Low Magnification Images Research Articles

Archaeometric characterisation and origin of black coatings on prehistoric pottery

Black or dark grey vessel surfaces with characteristic metallic lustre are recognised from various archaeological contexts throughout Europe. This feature is commonly attributed to the application of graphite-bearing coatings onto ceramic vessels. However, recent experimental studies have shown that a very similar visual effect can alternatively be achieved by polishing and, subsequently, firing in a reducing atmosphere (so-called smudging). In this study, experimentally manufactured ceramics as well as samples of prehistoric ware (dated to the Neolithic Age, late Bronze Age and early Iron Age) have been analysed to find the distinguishing features between graphite-based and non-mineral black coatings. In the low-magnification BSE images the well-preserved graphite-coated surfaces are characterized by a distinctive spotty pattern with numerous dark grey angular fragments of monomineralic or nearly monomineralic metamorphic rocks scattered in the clay groundmass. Provided that polished thin sections are skilfully prepared, individual graphite plates can be easily recognized in the near-surface layer of the sherds using polarized reflected light microscopy. A relatively homogeneous appearance is typical of the low-magnification BSE images of surfaces coated by smudging. At higher magnifications, presence of blurred darkish stains (presumably organic-derived, as indicated by elevated levels of biogenic elements) is characteristic of this kind of non-mineral black coatings. SEM-EDS and optical microscopy have been supplemented by micro-Raman spectroscopy, which allows differentiation between various carbonaceous phases.

The Role of MicroRNA-21 in Venous Neointimal Hyperplasia: Implications for Targeting miR-21 for VNH Treatment.

The molecular mechanism of hemodialysis access arteriovenous fistula (AVF) failure due to venous neointimal hyperplasia (VNH) is not known. The role of microRNA-21 (miR-21) in VNH associated with AVF failure was investigated by performing in vivo and in vitro experiments. In situ hybridization results revealed that miR-21 expression increased and was associated with fibroblasts in failed AVFs from patients. In a murine AVF model, qRT-PCR gene expression results showed a significant increase in miR-21 and a decrease in miR-21 target genes in graft veins (GVs) compared to contralateral veins in mouse AVF. miR-21 knockdown in GVs was performed using a lentivirus-mediated small hairpin RNA (shRNA), and this improved AVF patency with a decrease in neointima compared to control GVs. Moreover, loss of miR-21 in GVs significantly decreased the Tgfβ1, Col-Ia, and Col-Iva genes. Immunohistochemistry demonstrated a significant decrease in myofibroblasts and proliferation with an increase in terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining in miR-21-knockdown vessels, along with a decrease in hypoxia-inducible factor-1 alpha (HIF-1α) and phospho-SMAD2 (pSMAD-2) and phospho-SMAD3 (pSMAD-3) and an increase in phosphatase and tensin homolog (PTEN) staining. Hypoxic fibroblast knockdown for miR-21 showed a significant decrease in Tgfβ-1 expression and pSMAD-2 and -3 levels and a decrease in myofibroblasts. These results indicate that miR-21 upregulation causes VNH formation by fibroblast-to-myofibroblast differentiation.

Evaluation of Region of Interest Digital Cytology Compared to Light Microscopy for Veterinary Medicine.

Digital microscopy (DM) has been employed for primary diagnosis in human medicine and for research and teaching applications in veterinary medicine, but there are few veterinary DM validation studies. Region of interest (ROI) digital cytology is a subset of DM that uses image-stitching software to create a low-magnification image of a slide, then selected ROI at higher magnification, and stitches the images into a relatively small file of the embedded magnifications. This study evaluated the concordance of ROI-DM compared to traditional light microscopy (LM) between 2 blinded clinical pathologists. Sixty canine and feline cytology samples from a variety of anatomic sites, including 31 cases of malignant neoplasia, 15 cases of hyperplastic or benign neoplastic lesions, and 14 infectious/inflammatory lesions, were evaluated. Two separate nonblinded adjudicating clinical pathologists evaluated the reports and diagnoses and scored each paired case as fully concordant, partially concordant, or discordant. The average overall concordance (full and partial concordance) for both pathologists was 92%. Full concordance was significantly higher for malignant lesions than benign. For the 40 neoplastic lesions, ROI-DM and LM agreed on general category of tumor type in 78 of 80 cases (98%). ROI-DM cytology showed robust concordance with the current gold standard of LM cytology and is potentially a viable alternative to current LM cytology techniques.

