The simulator has been applied to a geometric model based on experimentally determined positions and fluctuations of DNA and the distribution of cohesin and condensin in the budding yeast centromere. Simulations of chromatin in differing initial configurations reveal novel principles for understanding the structure and function of a eukaryotic centromere. The entropic position of DNA loops mirrors their experimental position, consistent with their radial displacement from the spindle axis. The barrel-like distribution of cohesin complexes surrounding the central spindle in metaphase is a consequence of the size of the DNA loops within the pericentromere to which cohesin is bound. Linkage between DNA loops of different centromeres is requisite to recapitulate experimentally determined correlations in DNA motion. The consequences of radial loops and cohesin and condensin binding are to stiffen the DNA along the spindle axis, imparting an active function to the centromere in mitosis., (© 2016 Lawrimore et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).)

The large amount video data produced by multi-channel, high-resolution microscopy system drives the need for a new high-performance domain-specific video compression technique. We describe a novel compression method for video microscopy data. The method is based on Pearson's correlation and mathematical morphology. The method makes use of the point-spread function (PSF) in the microscopy video acquisition phase. We compare our method to other lossless compression methods and to lossy JPEG, JPEG2000, and H.264 compression for various kinds of video microscopy data including fluorescence video and brightfield video. We find that for certain data sets, the new method compresses much better than lossless compression with no impact on analysis results. It achieved a best compressed size of 0.77% of the original size, 25× smaller than the best lossless technique (which yields 20% for the same video). The compressed size scales with the video's scientific data content. Further testing showed that existing lossy algorithms greatly impacted data analysis at similar compression sizes., (© 2015 Wiley Periodicals, Inc.)

The progression of breast cancer is known to be affected by stromal cells within the local microenvironment. Here we study the effect of stromal fibroblasts on the in-place motions (motility) of mammary epithelial cells within organoids in 3D co-culture, inferred from the speckle fluctuation spectrum using optical coherence tomography (OCT). In contrast to Brownian motion, mammary cell motions exhibit an inverse power-law fluctuation spectrum. We introduce two complementary metrics for quantifying fluctuation spectra: the power-law exponent and a novel definition of the motility amplitude, both of which are signal- and position-independent. We find that the power-law exponent and motility amplitude are positively ( p <0.001) and negatively ( p <0.01) correlated with the density of stromal cells in 3D co-culture, respectively. We also show how the hyperspectral data can be visualized using these metrics to observe heterogeneity within organoids. This constitutes a simple and powerful tool for detecting and imaging cellular functional changes with OCT.

The centromere is the DNA locus that dictates kinetochore formation and is visibly apparent as heterochromatin that bridges sister kinetochores in metaphase. Sister centromeres are compacted and held together by cohesin, condensin, and topoisomerase-mediated entanglements until all sister chromosomes bi-orient along the spindle apparatus. The establishment of tension between sister chromatids is essential for quenching a checkpoint kinase signal generated from kinetochores lacking microtubule attachment or tension. How the centromere chromatin spring is organized and functions as a tensiometer is largely unexplored. We have discovered that centromere chromatin loops generate an extensional/poleward force sufficient to release nucleosomes proximal to the spindle axis. This study describes how the physical consequences of DNA looping directly underlie the biological mechanism for sister centromere separation and the spring-like properties of the centromere in mitosis., (© 2015 Lawrimore et al.)

We have developed new procedures to examine the early steps in fibrin polymerization. First, we isolated fibrinogen monomers from plasma fibrinogen by gel filtration. Polymerization of fibrinogen monomers differed from that of plasma fibrinogen. The formation of protofibrils was slower and the transformation of protofibrils to fibers faster for the fibrinogen monomers. Second, we used formaldehyde to terminate the polymerization reactions. The formaldehyde-fixed products obtained at each time point were examined by dynamic light scattering and transmission electron microscopy (TEM). The data showed the formaldehyde-fixed products were stable and representative of the reaction intermediates. TEM images showed monomers, short oligomers, protofibrils, and thin fibers. The amount and length of these species varied with time. Short oligomers were less than 5% of the molecules at all times. Third, we developed models that recapitulate the TEM images. Fibrin monomer models were assembled into protofibrils, and protofibrils were assembled into two-strand fibers using Chimera software. Monomers were based on fibrinogen crystal structures, and the end-to-end interactions between monomers were based on D-dimer crystal structures. Protofibrils assembled from S-shaped monomers through asymmetric D:D interactions were ordered helical structures. Fibers were modeled by duplicating a protofibril and rotating the duplicate 120° around its long axis. No specific interactions were presumed. The two protofibrils simply twisted around one another to form a fiber. This model suggests that the conformation of the protofibril per se promotes the assembly into fibers. These findings introduce a novel mechanism for fibrin assembly that may be relevant to other biopolymers.

