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Laser capture microscopy

Jonathan M Grimes, David R Hall, Alun W Ashton, Gwyndaf Evans, Robin L Owen, Armin Wagner, Katherine E McAuley, Frank von Delft, Allen M Orville, Thomas Sorensen, Martin A Walsh, Helen M Ginn, David I Stuart
Macromolecular crystallography (MX) has been a motor for biology for over half a century and this continues apace. A series of revolutions, including the production of recombinant proteins and cryo-crystallography, have meant that MX has repeatedly reinvented itself to dramatically increase its reach. Over the last 30 years synchrotron radiation has nucleated a succession of advances, ranging from detectors to optics and automation. These advances, in turn, open up opportunities. For instance, a further order of magnitude could perhaps be gained in signal to noise for general synchrotron experiments...
February 1, 2018: Acta Crystallographica. Section D, Structural Biology
G Fichi, S Carboni, J E Bron, J Ireland, M J Leaver, G Paladini
Ciliates have been reported as pathogens of many species of economically important bivalves. Mussel protozoan X (MPX), is an uncharacterised intracellular ciliate of mussels and has been widely reported in Mytilus spp. around the world. In order to characterise this ciliate, Mytilus edulis samples were collected from a site on the West coast of Scotland, and four different fixatives for histological examination were tested. Fresh preparations of mussel digestive glands were also examined by laser scanning confocal microscopy...
March 1, 2018: Journal of Invertebrate Pathology
Akiko Kunita, Vanessa Baeriswyl, Claudia Meda, Erik Cabuy, Kimiko Takeshita, Enrico Giraudo, Andreas Wicki, Masashi Fukayama, Gerhard Christofori
Tumor invasion is a critical first step in the organismic dissemination of cancer cells and the formation of metastasis in distant organs, the most important prognostic factor and the actual cause of death in most of the cancer patients. We report here that the cell surface protein podoplanin (PDPN), a potent inducer of cancer cell invasion, is conspicuously expressed by the invasive front of squamous cell carcinomas (SCC) of the cervix in patients and in the transgenic HPV/E2 mouse model of cervical cancer...
February 16, 2018: American Journal of Pathology
Adriele Prina-Mello, Namrata Jain, Baiyun Liu, Jason I Kilpatrick, Melissa A Tutty, Alan P Bell, Suzanne P Jarvis, Yuri Volkov, Dania Movia
Alternative models such as three-dimensional (3D) cell cultures represent a distinct milestone towards capturing the realities of cancer biology in vitro and reduce animal experimentation in the preclinical stage of drug discovery. Significant work remains to be done to understand how substrates used in in vitro alternatives influence cancer cells phenotype and drug efficacy responses, so that to accurately link such models to specific in vivo disease scenarios. Our study describes how the morphological, mechanical and biochemical properties of adenocarcinoma (A549) cells change in response to a 3D environment and varying substrates...
February 2018: Tissue & Cell
Sneha Jani
Bacterial chemotaxis in response to continuous chemical gradients has been extensively studied at the individual cell and population levels using a variety of well-established in vitro methods (Englert et al., Microfluidic techniques for the analysis of bacterial chemotaxis. Methods Mol Biol 571:1-23, 2009). In nature, bacteria are surrounded by heterogeneous chemical gradients; hence, it is essential to understand chemotaxis behavior under such conditions. Here, we describe a setup that allows visualization of the chemotaxis response of motile cells to the complex microenvironment of a biofilm maintained under static conditions...
2018: Methods in Molecular Biology
Suge Zhang, Hongxia Sun, Hongbo Chen, Qian Li, Aijiao Guan, Lixia Wang, Yunhua Shi, Shujuan Xu, Meirong Liu, Yalin Tang
BACKGROUND: Direct detection of G-quadruplexes in human cells has become an important issue due to the vital role of G-quadruplex related to biological functions. Despite several probes have been developed for detection of the G-quadruplexes in cytoplasm or whole cells, the probe being used to monitor the nucleolar G-quadruplexes is still lacking. METHODS: Formation of the nucleolar G-quadruplex structures was confirmed by using circular dichroism (CD) spectroscopy...