Synergistic Hybrid MoS2/Fe2O3 Composites for Efficient Hydrogen Evolution in Alkaline Media

It is highly desirable to generate H2 by splitting water (2H2O → O2 + 2H2), because the storage of a large amount of energy (4.92 eV) can be realized in the molecular transformation with no emission of greenhouse gases and other pollutants.(1) The hydrogen evolution reaction (HER) is the cathodic half reaction of water splitting, which is the actual process that produces H2 needed to fuel the purported "hydrogen-based economy".(2) HER on earth-abundant molybdenum disulfide (MoS2) in acidic media is a robust process, but it is kinetically retarded in alkaline media. Thus, improving the sluggish kinetics for HER in alkaline media is crucial for advancing the performance of water-alkali electrolyzers. Here, we demonstrate a dramatic enhancement of HER kinetics in base by judiciously hybridizing vertical MoS2 sheets with another earth-abundant material, iron oxide (Fe2O3). The field-emission scanning electron microscopy (FESEM) images in Figure 1a show that the MoS2/Fe2O3 composite is composed of a particle-like structure of Fe2O3 grown closely on the edge surfaces of the vertically aligned MoS2 sheets. The highly ordered lattice planes in the high-resolution TEM (HRTEM) image of MoS2/Fe2O3 composite are clearly visible in Figure 1b. The d-spacing of 0.63 nm and 0.37 nm, which are corresponds to the (002) and (012) facets of MoS2 and Fe2O3 crystals, respectively, suggests the formation of an interface between the (002) facet of MoS2 and the neighboring (012) facet of Fe2O3 in the composite. Benefiting from the desirable structural characteristics, the MoS2/Fe2O3 interfaces synergistically favor the chemisorption of H (on MoS2) and OH (on Fe2O3), and thus can effectively accelerate the water dissociation step and thus the overall HER catalysis. The resultant MoS2/Fe2O3 heterostructures exhibit much better HER performance than bare MoS2 and outstanding stability in base (Figure 1c and d). Our synergistic hybridization of MoS2 with metal oxide materials enlightens the design and fabrication of promising heterostructures for hydrogen evolution in alkaline media. Figure Caption Figure 1. (a) High- and (inset) low-magnification FESEM images of MoS2/Fe2O3 composite. (b) P High-magnification TEM image of the MoS2/Fe2O3 composite, showing their interfaces. (c) Polarization curves of the MoS2, Fe2O3 and MoS2/Fe2O3 composite catalysts in 1M KOH solution at a scan rate of 5 mV/s. (d) Chronopotentiometry responses (η ~ t) recorded from MoS2/NiCo-LDH composite at high current densities of -10 mA/cm2. References J. L. Dempsey, B. S. Brunschwig, J. R. Winkler, H. B. Gray, Hydrogen evolution catalyzed by cobaloximes. Acc. Chem. Res. 42, 1995-2004 (2009).J. Hu et al., Hydrogen evolution electrocatalysis with binary-nonmetal transition metal compounds. J. Mater. Chem. A 5, 5995-6012 (2017). Figure 1

Multi-level magnification correlation hashing for scalable histopathological image retrieval

Content-based Medical Image Retrieval (CBMIR) presents promising results in computer-aided diagnosis which provides the clinicians with interpretative guidance based on the visual similarity. In this paper, we focus on the tasks of histopathological image retrieval for breast cancer diagnosis. Instead of the holistic-based and cell-based methods proposed in previous works, we propose a multi-magnification correlation hashing (MMCH) framework that learns the discriminative binary codes for histopathological images. The patch-link graph is constructed with both low and high magnification images where the semantic links are propagated from the pre-labeled patches to unlabeled data. We then learn the binary hashing codes based on the global labels from the low-magnification images with structure-regularization. The experiments on BreakHis dataset and PLOSONE dataset demonstrate that MMCH outperforms the previous hashing methods on histopathological image retrieval.

Densely-Connected Multi-Magnification Hashing for Histopathological Image Retrieval.