Using random per-element luminance modulation can increase the visual salience of details in a range of visualizations (2D, 3D, and ND scalar, vector, and tensor fields). Although random luminance has been used in specific designs, its wide applicability isn't reflected in visualizations, perhaps because it hasn't yet been presented as a cross-cutting technique. Adding random-luminance contrast can benefit both static and animated visualizations. The article presents perceptual reasons for this technique's effectiveness. This article has two accompanying videos, at http://youtu.be/TTaSFMvBgvg and http://youtu.be/Rx1oPMTpPA4, showing animations of cones moving through a weather simulation, with and without random luminance modulation.

Background: Because of the difficulties involved in learning and using 3D modeling and rendering software, many scientists hire programmers or animators to create models and animations. This both slows the discovery process and provides opportunities for miscommunication. Working with multiple collaborators, a tool was developed (based on a set of design goals) to enable them to directly construct models and animations., Results: SketchBio is presented, a tool that incorporates state-of-the-art bimanual interaction and drop shadows to enable rapid construction of molecular structures and animations. It includes three novel features: crystal-by-example, pose-mode physics, and spring-based layout that accelerate operations common in the formation of molecular models. Design decisions and their consequences are presented, including cases where iterative design was required to produce effective approaches., Conclusions: The design decisions, novel features, and inclusion of state-of-the-art techniques enabled SketchBio to meet all of its design goals. These features and decisions can be incorporated into existing and new tools to improve their effectiveness.

In mitosis, the pericentromere is organized into a spring composed of cohesin, condensin, and a rosette of intramolecular chromatin loops. Cohesin and condensin are enriched in the pericentromere, with spatially distinct patterns of localization. Using model convolution of computer simulations, we deduce the mechanistic consequences of their spatial segregation. Condensin lies proximal to the spindle axis, whereas cohesin is radially displaced from condensin and the interpolar microtubules. The histone deacetylase Sir2 is responsible for the axial position of condensin, while the radial displacement of chromatin loops dictates the position of cohesin. The heterogeneity in distribution of condensin is most accurately modeled by clusters along the spindle axis. In contrast, cohesin is evenly distributed (barrel of 500-nm width × 550-nm length). Models of cohesin gradients that decay from the centromere or sister cohesin axis, as previously suggested, do not match experimental images. The fine structures of cohesin and condensin deduced with subpixel localization accuracy reveal critical features of how these complexes mold pericentric chromatin into a functional spring.

The budding yeast kinetochore is ~68 nm in length with a diameter slightly larger than a 25 nm microtubule. The kinetochores from the 16 chromosomes are organized in a stereotypic cluster encircling central spindle microtubules. Quantitative analysis of the inner kinetochore cluster (Cse4, COMA) reveals structural features not apparent in singly attached kinetochores. The cluster of Cse4-containing kinetochores is physically larger perpendicular to the spindle axis relative to the cluster of Ndc80 molecules. If there was a single Cse4 (molecule or nucleosome) at the kinetochore attached to each microtubule plus end, the cluster of Cse4 would appear geometrically identical to Ndc80. Thus, the structure of the inner kinetochore at the surface of the chromosomes remains unsolved. We have used point fluorescence microscopy and statistical probability maps to deduce the two-dimensional mean position of representative components of the yeast kinetochore relative to the mitotic spindle in metaphase. Comparison of the experimental images to three-dimensional architectures from convolution of mathematical models reveals a pool of Cse4 radially displaced from Cse4 at the kinetochore and kinetochore microtubule plus ends. The pool of displaced Cse4 can be experimentally depleted in mRNA processing pat1Δ or xrn1Δ mutants. The peripheral Cse4 molecules do not template outer kinetochore components. This study suggests an inner kinetochore plate at the centromere-microtubule interface in budding yeast and yields information on the number of Ndc80 molecules at the microtubule attachment site., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

