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This protocol focuses on measuring the microscope's lateral and axial resolution, essential for reporting size measurements of near-resolution limit objects or distances between them. Here resolution is expressed as the Full Width at Half Maximum (FWHM) of a measured Point Spread Function (PSF) of sub-resolution size beads. PSF is highly related to objective quality and condition, but also depends strongly upon other parameters ranging from sample preparation to signal detection. Monitoring PSF over time will identify possible aberrations in the system (e.g., damaged, unclean objective, defective or not adapted oil, etc.). We define test sample preparation, image acquisition, and data analysis protocols for point scanning and spinning disk confocal microscopes.

Advanced in vitro tissue chip models can reduce and replace animal experimentation and may eventually support ‘on-chip’ clinical trials. To realize this potential, however, tissue chip platforms must be both mass-produced and reconfigurable to allow for customized design. To address these unmet needs, we introduce an extension of our µSiM (microdevice featuring a silicon-nitride membrane) platform. The modular µSiM (m-µSiM) uses mass-produced components to enable rapid assembly and reconfiguration by laboratories without knowledge of microfabrication. We demonstrate the utility of the m-µSiM by establishing an hiPSC-derived blood-brain barrier (BBB) in bioengineering and non-engineering, brain barriers focused laboratories. We develop and validate in situ and sampling-based assays of small molecule diffusion as a measure of barrier function. BBB properties show excellent interlaboratory agreement and match expectations from literature, validating the m-µSiM as a platform for barrier models and demonstrating successful dissemination of components and protocols. We then demonstrate the ability to quickly reconfigure the m-µSiM for co-culture and immune cell transmigration studies through addition of accessories and/or quick exchange of components. Because the development of modified components and accessories is easily achieved, custom designs of the m-µSiM should be accessible to any laboratory desiring a barrier-style tissue chip platform.

We present a novel deep learning approach to reconstruct confocal microscopy stacks from single light field images. To perform the reconstruction, we introduce LFMNet, a fully convolutional neural network architecture inspired by the U-Net design. A property of LFMNet is its ability to reconstruct with near confocal accuracy a $112 \times 112 \times 57.6 \,\mu {\rm m}^3$ volume ( $1287\times 1287\times 64$ voxels) in $\text{50}\,ms$ given a single light field image of $1287\times 1287$ pixels, thus dramatically reducing 720-fold the capture time compared to confocal scanning of assays at the same volumetric resolution and 64-fold the required storage. To prove its applicability in life sciences, our approach is evaluated both quantitatively and qualitatively on mouse brain slices with fluorescently labeled blood vessels. Because of the drastic reduction in image acquisition time, our setup and method are suitable for imaging highly dynamic and light-sensitive events, and in closed-loop systems, where latency due to the reconstruction time is crucial. We provide empirical analysis of the optical design, of the network architecture and of our training procedure to reconstruct confocal-like volumes for a given target depth range. To train our network, we built a data set of 362 light field images of the mouse brain sample and the corresponding aligned set of 3D confocal scans, which we use as ground truth. The data set will be made available for research purposes.

In April 2020, the QUality Assessment and REProducibility for Instruments and Images in Light Microscopy (QUAREP-LiMi) initiative was formed. This initiative comprises imaging scientists from academia and industry who share a common interest in achieving a better understanding of the performance and limitations of microscopes and improved quality control (QC) in light microscopy. The ultimate goal of the QUAREP-LiMi initiative is to establish a set of common QC standards, guidelines, metadata models, and tools, including detailed protocols, with the ultimate aim of improving reproducible advances in scientific research. This White Paper 1) summarizes the major obstacles identified in the field that motivated the launch of the QUAREP-LiMi initiative; 2) identifies the urgent need to address these obstacles in a grassroots manner, through a community of stakeholders including, researchers, imaging scientists, bioimage analysts, bioimage informatics developers, corporate partners, funding agencies, standards organizations, scientific publishers, and observers of such; 3) outlines the current actions of the QUAREP-LiMi initiative, and 4) proposes future steps that can be taken to improve the dissemination and acceptance of the proposed guidelines to manage QC. To summarize, the principal goal of the QUAREP-LiMi initiative is to improve the overall quality and reproducibility of light microscope image data by introducing broadly accepted standard practices and accurately captured image data metrics., Comment: 17 pages, 3 figures, shortened abstract, Co-Lead Authors: Glyn Nelson and Ulrike Boehm, Corresponding author: Roland Nitschke