Content-based medical image retrieval is an important computer-aided diagnosis technique providing the clinicians with interpretative references based on visual similarity. In this paper, we focus on the tasks of histopathological image retrieval for breast cancer diagnosis. The densely-connected multi-magnification (DCMMH) framework is proposed to generate the discriminative binary codes by exploiting the histopathological images with multiple magnification factors. The low-magnification images are boosted by the accumulated similarity based on local patches that also regularize the feature learning of high-magnification images. In order to fully utilize the information across different magnification levels, a densely-connected architecture is finally deployed for high-low magnification pairs of datasets. Experiments on BreakHis dataset demonstrate that, DCMMH outperforms the previous hashing methods on histopathological image retrieval.

LASIE: Large Area Spectroscopic Imaging Ellipsometry for Characterizing Multi-Layered Film Structures

In this investigation, we describe a spectroscopic imaging ellipsometry for large area measurements, named as large area spectroscopic ellipsometry (LASIE). LASIE uses a broadband light source and an imaging spectrometer in order to obtain the spectral-spatial intensity images corresponding to a measurement line of a specimen and it can characterize the 3D multi-film structures with the aid of lateral scanning. Opposed to the typical high resolution imaging ellipsometry with the small field of view (FOV), LASIE uses a low magnification imaging lens to enlarge the measurement area and line profiles of multi-layered film structure can be reconstructed at a single operation based on the operation of the spectroscopic imaging spectrometer. In the experiment, 3- and 4-layered film specimen were measured after the system calibration and the 3D film thickness profiles of all film layers were obtained with 1 nm repeatability.

Ga2O3/Si and Al2O3/Si Room-Temperature Wafer Bonding Using in-Situ Deposited Si Thin Film

Recently, β-Ga2O3 has caught attentions in the field of high-voltage power-device applications due to its excellent material properties and suitability for mass production1). Thermal conductivity of Ga2O3 is, however, low, and bonding Ga2O3 to high-thermal conductivity substrate material is desired to improve heat dissipation of power devices. For the bonding between dissimilar materials, low temperature process is desirable to avoid the problems caused by thermal expansion mismatch. Surface activated bonding (SAB), which can achieve strong bonding without any heat treatments, is one of the candidates for the purpose, however bonding strength of oxide materials is low compared to metals and semiconductors2). In this paper, we propose bonding process using in-situ deposition of Si thin film on oxide materials, which can be performed by a standard SAB apparatus without any modification. By the process, we tried bonding of Ga2O3 and Al2O3 to Si. In the bonding experiments, a 15 x 10-mm2, 500-μm-thick, single-side polished chip of (010) β-Ga2O3 (TAMURA Corporation) was used. 100-mm-diameter, 650-μm-thick, single-side-polished, c-face sapphire wafers and 100-mm-diameter, 400-μm-thick, double-side-polished, (100) Si wafers were also used. A pair of Ga2O3 and Si specimens were introduced into an apparatus for the surface activated bonding (MWB-12ST, Mitsubishi Heavy Industries Machine Tool Co. Ltd.). After setting specimens on stages in the bonding chamber, Ar fast atom beam was irradiated to the Ga2O3 chip for 10 seconds to remove adsorbed surface contaminants. Then, the Ar fast atom beam was irradiated to the Si wafer for 60 seconds to remove surface contaminants and an oxide layer. During this surface activation step for Si, some of sputtered Si atoms were deposited on the Ga2O3 surface and form a thin Si layer. Immediately after the surface activation and Si layer deposition step, Ga2O3 and Si specimens were mated in the vacuum and pressed. Sapphire and Si wafers were bonded by the same procedure. The Ga2O3 chip was successfully bonded to the Si wafer except the area near the edge of the chip as shown in figures. The unbonded area is supposed to be caused by the edge-sagging formed during polishing process. Sapphire wafers were also successfully bonded to Si wafers. The Ga2O3/Si bonded specimen was cut into two pieces at the center of the Ga2O3 chip by dicing saw, and strength of Ga2O3/Si bonded specimen was evaluated by the dicing test from Si side. No cleavage from the bonding interface was observed after dicing test (lower-left in figures). Sapphire/Si bonding strength was evaluated by the blade test 3) as well as the dicing test. The bonding was so strong that no crack initiated at the bonding interface and Si wafer was cracked by blade insertion. Micro structure of the Ga2O3/Si and sapphire/Si interface was investigated by TEM. No cleavage was observed in low magnification images. In a HRTEM image of Ga2O3/Si interface (lower-right in figures), an amorphous intermediate layer was observed. Considering bonding step, the intermediate layer was resulted from deposited Si layer on Ga2O3 surface and damaged layer on Si surface formed during Ar fast beam irradiation4). In-situ deposition of Si thin film on oxide wafer is proved to be effective for strong bonding formation at room-temperature. Ga2O3 and sapphire were successfully bonded to Si wafers. This process is expected to provide a promising method to improve heat dissipation in Ga2O3 devices, as well as integration of oxides to other materials with large thermal expansion mismatch. This work is partly supported by International Joint Research Program for Innovative Energy Technology, METI. 1) M. Higashiwaki, K. Sasaki, T. Kamimura, M. Hoi Wong, D. Krishnamurthy, A. Kuramata, T. Masui, and S. Yamakoshi, Appl. Phys. Lett., 103(2013), 1. 2) H. Takagi, R. Maeda, T. R. Chung, and T. Suga, Sensors Actuators, A 70(1998), 164. 3) W. P. Maszara, G. Goetz, A. Caviglia, and J. B. McKitterick, J. Appl. Phys., 64(1988), 4943. 4) H. Takagi, R. Maeda, N. Hosoda, and T. Suga, Japanese J. Appl. Physics, 38(1999), 1589. Figure 1