The mechanisms by which sister chromatids maintain biorientation on the metaphase spindle are critical to the fidelity of chromosome segregation. Active force interplay exists between predominantly extensional microtubule-based spindle forces and restoring forces from chromatin. These forces regulate tension at the kinetochore that silences the spindle assembly checkpoint to ensure faithful chromosome segregation. Depletion of pericentric cohesin or condensin has been shown to increase the mean and variance of spindle length, which have been attributed to a softening of the linear chromatin spring. Models of the spindle apparatus with linear chromatin springs that match spindle dynamics fail to predict the behavior of pericentromeric chromatin in wild-type and mutant spindles. We demonstrate that a nonlinear spring with a threshold extension to switch between spring states predicts asymmetric chromatin stretching observed in vivo. The addition of cross-links between adjacent springs recapitulates coordination between pericentromeres of neighboring chromosomes.

An ensemble is a collection of related datasets. Each dataset, or member, of an ensemble is normally large, multidimensional, and spatio-temporal. Ensembles are used extensively by scientists and mathematicians, for example, by executing a simulation repeatedly with slightly different input parameters and saving the results in an ensemble to see how parameter choices affect the simulation. To draw inferences from an ensemble, scientists need to compare data both within and between ensemble members. We propose two techniques to support ensemble exploration and comparison: a pairwise sequential animation method that visualizes locally neighboring members simultaneously, and a screen door tinting method that visualizes subsets of members using screen space subdivision. We demonstrate the capabilities of both techniques, first using synthetic data, then with simulation data of heavy ion collisions in high-energy physics. Results show that both techniques are capable of supporting meaningful comparisons of ensemble data.

By definition, an ensemble is a set of surfaces or volumes derived from a series of simulations or experiments. Sometimes the series is run with different initial conditions for one parameter to determine parameter sensitivity. The understanding and identification of visual similarities and differences among the shapes of members of an ensemble is an acute and growing challenge for researchers across the physical sciences. More specifically, the task of gaining spatial understanding and identifying similarities and differences between multiple complex geometric data sets simultaneously has proved challenging. This paper proposes a comparison and visualization technique to support the visual study of parameter sensitivity. We present a novel single-image view and sampling technique which we call Ensemble Surface Slicing (ESS). ESS produces a single image that is useful for determining differences and similarities between surfaces simultaneously from several data sets. We demonstrate the usefulness of ESS on two real-world data sets from our collaborators.

We present three extensions to parallel coordinates that increase the perceptual salience of relationships between axes in multivariate data sets: (1) luminance modulation maintains the ability to preattentively detect patterns in the presence of overplotting, (2) adding a one-vs.-all variable display highlights relationships between one variable and all others, and (3) adding a scatter plot within the parallel-coordinates display preattentively highlights clusters and spatial layouts without strongly interfering with the parallel-coordinates display. These techniques can be combined with one another and with existing extensions to parallel coordinates, and two of them generalize beyond cases with known-important axes. We applied these techniques to two real-world data sets (relativistic heavy-ion collision hydrodynamics and weather observations with statistical principal component analysis) as well as the popular car data set. We present relationships discovered in the data sets using these methods.

Correlative microscopy is a sophisticated approach that combines the capabilities of typically separate, but powerful microscopy platforms: often including, but not limited, to conventional light, confocal and super-resolution microscopy, atomic force microscopy, transmission and scanning electron microscopy, magnetic resonance imaging and micro/nano CT (computed tomography). When targeting rare or specific events within large populations or tissues, correlative microscopy is increasingly being recognized as the method of choice. Furthermore, this multi-modal assimilation of technologies provides complementary and often unique information, such as internal and external spatial, structural, biochemical and biophysical details from the same targeted sample. The development of a continuous stream of cutting-edge applications, probes, preparation methodologies, hardware and software developments will enable realization of the full potential of correlative microscopy., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