February 1, 2018: Biochimica et Biophysica Acta
David J Glugla, Madeline B Chosy, Marvin D Alim, Amy C Sullivan, Robert R McLeod
Precise direct-write lithography of 3D waveguides or diffractive structures within the volume of a photosensitive material is hindered by the lack of metrology that can yield predictive models for the micron-scale refractive index profile in response to a range of exposure conditions. We apply the transport of intensity equation in conjunction with confocal reflection microscopy to capture the complete spatial frequency spectrum of isolated 10 μm-scale gradient-refractive index structures written by single-photon direct-write laser lithography...
January 22, 2018: Optics Express
Yosif Manavski, Tina Lucas, Simone F Glaser, Lena Dorsheimer, Stefan Günther, Thomas Braun, Michael A Rieger, Andreas M Zeiher, Reinier A Boon, Stefanie Dimmeler
RATIONALE: Vascularization is critical to maintain organ function. Although many molecular pathways were shown to control vessel growth, the genuine process of capillary formation under different conditions is unclear. OBJECTIVE: Here, we elucidated whether clonal expansion contributes to vessel growth by using Confetti mice for genetic tracing of clonally expanding endothelial cells (ECs). METHODS AND RESULTS: In postnatal retina angiogenesis, we predominantly observed random distribution of fluorescence labeled ECs indicative of random integration or cell mixing...
March 2, 2018: Circulation Research
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
Brian E Walczak, Matthew S Nies, Darrin J Trask, Scott Hetzel, Patrick J Roney, Matthew W Squire, Geoffrey S Baer
BACKGROUND: Insertion force has been shown to significantly reduce chondrocyte viability during osteochondral allograft transplantation. How graft size influences the required insertion force and chondrocyte viability has yet to be determined. Hypothesis/Purpose: The purpose was to characterize how graft size influences insertion force requirements and chondrocyte viability during osteochondral transplantation. The hypothesis was that larger grafts would require greater force and reduce chondrocyte viability...
January 1, 2018: American Journal of Sports Medicine
D G Norman, D G Watson, B Burnett, P M Fenne, M A Williams
Toolmark analysis involves examining marks created on an object to identify the likely tool responsible for creating those marks (e.g., a knife). Although a potentially powerful forensic tool, knife mark analysis is still in its infancy and the validation of imaging techniques as well as quantitative approaches is ongoing. This study builds on previous work by simulating real-world stabbings experimentally and statistically exploring quantitative toolmark properties, such as cut mark angle captured by micro-CT imaging, to predict the knife responsible...
December 30, 2017: Forensic Science International
José A Rodrigo, Juan M Soto, Tatiana Alieva
The refractive index (RI) is an important optical characteristic that is often exploited in label-free microscopy for analysis of biological objects. A technique for 3D RI reconstruction of living cells has to be fast enough to capture the cell dynamics and preferably needs to be compatible with standard wide-field microscopes. To solve this challenging problem, we present a technique that provides fast measurement and processing of data required for real-time 3D visualization of the object RI. Specifically, the 3D RI is reconstructed from the measurement of bright-field intensity images, axially scanned by a high-speed focus tunable lens mounted in front of a sCMOS camera, by using a direct deconvolution approach designed for partially coherent light microscopy in the non-paraxial regime...
December 1, 2017: Biomedical Optics Express
Carolina Stenfeldt, Michael Eschbaumer, George R Smoliga, Luis L Rodriguez, James Zhu, Jonathan Arzt
Long-term persistent viral infections cause substantial morbidity and associated economic losses in human and veterinary contexts. Yet, the mechanisms associated with establishment of persistent infections are poorly elucidated. We investigated immunomodulatory mechanisms associated with clearance versus persistence of foot-and-mouth disease virus (FMDV) in micro-dissected compartments of the bovine nasopharynx by microarray. The use of laser-capture microdissection allowed elucidation of differential gene regulation within distinct anatomic compartments critical to FMDV infection...
December 19, 2017: Scientific Reports
Ryohsuke Kohmoto, Takatoshi Kobayashi, Takaki Sato, Daisaku Kimura, Masanori Fukumoto, Kensuke Tajiri, Teruyo Kida, Tsunehiko Ikeda
BACKGROUND: To report a case of proliferative diabetic retinopathy (PDR) exhibiting the appearance of scintillating particles presumed to be crystallin inside the intravitreal cavity after laser photocoagulation. CASE PRESENTATION: A 56-year-old male patient presented at our outpatient clinic after becoming aware of decreased vision in his right eye. Ocular examination performed at the patient's initial visit revealed a massive preretinal macular hemorrhage due to PDR in his right eye...