Robotic Remote Controlled Stereo Slit Lamp.

PurposeOur purpose was to develop a robotic remotely operated stereo slit lamp system allowing three-dimensional stereo viewing and recording of the patient's examination via local area network, Internet, and satellite.MethodsA commercial slit lamp was modified to accept motors and servos to permit control of all optical and mechanical components of the device. The custom graphical user interface with dual high-resolution real-time stereoscopic imaging, control/position indicators, overview video, and audio were transmitted via local area network, Internet, and satellite. Under University of Miami Institutional Review Board authorization, Internet connectivity enabled multiple examiners to simultaneously view and control the slit lamp and to collaboratively discuss diagnosis and treatment options. The remote clinicians used a tablet, laptop, or desktop computer to view and control the slit lamp.ResultsThe network, Internet, satellite-connected system was controllable from the United States, Europe, and Canada while acquiring high-resolution, real-time video in all subjects. Control of the slit lamp through Ethernet, WiFi, and 4G exhibited total system latencies of 464 ± 58, 483 ± 64, and 870 ± 66 milliseconds when transmitting within the continent, and Ethernet control exhibited a latency of 606 ± 130 milliseconds when transmitting between continents. High- and low-magnification images of healthy volunteers were acquired by a remote clinician.ConclusionsThe robotic remotely operated stereo slit lamp system allows three-dimensional stereo viewing and recording of the patient's examination via local area network, Internet, and satellite.Translational RelevanceThe robotic remotely controlled stereo slit lamp system enables remote examination of human subjects.

Natural End Member Samples of Pyrope and Grossular: A Cathodoluminescence-Microscopy and -Spectra Case Study

Garnet is one of the most significant minerals in metamorphic rocks, that provides key information on prograde, peak-metamorphic and retrograde parts of the pressure-temperature (PT) path. Such results require a detailed knowledge of its different growth domains. For iron-poor compositions, the cathodoluminescence (CL) microscopy is an important and often overlooked method and allows to identify the internal structures of all garnet grains in one thin section within only a few seconds. The advantage of the CL-microscope is to deliver low magnification images in true color, not only of garnet but also, for instance, of other rock forming silicates, carbonates, sulfates, etc., of metamorphic, but also of sedimentary and magmatic origin, using polished thin sections. Internal structures of grossular from Mexico and pyrope from the Italian Alps were characterized and visualized by CL-microscopy. The different growth domains were additionally studied using CL-spectra and electron microprobe (EMP) analysis. Grossular shows a patchy zonation in its core while in mantle and rim zones oscillatory zoning is observed. It contains zones of anomalous birefringence, zones of orange and bluish luminescence and zones lacking luminescence. Different but low amounts of the activator elements Mn2+ and Eu2+ are responsible for the orange and bluish luminescent domains. Pyrope is also characterized by oscillatory growth zones, shows a dull luminescent core with a change of crystal morphology during growth, and displays an increase of brightness from core towards rim—the outermost rim, however, is lacking luminescence. The different luminescent zones are characterized by different amounts of Dy3+, Tb3+, Sm3+ and Sm2+ as activator elements. Because of slow diffusion rates of activators such as the REEs Sm, Dy and Tb, it can be still possible to visualize possible prograde and/or peak pressure stage growth domains of garnet, even if later high temperature events may have homogenized the major element profiles. Such domains may help to identify respective assemblages of mineral inclusions, and hence these results can represent an integral part of a detailed PT path. Thus the CL-information can be used as an important pathfinder prior to supplementary investigations, as for instance EMP, ion probe, mineral or fluid inclusion studies.