A clot's function is to achieve hemostasis by resisting fluid flow. Permeability is the measurement of a clot's hemostatic potential. It is sensitive to a wide range of biochemical parameters and pathologies. In this work, we consider the hydrodynamic phenomenon that reduces the mobility of fluid near the fiber surfaces. This no-slip boundary condition both defines the gel's permeability and suppresses nanoparticle diffusion in gel interstices. Here we report that, unlike previous work where steric effects also hindered diffusion, our system-nanoparticles in fibrin gel-was subject exclusively to hydrodynamic diffusion suppression. This result enabled an automated, high-throughput permeability assay that used small clot volumes. Permeability was derived from nanoparticle diffusion using the effective medium theory, and showed one-to-one correlation with measured permeability. This technique measured permeability without quantifying gel structure, and may therefore prove useful for characterizing similar materials (e.g., extracellular matrix) where structure is uncontrolled during polymerization and difficult to measure subsequently. We also report that PEGylation reduced, but did not eliminate, the population of immobile particles. We studied the forces required to pull stuck PEG particles free to confirm that the attachment is a result of neither covalent nor strong electrostatic binding, and discuss the relevance of this force scale to particle transport through physiological clots., (Copyright © 2011 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Sister chromatid cohesion provides the mechanistic basis, together with spindle microtubules, for generating tension between bioriented chromosomes in metaphase. Pericentric chromatin forms an intramolecular loop that protrudes bidirectionally from the sister chromatid axis. The centromere lies on the surface of the chromosome at the apex of each loop. The cohesin and condensin structural maintenance of chromosomes (SMC) protein complexes are concentrated within the pericentric chromatin, but whether they contribute to tension-generating mechanisms is not known. To understand how pericentric chromatin is packaged and resists tension, we map the position of cohesin (SMC3), condensin (SMC4), and pericentric LacO arrays within the spindle. Condensin lies proximal to the spindle axis and is responsible for axial compaction of pericentric chromatin. Cohesin is radially displaced from the spindle axis and confines pericentric chromatin. Pericentric cohesin and condensin contribute to spindle length regulation and dynamics in metaphase. Together with the intramolecular centromere loop, these SMC complexes constitute a molecular spring that balances spindle microtubule force in metaphase.

Conveying data uncertainty in visualizations is crucial for preventing viewers from drawing conclusions based on untrustworthy data points. This paper proposes a methodology for efficiently generating density plots of uncertain multivariate data sets that draws viewers to preattentively identify values of high certainty while not calling attention to uncertain values. We demonstrate how to augment scatter plots and parallel coordinates plots to incorporate statistically modeled uncertainty and show how to integrate them with existing multivariate analysis techniques, including outlier detection and interactive brushing. Computing high quality density plots can be expensive for large data sets, so we also describe a probabilistic plotting technique that summarizes the data without requiring explicit density plot computation. These techniques have been useful for identifying brain tumors in multivariate magnetic resonance spectroscopy data and we describe how to extend them to visualize ensemble data sets.

The image registration framework in the Insight Tookit offers a powerful body of code for finding the optimal spatial transform that registers one image with another. However, ITK currently lacks a way to fit parametric models of image pixel values to an input image. Such a capability is necessary in certain applications such as sub-resolution localization of molecules in fluorescence microscopy. This document describes new classes that enable the use of the registration framework to provide this capability. We describe a new base class, itk::ParametricImageSource, that defines an interface for parametric image sources. An adapter class itk::ImageToParametricImageSourceMetric that enables itk::ParametricImageSources to be hooked into the registration framework is also described. An example adapter class that enables the existing itk::GaussianImageSource to be used for image fitting is presented, and we demonstrate use of the classes by fitting a 2D Gaussian function to an image generated by the itk::GaussianImageSource class.

As the structural backbone of blood clots, fibrin networks carry out the mechanical task of stemming blood flow at sites of vascular injury. These networks exhibit a rich set of remarkable mechanical properties, but a detailed picture relating the microscopic mechanics of the individual fibers to the overall network properties has not been fully developed. In particular, how the high strain and failure characteristics of single fibers affect the overall strength of the network is not known. Using a combined fluorescence/atomic force microscope nanomanipulation system, we stretched 2-D fibrin networks to the point of failure, while recording the strain of individual fibers. Our results were compared to a pair of model networks: one composed of linearly responding elements and a second of nonlinear, strain-stiffening elements. We find that strain-stiffening of the individual fibers is necessary to explain the pattern of strain propagation throughout the network that we observe in our experiments. Fiber strain-stiffening acts to distribute strain more equitably within the network, reduce strain maxima, and increase network strength. Along with its physiological implications, a detailed understanding of this strengthening mechanism may lead to new design strategies for engineered polymeric materials., (Copyright 2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

We present a user study quantifying the effectiveness of Scaled Data-Driven Spheres (SDDS), a multivariate three-dimensional data set visualization technique. The user study compares SDDS, which uses separate sets of colored sphere glyphs to depict variable values, to superquadric glyphs, an alternative technique that maps all variable values to a single glyph. User study participants performed tasks designed to measure their ability to estimate values of particular variables and identify relationships among variables. Results from the study show that users were significantly more accurate and faster for both tasks under the SDDS condition.