December 19, 2017: BMC Ophthalmology
Jesse K Adams, Vivek Boominathan, Benjamin W Avants, Daniel G Vercosa, Fan Ye, Richard G Baraniuk, Jacob T Robinson, Ashok Veeraraghavan
Modern biology increasingly relies on fluorescence microscopy, which is driving demand for smaller, lighter, and cheaper microscopes. However, traditional microscope architectures suffer from a fundamental trade-off: As lenses become smaller, they must either collect less light or image a smaller field of view. To break this fundamental trade-off between device size and performance, we present a new concept for three-dimensional (3D) fluorescence imaging that replaces lenses with an optimized amplitude mask placed a few hundred micrometers above the sensor and an efficient algorithm that can convert a single frame of captured sensor data into high-resolution 3D images...
December 2017: Science Advances
Arthur B Volovetsky, Vladimir S Sukhov, Irina V Balalaeva, Varvara V Dudenkova, Natalia Yu Shilyagina, Аlexey V Feofanov, Anastasija V Efremenko, Mikhail A Grin, Andrey F Mironov, Igor B Sivaev, Vladimir I Bregadze, Anna V Maslennikova
The necessary precondition for efficient boron neutron capture therapy (BNCT) is control over the content of isotope10 B in the tumor and normal tissues. In the case of boron-containing porphyrins, the fluorescent part of molecule can be used for quantitative assessment of the boron content. Study Objective: We performed a study of the biodistribution of the chlorin e ₆-Cobalt bis(dicarbollide) conjugate in carcinoma-bearing Balb/c mice using ex vivo fluorescence imaging, and developed a mathematical model describing boron accumulation and release based on the obtained experimental data...
November 28, 2017: International Journal of Molecular Sciences
Robert K Y Cheng, Rafael Abela, Michael Hennig
Past decades have shown the impact of structural information derived from complexes of drug candidates with their protein targets to facilitate the discovery of safe and effective medicines. Despite recent developments in single particle cryo-electron microscopy, X-ray crystallography has been the main method to derive structural information. The unique properties of X-ray free electron laser (XFEL) with unmet peak brilliance and beam focus allow X-ray diffraction data recording and successful structure determination from smaller and weaker diffracting crystals shortening timelines in crystal optimization...
November 8, 2017: Essays in Biochemistry
Norbert Kiss, Ádám Krolopp, Kende Lőrincz, András Bánvölgyi, Róbert Szipőcs, Norbert Wikonkál
Basal cell carcinoma (BCC) is the most common malignancy in Caucasians. Nonlinear microscopy has been previously utilized for the imaging of BCC, but the captured images do not correlate with H&E staining. Recently, Freudiger et al. introduced a novel method to visualize tissue morphology analogous to H&E staining, using coherent anti-Stokes Raman scattering (CARS) technique. In our present work, we introduce a novel algorithm to post-process images obtained from dual vibration resonance frequency (DVRF) CARS measurements to acquire high-quality pseudo H&E images of BCC samples...
November 4, 2017: Pathology Oncology Research: POR
Manikkuwadura Eranda Harshan De Silva, Alexis Ceecee Zhang, Amalia Karahalios, Holly Rose Chinnery, Laura Elizabeth Downie
INTRODUCTION: Laser scanning in vivo confocal microscopy (IVCM) enables non-invasive, high-resolution imaging of the cornea. In recent years, there has been a vast increase in researchers using laser scanning IVCM to image and quantify corneal nerve parameters. However, a range of methodological approaches have been adopted. The primary aim of this systematic review is to critically appraise the reported method(s) of primary research studies that have used laser scanning IVCM to quantify corneal sub-basal nerve plexus (SBNP) parameters in humans, and to examine corneal nerve parameters in healthy individuals...
November 3, 2017: BMJ Open
Imogen Sparkes, Rhiannon R White, Benjamin Coles, Stanley W Botchway, Andy Ward
Optical tweezers have been used to trap and micromanipulate several biological specimens ranging from DNA, macromolecules, organelles to single celled organisms. Using a combination of the refraction and scattering of laser light from a focused laser beam, refractile objects are physically captured and can be moved within the surrounding media. The technique is routinely used to determine biophysical properties such as the forces exerted by motor proteins. Here, we describe how optical tweezers combined with total internal reflection fluorescence (TIRF) microscopy can be used to assess physical interactions between organelles, more specifically the ER and Golgi bodies in plant cells...
2018: Methods in Molecular Biology
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