A fully automated image analysis method to quantify lung fibrosis in the bleomycin-induced rat model.

Intratracheal administration of bleomycin induces fibrosis in the lung, which is mainly assessed by histopathological grading that is subjective. Current literature highlights the need of reproducible and quantitative pulmonary fibrosis analysis. If some quantitative studies looked at fibrosis parameters separately, none of them quantitatively assessed both aspects: lung tissue remodeling and collagenization. To ensure reliable quantification, support vector machine learning was used on digitalized images to design a fully automated method that analyzes two important aspects of lung fibrosis: (i) areas having substantial tissue remodeling with appearance of dense fibrotic masses and (ii) collagen deposition. Fibrotic masses were identified on low magnification images and collagen detection was performed at high magnification. To insure a fully automated application the tissue classifier was trained on several independent studies that were performed over a period of four years. The detection method generates two different values that can be used to quantify lung fibrosis development: (i) percent area of fibrotic masses and (ii) percent of alveolar collagen. These two parameters were validated using independent studies from bleomycin- and saline-treated animals. A significant change of these lung fibrosis quantification parameters- increased amount of fibrotic masses and increased collagen deposition- were observed upon intratracheal administration of bleomycin and subsequent significant beneficial treatments effects were observed with BIBF-1120 and pirfenidone.

Failure Analysis of Heavy Ion-Irradiated Schottky Diodes

In this paper, we use high- and low-magnification optical microscope images, thermal infrared camera images, and scanning electron microscope images to identify failure locations in heavy ion-irradiated Schottky diodes. After failures have been identified, the parts were cross sectioned to describe the structure of the failures and energy dispersive X-ray spectroscopy was used to characterize the materials in the failure structures.

The integration of magnifications: A novel approach to obtain representative information about the pore space of mudrocks from SEM images

Abstract High resolution Scanning Electron Microscopy (SEM) imaging has been widely used to characterize the pore space of mudrocks. However, obtaining representative microfabric information has been limited by the highly heterogeneous microstructure of mudrocks, which contains pores and particles spanning several length scales. The integration of magnifications (IOM) method was developed to obtain representative information about the pore space of mudrocks. This method combines information from SEM images acquired at different magnifications to characterize the pore space. High magnification images with high resolution are used to quantify small pores and low magnification images with lower resolution are used to quantify large pores. These different scales are then mathematically combined to obtain a complete pore space representation. The method was validated using 2D Monte Carlo simulations applied on a synthetic 2D microstructure with a prescribed porosity and pore size distribution. The applicability of the method was illustrated by computing the porosity and pore size distribution of two naturally resedimented mudrocks. The IOM method provides a practical and economical alternative to the current cumbersome methodology that stitches together hundreds to thousands of SEM images to obtain large representative mosaics.

Time-lapsed, large-volume, high-resolution intravital imaging for tissue-wide analysis of single cell dynamics.

Pathologists rely on microscopy to diagnose disease states in tissues and organs. They utilize both high-resolution, high-magnification images to interpret the staining and morphology of individual cells, as well as low-magnification overviews to give context and location to these cells. Intravital imaging is a powerful technique for studying cells and tissues in their native, live environment and can yield sub-cellular resolution images similar to those used by pathologists. However, technical limitations prevent the straightforward acquisition of low-magnification images during intravital imaging, and they are hence not typically captured. The serial acquisition, mosaicking, and stitching together of many high-resolution, high-magnification fields of view is a technique that overcomes these limitations in fixed and ex vivo tissues. The technique however, has not to date been widely applied to intravital imaging as movements caused by the living animal induce image distortions that are difficult to compensate for computationally. To address this, we have developed techniques for the stabilization of numerous tissues, including extremely compliant tissues, that have traditionally been extremely difficult to image. We present a novel combination of these stabilization techniques with mosaicked and stitched intravital imaging, resulting in a process we call Large-Volume High-Resolution Intravital Imaging (LVHR-IVI). The techniques we present are validated and make large volume intravital imaging accessible to any lab with a multiphoton microscope.