Fibrin polymerizes into the fibrous network that is the major structural component of blood clots and thrombi. We demonstrate that fibrin from three different species can also spontaneously polymerize into extensive, molecularly thin, 2D sheets. Sheet assembly occurs in physiologic buffers on both hydrophobic and hydrophilic surfaces, but is routinely observed only when polymerized using very low concentrations of fibrinogen and thrombin. Sheets may have been missed in previous studies because they may be very short-lived at higher concentrations of fibrinogen and thrombin, and their thinness makes them very difficult to detect. We were able to distinguish fluorescently labeled fibrin sheets by polymerizing fibrin onto micro-patterned structured surfaces that suspended polymers 10 microm above and parallel to the cover-glass surface. We used a combined fluorescence/atomic force microscope system to determine that sheets were approximately 5 nm thick, flat, elastic and mechanically continuous. Video microscopy of assembling sheets showed that they could polymerize across 25-microm channels at hundreds of microm(2)/sec (approximately 10(13) subunits/s x M), an apparent rate constant many times greater than those of other protein polymers. Structural transitions from sheets to fibers were observed by fluorescence, transmission, and scanning electron microscopy. Sheets appeared to fold and roll up into larger fibers, and also to develop oval holes to form fiber networks that were "pre-attached" to the substrate and other fibers. We propose a model of fiber formation from sheets and compare it with current models of end-wise polymerization from protofibrils. Sheets could be an unanticipated factor in clot formation and adhesion in vivo, and are a unique material in their own right.

The ability to detect biological events at the single-molecule level provides unique biophysical insights. Back-focal-plane laser interferometry is a promising technique for nanoscale three-dimensional position measurements at rates far beyond the capability of standard video. We report an in situ calibration technique for back-focal-plane, low-power (nontrapping) laser interferometry. The technique does not rely on any a priori model or calibration knowledge, hence the name "agnostic". We apply the technique to track long-range (up to 100 microm) motion of a variety of particles, including magnetic beads, in three-dimensions with high spatiotemporal resolution ( approximately 2 nm, 100 micros). Our tracking of individual unlabeled vesicles revealed a previously unreported grouping of mean-squared displacement curves at short timescales (<10 ms). Also, tracking functionalized magnetic beads attached to a live cell membrane revealed an anchorage-dependent nonlinear response of the membrane. The software-based technique involves injecting small perturbations into the probe position by driving a precalibrated specimen-mounting stage while recording the quadrant photodetector signals. The perturbations and corresponding quadrant photodetector signals are analyzed to extract the calibration parameters. The technique is sufficiently fast and noninvasive that the calibration can be performed on-the-fly without interrupting or compromising high-bandwidth, long-range tracking of a particle.

Life is a mechanical process. Cells, tissues, and bodies must act within their environments to grow, divide, move, communicate, and defend themselves. The stiffness and viscosity of cells and biologic materials will vary depending upon a wide variety of variables including for example environmental conditions, activation of signaling pathways, stage of development, gene expression. By pushing and pulling cells or materials such as mucus or extracellular matrix, one can learn about their mechanical properties. By varying the conditions, signaling pathways or genetic background, one can also assess how the response of the cell or material is modulated by that pathway. Magnetic particles are available commercially in many useful sizes, magnetic contents, and surface chemistries. The variety of surface chemistries allow forces to be applied to a specimen through specific linkages such as receptors or particular proteins, allowing the biologist to ask fundamental questions about the role of those linkages in the transduction of force or motion. In this chapter, we discuss the use of a magnetic system designed to apply a wide range of forces and force patterns fully integrated into a high numerical aperture inverted fluorescence microscope. Fine, thin and flat magnetic poles allow the use of high magnification microscope objectives, and flexible software to control the direction and pattern of applied forces supports a variety of experimental situations. The system can be coupled with simple video acquisition for medium-bandwidth, two-dimensional particle tracking. Alternatively, the system can be coupled with a laser tracking and position feedback system for higher resolution, high bandwidth, three-dimensional tracking.