Networks of dry valleys (or balkas) and hollows in the upper reaches of fluvial basins in extraglacial areas in the Penultimate Glaciation (Marine Isotope Stage 6—MIS 6) regions of the East European Plain demonstrate clear incision/aggradation rhythms corresponding to global glacial/interglacial climate cycles. The first phase of each incision/aggradation rhythm began after the global glacial maximum and was characterized by a cool and humid climate, permafrost and sparse vegetation, when high surface runoff and active linear erosion formed a dense network of gullies. The second phase occurred at the glacial–interglacial transition and the subsequent interglacial period with its warm and humid climate and dense vegetation. This phase was distinguished by the partial filling of fluvial forms with slopewash deposits, the transformation of gullies into dry valleys (balkas) and the subsequent stabilization of fluvial forms marked by the formation of mature soils on the sides and bottoms of balkas. The third phase of the rapid accumulation of balkas developed during the cold and dry part of the next glacial epoch, resulting in the balkas becoming shallow hollows filled in with sediments. The last full incision/aggradation rhythm occurred in the late MIS 6 to mid-MIS 2. The erosion network formed during the late MIS 6 was almost completely filled by mid-MIS 2, and its manifestation in the modern topography is limited to a network of shallow hollows in the upper parts of the fluvial systems. The modern (incomplete) incision/aggradation rhythm began in the late MIS 2 and caused the formation of the modern erosion landscape in the upper reaches of fluvial systems. This rhythm is now in the stabilization phase, and the main accumulation phase of this rhythm is still far in the future.

In this study we analyzed the information about the presence of different types of anthropogenic objects (settlements, transport infrastructure, mining areas, etc.) in the Arctic zone of Russia. This information was taken from open Internet-sources: maps, cartographic projects, databases, schemes of regional development of the Russian Federation. Data analysis shows than only about 20% of Russian Arctic’s area is affected by economic development, meanwhile on the other 80% of the area there are practically no anthropogenic objects.The economic development of the Arctic region decreases from West to East of Russia. The Republic of Karelia is characterized by the highest economic development level (only 13,1% of the area are not affected by any economic activities), the lowest levels have Krasnoyarskiy krai (95,2%) and the Republic of Sakha (Yakutia) (87,2%). Data on the presence, position, and types of anthropogenic objects were subjected to the k-means method of cluster analysis in order to identify characteristic combinations of objects corresponding to different types of development. Within the Arctic zone of Russia six main types of economical use of the territory were identified. Each of these types was characterized by the dominance of a certain type of anthropogenic objects (settlements, roads, mining industry objects, oil and gas transport infrastructure, wood industry objects).Each type of the economical use of the territory is characterized by specific anthropogenic transformation of the topography of the area. The greatest transformation of the topography and geomorphological processes was found within the open mining areas. The least influence on the topography is connected with some of the linear transport structures (unpaved roads and underground gas pipelines). In general, economic activity in Russian Arctic is relatively low. Anthropogenic transformation of topography and geomorphic processes ​​is typical for the area about 667 thousand square km, that is about 18% of the total area of ​​the Russian Arctic.This study is supported by Russian Foundation for Basic Research (RFBR) Project № 18-05-60200 "Anthropogenic transformation of Arctic Landscapes for the last 100 years".

The paper describes the methodology and presents the results of the calculation of the spectral characteristics of the terrain for the territory of the Bolshezemelskaya and Malozemelskaya Tundra (the north of the European part of Russia). There are nine terrain clusters, differing in the topographic dissection pattern. Their geomorphological interpretation is given. Three clusters characterize mountainous and coarsely hilly terrain with the depth of dissection from 150 m and more. The others are low and elevated terrains with much lower depths of dissection (up to 50 m) and different patterns of interposition of landforms.

Histochemistry and Cell Biology, 128 (6), ISSN:0948-6143, ISSN:1432-119X

Of the various super-resolution techniques, stimulated emission depletion (STED) microscopy achieves the best temporal resolution at high spatial resolution, enabling live-cell imaging beyond the diffraction limit. However, STED and most other super-resolution imaging methods utilize a particular type of information extractable from the raw data, namely the positions of fluorophores. To expand on the use of super-resolution techniques, we report here the live-cell STED microscopy of a dynamic biosensor. Using the fluorescent H2O2 sensor HyPer2 for subdiffraction imaging, we were able not only to image filaments with superior resolution by localizing emission but also to trace H2O2 produced within living cell by monitoring brightness of the probe. STED microscopy of HyPer2 demonstrates potential utility of FP-based biosensors for super-resolution experiments in situ and in vivo.