A low cost mobile phone dark-field microscope for nanoparticle-based quantitative studies

Dark-field microscope (DFM) analysis of nanoparticle binding signal is highly useful for a variety of research and biomedical applications, but current applications for nanoparticle quantification rely on expensive DFM systems. The cost, size, limited robustness of these DFMs limits their utility for non-laboratory settings. Most nanoparticle analyses use high-magnification DFM images, which are labor intensive to acquire and subject to operator bias. Low-magnification DFM image capture is faster, but is subject to background from surface artifacts and debris, although image processing can partially compensate for background signal. We thus mated an LED light source, a dark-field condenser and a 20× objective lens with a mobile phone camera to create an inexpensive, portable and robust DFM system suitable for use in non-laboratory conditions. This proof-of-concept mobile DFM device weighs less than 400g and costs less than $2000, but analysis of images captured with this device reveal similar nanoparticle quantitation results to those acquired with a much larger and more expensive desktop DFMM system. Our results suggest that similar devices may be useful for quantification of stable, nanoparticle-based activity and quantitation assays in resource-limited areas where conventional assay approaches are not practical.

Corrigendum] Death receptor 5 expression is inversely correlated with prostate cancer progression.

During the preparation of the figures in the above article, the authors inadvertently duplicated in Fig.1B, a and b (high and low magnification images) the images that had already appeared as Figs.5A,a andc (high and low magnification images), respectively, of the following paper: Huerta-Yepez S, BaritakiS, Baay-GuzmanG, Hernandez-LunaMA, Hernandez-CuetoA, VegaMI and BonavidaB: Contribution of either YY1 or BclXL-induced inhibition by the NO-donor DETANONOate in the reversal of drug resistance, both invitro and invivo. Nitric Oxide29: 17-24, 2013. The revised version of Fig.1 containing the corrected data for Fig.1B,a andb (high and low magnification images; the YY1 data) is shown opposite protein expression. All those authors whom the corresponding author was able to contact have agreed to this Corrigendum. The authors regret this error, and apologize for any confusion that it may have caused. [the original article was published in the Molecular Medicine Reports 10: 2279-2286, 2014; DOI: 10.3892/mmr.2014.2504 ].

Novel low-dose imaging technique for characterizing atomic structures through scanning transmission electron microscope

To investigate dislocations or heterostructures across interfaces is now of great interest to condensed matter and materials scientists. With the advances in aberration-corrected electron optics, the scanning transmission electron microscope has demonstrated its excellent capability of characterizing atomic structures within nanomaterials, and well-resolved atomic-resolution images can be obtained through long-exposure data acquisition. However, the sample drifting, carbon contamination, and radiation damage hinder further analysis, such as deriving three-dimensional (3D) structures from a series of images. In this study, a method for obtaining atomic-resolution images with significantly reduced exposure time was developed, using which an original high-resolution image with approximately one tenth the electron dose can be obtained by combining a fast-scan high-magnification image and a slow-scan low-magnification image. The feasibility of obtaining 3D atomic structures using the proposed approach was demonstrated through multislice simulation. Finally, the feasibility and accuracy of image restoration were experimentally verified. This general method cannot only apply to electron microscopy but also benefit to image radiation-sensitive materials using various light sources.

Pluri-IQ: Quantification of Embryonic Stem Cell Pluripotency through anImage-Based Analysis Software.