Fluorescence microscopy provides a powerful method for localization of structures in biological specimens. However, aspects of the image formation process such as noise and blur from the microscope's point-spread function combine to produce an unintuitive image transformation on the true structure of the fluorescing molecules in the specimen, hindering qualitative and quantitative analysis of even simple structures in unprocessed images. We introduce FluoroSim, an interactive fluorescence microscope simulator that can be used to train scientists who use fluorescence microscopy to understand the artifacts that arise from the image formation process, to determine the appropriateness of fluorescence microscopy as an imaging modality in an experiment, and to test and refine hypotheses of model specimens by comparing the output of the simulator to experimental data. FluoroSim renders synthetic fluorescence images from arbitrary geometric models represented as triangle meshes. We describe three rendering algorithms on graphics processing units for computing the convolution of the specimen model with a microscope's point-spread function and report on their performance. We also discuss several cases where the microscope simulator has been used to solve real problems in biology.

Recent advances in optical-sectioning microscopy, along with novel fluorescent proteins and probes, give us the tools to image molecules and their interactions in space and time. Investigators using these tools routinely collect multichannel three-dimensional (3D) images and time series, but analyzing such complex datasets requires sophisticated visualization techniques. We here provide an overview of the principles and practices of 3D visualization of multichannel microscopic data. We also describe ImageSurfer, a new software package for volume visualization and data analysis. ImageSurfer is freely available (www.imagesurfer.org) and provides powerful interactive tools to explore and analyze complex multichannel 3D datasets. Although ImageSurfer is designed with fluorescent microscopy in mind, it is also effective for other types of data, including 3D datasets acquired by functional magnetic resonance imaging and EM tomography.

A vexing problem in cystic fibrosis (CF) pathogenesis has been to explain the high prevalence of Pseudomonas aeruginosa biofilms in CF airways. We speculated that airway surface liquid (ASL) hyperabsorption generates a concentrated airway mucus that interacts with P. aeruginosa to promote biofilms. To model CF vs. normal airway infections, normal (2.5% solids) and CF-like concentrated (8% solids) mucus were prepared, placed in flat chambers, and infected with an approximately 5 x 10(3) strain PAO1 P. aeruginosa. Although bacteria grew to 10(10) cfu/ml in both mucus concentrations, macrocolony formation was detected only in the CF-like (8% solids) mucus. Biophysical and functional measurements revealed that concentrated mucus exhibited properties that restrict bacterial motility and small molecule diffusion, resulting in high local bacterial densities with high autoinducer concentrations. These properties also rendered secondary forms of antimicrobial defense, e.g., lactoferrin, ineffective in preventing biofilm formation in a CF-like mucus environment. These data link airway surface liquid hyperabsorption to the high incidence of P. aeruginosa biofilms in CF via changes in the hydration-dependent physical-chemical properties of mucus and suggest that the thickened mucus gel model will be useful to develop therapies of P. aeruginosa biofilms in CF airways.

The transfer of electrons from one material to another is usually described in terms of energy conservation, with no attention being paid to momentum conservation. Here we present results on the junction resistance between a carbon nanotube and a graphite substrate and show that details of momentum conservation also can change the contact resistance. By changing the angular alignment of the atomic lattices, we found that contact resistance varied by more than an order of magnitude in a controlled and reproducible fashion, indicating that momentum conservation, in addition to energy conservation, can dictate the junction resistance in graphene systems such as carbon nanotube junctions and devices.

The nanoManipulator system adds a virtual reality interface to an atomic force microscope (AFM), thus providing a tool that enables the user not only to image but also to manipulate nanometer-sized molecular structures. As the AFM tip scans the surface of these structures, the tip-sample interaction forces are monitored, which in turn provide information about the frictional, mechanical, and topological properties of the sample. Computer graphics are used to reconstruct the surface for the user, with color or contours overlaid to indicate additional data sets. Moreover, by means of a force-feedback pen, which is connected to the scanning tip via software, the user can touch the surface under investigation to feel it and to manipulate objects on it. This system has been used to investigate carbon nanotubes, fibrin, DNA, adenovirus, and tobacco mosaic virus. Nanotubes have been bent, translated, and rotated to understand their mechanical properties and to investigate friction on the molecular level. AFM lithography is being combined with the nanoManipulator to investigate the electromechanical properties of carbon nanotubes. The rupture forces of fibrin and DNA have been measured. This article discusses how some of the graphics and interface features of the nanoManipulator made these novel investigations possible. Visitors have used the system to examine chromosomes, bacterial pili fibers, and nanochain aggregates (NCAs). Investigators are invited to apply to use the system as described on the web at http:@www.cs.unc.edu/Research/nano/doc/biovis it.html.

