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High throughput microscopy

Rui Yan, Seonah Moon, Samuel J Kenny, Ke Xu
As an elegant integration of the spatial and temporal dimensions of single-molecule fluorescence, single-molecule localization microscopy (SMLM) overcomes the diffraction-limited resolution barrier of optical microscopy by localizing single molecules that stochastically switch between fluorescent and dark states over time. While this type of super-resolution microscopy (SRM) technique readily achieves remarkable spatial resolutions of ∼10 nm, it typically provides no spectral information. Meanwhile, current scanning-based single-location approaches for mapping the positions and spectra of single molecules are limited by low throughput and are difficult to apply to densely labeled (bio)samples...
February 14, 2018: Accounts of Chemical Research
Deepti K Pillai, Elva Cha, Derek Mosier
Bovine respiratory disease (BRD) is a major problem for the cattle industry that is triggered by various environmental stressors, pathogens and host responses. Mannheimia hemolytica, an important bacterial component of BRD, are present within the nasopharayngeal region of normal calves as commensal biofilm communities. However, following stress there are changes in the nasopharyngeal microenvironment that triggers the transition of the commensal M. haemolytica into a pulmonary pathogen. The factors responsible for this transition in- vivo are unknown...
February 2018: Veterinary Microbiology
T J Heger, Ian J W Giesbrecht, Julia Gustavsen, Javier Del Campo, Colleen T E Kellogg, Kira M Hoffman, Ken Lertzman, William W Mohn, Patrick J Keeling
Although previous studies, mostly based on microscopy analyses of a few groups of protists, have suggested that protists are abundant and diverse in litter and moss habitats, the overall diversity of moss and litter associated protists remains elusive. Here, high-throughput environmental sequencing was used to characterize the diversity and community structure of litter and moss-associated protists along a gradient of soil drainage and forest primary productivity in a temperate rainforest in British Columbia...
February 7, 2018: Environmental Microbiology
Dushan N Wadduwage, Vijay Raj Singh, Heejin Choi, Zahid Yaqoob, Hans Heemskerk, Paul Matsudaira, Peter T C So
Imaging Fourier-transform spectroscopy (IFTS) is a powerful method for biological hyperspectral analysis based on various imaging modalities, such as fluorescence or Raman. Since the measurements are taken in the Fourier space of the spectrum, it can also take advantage of compressed sensing strategies. IFTS has been readily implemented in high-throughput, high-content microscope systems based on wide-field imaging modalities. However, there are limitations in existing wide-field IFTS designs. Non-common-path approaches are less phase-stable...
May 20, 2017: Optica
M Gesley, R Puri
A high throughput spectral image microscopy system is configured for rapid detection of rare cells in large populations. To overcome flow cytometry rates and use of fluorophore tags, a system architecture integrates sample mechanical handling, signal processors, and optics in a non-confocal version of light absorption and scattering spectroscopic microscopy. Spectral images with native contrast do not require the use of exogeneous stain to render cells with submicron resolution. Structure may be characterized without restriction to cell clusters of differentiation...
January 2018: Review of Scientific Instruments
Clare Bird, Kate F Darling, Ann D Russell, Jennifer S Fehrenbacher, Catherine V Davis, Andrew Free, Bryne T Ngwenya
Uncovering the complexities of trophic and metabolic interactions among microorganisms is essential for the understanding of marine biogeochemical cycling and modelling climate-driven ecosystem shifts. High-throughput DNA sequencing methods provide valuable tools for examining these complex interactions, although this remains challenging, as many microorganisms are difficult to isolate, identify and culture. We use two species of planktonic foraminifera from the climatically susceptible, palaeoceanographically important genus Neogloboquadrina, as ideal test microorganisms for the application of 16S rRNA gene metabarcoding...
2018: PloS One
Julia Niessl, Amy E Baxter, Daniel E Kaufmann
PURPOSE OF REVIEW: The long-lived HIV reservoir remains a major obstacle for an HIV cure. Current techniques to analyze this reservoir are generally population-based. We highlight recent developments in methods visualizing HIV, which offer a different, complementary view, and provide indispensable information for cure strategy development. RECENT FINDINGS: Recent advances in fluorescence in situ hybridization techniques enabled key developments in reservoir visualization...