SummaryImage-based assays, such as alkaline phosphatase staining or immunocytochemistry for pluripotent markers, are common methods used in the stem cell field to assess pluripotency. Although an increased number of image-analysis approaches have been described, there is still a lack of software availability to automatically quantify pluripotency in large images after pluripotency staining. To address this need, we developed a robust and rapid image processing software, Pluri-IQ, which allows the automatic evaluation of pluripotency in large low-magnification images. Using mouse embryonic stem cells (mESC) as a model, we combined an automated segmentation algorithm with a supervised machine-learning platform to classify colonies as pluripotent, mixed, or differentiated. In addition, Pluri-IQ allows the automatic comparison between different culture conditions. This efficient user-friendly open-source software can be easily implemented in images derived from pluripotent cells or cells that express pluripotent markers (e.g., OCT4-GFP) and can be routinely used, decreasing image assessment bias.

Abstract 879: Multi-scale time-lapse intravital imaging of soft tissues to map single cell behavior

Abstract After more than 10 years of research into the tumor micro-environment and the sources of tumor micro-heterogeneity, it is becoming increasingly clear that in addition to driver mutations, the tumor microenvironment determines tumor metastatic phenotype. This research has led to a new understanding of the impact of tumor microenvironments and their heterogeneity upon tumor cell proliferation, dissemination, dormancy, and survival. A full understanding of this heterogeneity, both temporally and spatially, how it supports tumor cell dissemination, dormancy and eventual further metastatic growth, and how it responds to therapeutic interventions, is crucial. Traditional single parameter studies have not been as productive as hoped in revealing the above relationships and have highlighted the need to understand tumors as integrated systems of genes, gene networks, and intracellular interactions, particularly with regard to the interplay between cells and their immediate microenvironment. To accomplish this, we have developed a new technology in the form of large volume intravital imaging using multiphoton intravital microscopy (MIVM) where images of large tumor volumes are stitched together to form a comprehensive record of the genes, gene networks, and intracellular interactions that occur throughout many tumor microenvironments at single cell resolution. Here, we report a protocol for obtaining large area high-resolution mosaic imaging in living animals. The protocol is composed of surgical techniques for the stabilization of soft tissues including mammary gland, lymph node, liver, and lung and the acquisition of multiple high-magnification tiles that are stitched together to form a large-area low-magnification image. We have developed specific protocols for the surgery as well as tools for tissue stabilization and the acquisition and stitching of the images to generate very large MVIM data sets. We have interfaced these very large MVIM image data sets with support vector machine (SVM) classification, a nonlinear, multiparametric classification algorithm suitable for analysis of systems with arbitrary distributions and/or non-linear parameter’s correlations. To define the combinations of microenvironment parameters where tumor cell phenotypes of interest are likely to occur, we used these microenvironment parameters as an “input” for the SVM classifier to identify associated tumor cell phenotypes as the classifier’s “output”. Using this approach at widely varying temporal and spatial scales (from minutes to weeks and from sub-cellular to tissue wide) and at different stages (early carcinoma on to metastasis), has resulted in a catalog of associations between microenvironment conditions and tumor cell phenotypes associated with progression. These associations resulted in new insights into the mechanisms of metastasis. Citation Format: Jessica M. Pastoriza, Maria Soledad Sosa, Kathryn Harper, Julio Aguirre-Ghiso, Maja H. Oktay, David Entenberg, Yarong Wang, John S. Condeelis, Aviv Bergman, Mihaela Skobe, Benedicte Lenoir. Multi-scale time-lapse intravital imaging of soft tissues to map single cell behavior [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 879. doi:10.1158/1538-7445.AM2017-879

Determining the structure of graphene-based flakes from their morphotype

Graphene-based flakes of various origins were investigated by transmission electron microscopy. For each of them, the morphotype was described, and then both the related structure (stacking type) and nanotexture (degree of perfection of the graphene layers) were described as well. Two kinds of clearly discriminated morphotypes were found (‘crumpled’ and ‘folded’), for objects with similar overall dimensions (in the range of several micrometers large) and similar multi-graphene nature (with the number of stacked graphene layers in the range of 2–15, sometimes more). It is found that the morphological discrepancies relate to the structural discrepancies. Typically, whether the structure is either turbostratic or graphitic makes the flake morphotype be either crumpled or folded, as a consequence of preferred folding directions related to lattice periodicities. In this regard, structure prevails over nanotexture. This strict relationship implies that the 2D (turbostratic) or 3D (graphitic) nature of the stacking in graphene-based flakes whose determination usually requires diffraction studies may be merely deduced from low magnification images of their morphotypes.