Understanding the relative motion of objects in contact is essential for controlling macroscopic lubrication and adhesion, for comprehending biological macromolecular interfaces, and for developing submicrometre-scale electromechanical devices. An object undergoing lateral motion while in contact with a second object can either roll or slide. The resulting energy loss and mechanical wear depend largely on which mode of motion occurs. At the macroscopic scale, rolling is preferred over sliding, and it is expected to have an equally important role in the microscopic domain. Although progress has been made in our understanding of the dynamics of sliding at the atomic level, we have no comparable insight into rolling owing to a lack of experimental data on microscopic length scales. Here we produce controlled rolling of carbon nanotubes on graphite surfaces using an atomic force microscope. We measure the accompanying energy loss and compare this with sliding. Moreover, by reproducibly rolling a nanotube to expose different faces to the substrate and to an external probe, we are able to study the object over its complete surface.

: In many cases in experimental science, the instrument interface becomes a limiting factor in the efficacy of carrying out unusual experiments or prevents the complete understanding of the acquired data. We have developed an advanced interface for scanning probe microscopy (SPM) that allows intuitive rendering of data sets and natural instrument control, all in real time. The interface, called the nanoManipulator, combines a high-performance graphics engine for real-time data rendering with a haptic interface that places the human operator directly into the feedback loop that controls surface manipulations. Using a hand-held stylus, the operator moves the stylus laterally, directing the movement of the SPM tip across the sample. The haptic interface enables the user to "feel" the surface by forcing the stylus to move up and down in response to the surface topography. In this way the user understands the immediate location of the tip on the sample and can quickly and precisely maneuver nanometer-scale objects. We have applied this interface to studies of the mechanical properties of nanotubes and to substrate-nanotube interactions. The mechanical properties of carbon nanotubes have been demonstrated to be extraordinary. They have an elastic modulus rivaling that of the stiffest material known, diamond, while maintaining a remarkable resistance to fracture. We have used atomic-force microscopy (AFM) to manipulate the nanotubes through a series of configuration that reveal buckling behavior and high-strain resilience. Nanotubes also serve as test objects for nanometer-scale contact mechanics. We have found that nanotubes will roll under certain conditions. This has been determined through changes in the images and through the acquisition of lateral force during manipulation. The lateral force data show periodic stick-slip behavior with a periodicity matching the perimeter of the nanotube.

The curling of a graphitic sheet to form carbon nanotubes produces a class of materials that seem to have extraordinary electrical and mechanical properties. In particular, the high elastic modulus of the graphite sheets means that the nanotubes might be stiffer and stronger than any other known material, with beneficial consequences for their application in composite bulk materials and as individual elements of nanometre-scale devices and sensors. The mechanical properties are predicted to be sensitive to details of their structure and to the presence of defects, which means that measurements on individual nanotubes are essential to establish these properties. Here we show that multiwalled carbon nanotubes can be bent repeatedly through large angles using the tip of an atomic force microscope, without undergoing catastrophic failure. We observe a range of responses to this high-strain deformation, which together suggest that nanotubes are remarkably flexible and resilient.

We present our results on the manipulation of individual viruses using an advanced interface for atomic force microscopes (AFMs). We show that the viruses can be dissected, rotated, and translated with great facility. We interpret the behavior of tobacco mosaic virus with a mechanical model that makes explicit the competition between sample-substrate lateral friction and the flexural rigidity of the manipulated object. The manipulation behavior of tobacco mosaic virus on graphite is shown to be consistent with values of lateral friction observed on similar interfaces and the flexural rigidity expected for macromolecular assemblies. The ability to manipulate individual samples broadens the scope of possible studies by providing a means for positioning samples at specific binding sites or predefined measuring devices. The mechanical model provides a framework for interpreting quantitative measurements of virus binding and mechanical properties and for understanding the constraints on the successful, nondestructive AFM manipulation of delicate samples.