January 27, 2018: Current HIV/AIDS Reports
Mansour S Alturki, Ngolui Rene Fuanta, Madison A Jarrard, Judith V Hobrath, Douglas C Goodwin, Thankhoe A Rants'o, Angela I Calderón
Single dose high-throughput screening (HTS) followed by dose-response evaluations is a common strategy for the identification of initial hits for further development. Early identification and exclusion of false positives is a cost-saving and essential step in early drug discovery. One of the mechanisms of false positive compounds is the formation of aggregates in assays. This study evaluates the mechanism(s) of inhibition of a set of 14 compounds identified previously as actives in Mycobacterium tuberculosis (Mt) cell culture screening and in vitro actives in Mt shikimate kinase (MtSK) assay...
December 5, 2017: Bioorganic & Medicinal Chemistry Letters
Quigly Dragotakes, Arturo Casadevall
The purpose of this technique is to provide a consistent, accurate, and manageable process for large numbers of polysaccharide capsule measurements. First, a threshold image is generated based on intensity values uniquely calculated for each image. Then, circles are detected based on contrast between the object and background using the well-established Circle Hough Transformation (CHT) algorithm. Finally, the detected cell capsules and bodies are matched according to center coordinates and radius size, and data is exported to the user in a manageable spreadsheet...
January 11, 2018: Journal of Visualized Experiments: JoVE
Lucas Moritz Wiggenhauser, Katharina Kohl, Nadine Dietrich, Hans-Peter Hammes, Jens Kroll
Diabetic retinopathy is the leading cause of blindness among middle-aged adults. The rising prevalence of diabetes worldwide will make the prevention of diabetic microvascular complications one of the key research fields of the next decades. Specialized, targeted therapy and novel therapeutic drugs are needed to manage the increasing number of patients at risk of vision-loss. The zebrafish is an established animal model for developmental research questions with increasing relevance for modeling metabolic multifactorial disease processes...
December 26, 2017: Journal of Visualized Experiments: JoVE
Isaac Martens, Elizabeth A Fisher, Dan Bizzotto
The characterization of bio-functionalized surfaces such as alkanethiol self-assembled monolayers (SAMs) on gold modified with DNA or other biomolecules is a challenging analytical problem and access to a routine method is desirable. Despite substantial investigation from structural and mechanistic perspectives, robust and high-throughput metrology tools for SAMs remain elusive but essential for the continued development of these devices. We demonstrate that scanning electron microscopy (SEM) can provide excellent contrast of the molecular interface at every step of SAM functionalization...
January 24, 2018: Langmuir: the ACS Journal of Surfaces and Colloids
Karthika Suryaletha, Joby John, Megha Periyappilly Radhakrishnan, Sanil George, Sabu Thomas
Chronic diabetic foot is a global burden affecting millions of people, and the chronicity of an ulcer is directly linked to the diverse bacterial burden and its biofilm mode of infection. The bacterial diversity of 100 chronic diabetic ulcer samples was profiled via traditional culturing method as well as metagenomic approach by sequencing the 16S rRNA V3 hyper-variable region on Illumina Miseq Platform (Illumina, Inc., San Diego, CA). All the relevant clinical metadata, including duration of diabetes, grade of ulcer, presence of neuropathy, and glycaemic level, were noted and correlated with the microbiota...
January 22, 2018: International Wound Journal
A Mira
Understanding the microbiology of dental caries is not a mere academic exercise; it provides the basis for preventive, diagnostic, and treatment strategies and gives the dentist a theoretical framework to become a better professional. The last years have seen the development of new research methodologies, ranging from high-throughput sequencing or "omics" techniques to new fluorescence microscopy applications and microfluidics, which have allowed the study of the oral microbiome to an unprecedented level of detail...
February 2018: Advances in Dental Research
Sabrina Simoncelli, Yi Li, Emiliano Cortes, Stefan A Maier
Self-assembly processes allow designing and creating complex nanostructures using molecules as building blocks and surfaces as scaffolds. This autonomous driven construction is possible due to a complex thermodynamic balance of molecule-surface interactions. As such, nanoscale guidance and control over this process is hard to achieve. Here we use the highly localized light-to-chemical-energy conversion of plasmonic materials to spatially cleave Au-S bonds on pre-determined locations within a single nanoparticle, enabling a high degree of control over this archetypal system for molecular self-assembly...
January 18, 2018: ACS Nano
F Merola, P Memmolo, L Miccio, M Mugnano, P Ferraro
High-throughput single-cell analysis is a challenging target for implementing advanced biomedical applications. An excellent candidate for this aim is label-free tomographic phase microscopy (TPM). In this paper, some of the methods used to obtain TPM are reviewed, analyzing advantages and disadvantages of each of them. Moreover, an alternative tomographic technique is described for live cells analysis, and future trends of the method are foreseen. In particular, by exploiting random rolling of cells while they are flowing along a microfluidic channel, it is possible to obtain phase-contrast tomography thus obtaining complete retrieval of both 3D-position and orientation of rotating cells...
January 13, 2018: Methods: a Companion to Methods in Enzymology
Jianxin Sun, L Moore, Wei Xue, James Kim, Maciej Zborowski, Jeffrey J Chalmers
Magnetic separation of cells has been, and continues to be, widely used in a variety of applications, ranging from healthcare diagnostics to detection of food contamination. Typical, these technologies require cells labeled with antibody magnetic particle conjugate and a high magnetic energy gradient created in the flow containing the labeled cells (i.e. a column packed with magnetically inducible material), or dense packing of magnetic particles next to the flow cell. Such designs while creating high magnetic energy gradients, are not amenable to easy, highly detailed, mathematic characterization...
January 16, 2018: Biotechnology and Bioengineering
Csilla Brasko, Kevin Smith, Csaba Molnar, Nora Farago, Lili Hegedus, Arpad Balind, Tamas Balassa, Abel Szkalisity, Farkas Sukosd, Katalin Kocsis, Balazs Balint, Lassi Paavolainen, Marton Z Enyedi, Istvan Nagy, Laszlo G Puskas, Lajos Haracska, Gabor Tamas, Peter Horvath
Quantifying heterogeneities within cell populations is important for many fields including cancer research and neurobiology; however, techniques to isolate individual cells are limited. Here, we describe a high-throughput, non-disruptive, and cost-effective isolation method that is capable of capturing individually targeted cells using widely available techniques. Using high-resolution microscopy, laser microcapture microscopy, image analysis, and machine learning, our technology enables scalable molecular genetic analysis of single cells, targetable by morphology or location within the sample...
January 15, 2018: Nature Communications
Xuan Cao, Gang Lei, Jingjing Feng, Qi Pan, Xiaodong Wen, Yan He
Creating color difference and improving the color resolution in digital imaging is crucial for better application of color analy-sis. Herein, a novel color modulation analysis strategy was developed by using a homemade tunable multiband laser illumi-nation device, in which the portions of R, G, and B components of the illumination light are discretionarily adjustable, and hence the sample color could be visually modulated continuously in the RGB color space. Through this strategy, the color appearance of single gold nanorods (AuNRs) under dark-field microscopy, was migrated from the spectrally insensitive red region to the spectrally sensitive green-yellow-red region...
January 15, 2018: Analytical Chemistry
Anne W Lucky, Neha Dagaonkar, Karen Lammers, Ammar Husami, Diane Kissell, Kejian Zhang
BACKGROUND: Historically, diagnosis of epidermolysis bullosa has required skin biopsies for electron microscopy, direct immunofluorescence to determine which gene(s) to choose for genetic testing, or both. METHODS: To avoid these invasive tests, we developed a high-throughput next-generation sequencing (NGS)-based diagnostic assay called EBSEQ that allows simultaneous detection of mutations in 21 genes with known roles in epidermolysis bullosa pathogenicity. Mutations are confirmed with traditional Sanger sequencing...
January 15, 2018: Pediatric Dermatology
Di Jin, Renjie Zhou, Zahid Yaqoob, Peter T C So
Optical diffraction tomography (ODT) is an emerging microscopy technique for three-dimensional (3D) refractive index (RI) mapping of transparent specimens. Recently, the digital micromirror device (DMD) based scheme for angle-controlled plane wave illumination has been proposed to improve the imaging speed and stability of ODT. However, undesired diffraction noise always exists in the reported DMD-based illumination scheme, which leads to a limited contrast ratio of the measurement fringe and hence inaccurate RI mapping...
January 8, 2018: Optics Express
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