Geometry Processing of Conventionally Produced Mouse Brain Slice Images

Brain mapping research in most neuroanatomical laboratories relies on conventional processing techniques, which often introduce histological artifacts such as tissue tears and tissue loss. In this paper we present techniques and algorithms for automatic registration and 3D reconstruction of conventionally produced mouse brain slices in a standardized atlas space. This is achieved first by constructing a virtual 3D mouse brain model from annotated slices of Allen Reference Atlas (ARA). Virtual re-slicing of the reconstructed model generates ARA-based slice images corresponding to the microscopic images of histological brain sections. These image pairs are aligned using a geometric approach through contour images. Histological artifacts in the microscopic images are detected and removed using Constrained Delaunay Triangulation before performing global alignment. Finally, non-linear registration is performed by solving Laplace's equation with Dirichlet boundary conditions. Our methods provide significant improvements over previously reported registration techniques for the tested slices in 3D space, especially on slices with significant histological artifacts. Further, as an application we count the number of neurons in various anatomical regions using a dataset of 51 microscopic slices from a single mouse brain. This work represents a significant contribution to this subfield of neuroscience as it provides tools to neuroanatomist for analyzing and processing histological data.Comment: 14 pages, 11 figure

Comparison of image analysis software packages in the assessment of adhesion of microorganisms to mucosal epithelium using confocal laser scanning microscopy

We have compared current image analysis software packages in order to find the most useful one for assessing microbial adhesion and inhibition of adhesion to tissue sections. We have used organisms of different sizes, the bacterium Helicobacter pylori and the yeast Candida albicans. Adhesion of FITC-labelled H. pylori and C. albicans was assessed by confocal microscopy. Four different Image analysis software packages, NIH-Image, IP Lab, Image Pro+, and Metamorph, were compared for their ability to quantify adhesion of the two organisms and several quantification methods were devised for each package. For both organisms, the dynamic range that could be detected by the software packages was 1×106?1×109 cells/ml. Of the four software packages tested, our results showed that Metamorph software, using our ?Region of Interest? method, with the software's ?Standard Area Method? of counting, was the most suitable for quantifying adhesion of both organisms because of its unique ability to separate clumps of microbial cells. Moreover, fewer steps were required. By pre-incubating H. pylori with the glycoconjugate Lewis b-HSA, an inhibition of binding of 48.8% was achieved using 250 ?g/ml Lewis b-HSA. The method we have devised using Metamorph software, provides a simple, quick and accurate way of quantifying adhesion and inhibition of adhesion of microbial cells to the epithelial surface of tissue sections. The method can be applied to organisms ranging in size from small bacteria to larger yeast cells

Colocalization of neurons in optical coherence microscopy and Nissl-stained histology in Brodmann’s area 32 and area 21

Published in final edited form as: Brain Struct Funct. 2019 January ; 224(1): 351–362. doi:10.1007/s00429-018-1777-z.Optical coherence tomography is an optical technique that uses backscattered light to highlight intrinsic structure, and when applied to brain tissue, it can resolve cortical layers and fiber bundles. Optical coherence microscopy (OCM) is higher resolution (i.e., 1.25 µm) and is capable of detecting neurons. In a previous report, we compared the correspondence of OCM acquired imaging of neurons with traditional Nissl stained histology in entorhinal cortex layer II. In the current method-oriented study, we aimed to determine the colocalization success rate between OCM and Nissl in other brain cortical areas with different laminar arrangements and cell packing density. We focused on two additional cortical areas: medial prefrontal, pre-genual Brodmann area (BA) 32 and lateral temporal BA 21. We present the data as colocalization matrices and as quantitative percentages. The overall average colocalization in OCM compared to Nissl was 67% for BA 32 (47% for Nissl colocalization) and 60% for BA 21 (52% for Nissl colocalization), but with a large variability across cases and layers. One source of variability and confounds could be ascribed to an obscuring effect from large and dense intracortical fiber bundles. Other technical challenges, including obstacles inherent to human brain tissue, are discussed. Despite limitations, OCM is a promising semi-high throughput tool for demonstrating detail at the neuronal level, and, with further development, has distinct potential for the automatic acquisition of large databases as are required for the human brain.Accepted manuscrip

Evaluation of Dynamic Cell Processes and Behavior Using Video Bioinformatics Tools

Just as body language can reveal a person’s state of well-being, dynamic changes in cell behavior and morphology can be used to monitor processes in cultured cells. This chapter discusses how CL-Quant software, a commercially available video bioinformatics tool, can be used to extract quantitative data on: (1) growth/proliferation, (2) cell and colony migration, (3) reactive oxygen species (ROS) production, and (4) neural differentiation. Protocols created using CL-Quant were used to analyze both single cells and colonies. Time-lapse experiments in which different cell types were subjected to various chemical exposures were done using Nikon BioStations. Proliferation rate was measured in human embryonic stem cell colonies by quantifying colony area (pixels) and in single cells by measuring confluency (pixels). Colony and single cell migration were studied by measuring total displacement (distance between the starting and ending points) and total distance traveled by the colonies/cells. To quantify ROS production, cells were pre-loaded with MitoSOX Red™, a mitochondrial ROS (superoxide) indicator, treated with various chemicals, then total intensity of the red fluorescence was measured in each frame. Lastly, neural stem cells were incubated in differentiation medium for 12 days, and time lapse images were collected daily. Differentiation of neural stem cells was quantified using a protocol that detects young neurons. CLQuant software can be used to evaluate biological processes in living cells, and the protocols developed in this project can be applied to basic research and toxicological studies, or to monitor quality control in culture facilities

A data extraction system for underwater particle holography

Pulsed laser holography is an extremely powerful technique for the study of particle fields as it allows instantaneous, noninvasive high-resolution recording of substantial volumes. By replaying the real image one can obtain the size, shape, position and - if multiple exposures are made - velocity of every object in the recorded field. Manual analysis of large volumes containing thousands of particles is, however, an enormous and time-consuming task, with operator fatigue an unpredictable source of errors. Clearly the value of holographic measurements also depends crucially on the quality of the reconstructed image: not only will poor resolution degrade size and shape measurements, but aberrations such as coma and astigmatism can change the perceived centroid of a particle, affecting position and velocity measurements. For large-scale applications of particle field holography, specifically the in situ recording of marine plankton with 'HoloCam,' we have developed an automated data extraction system that can be readily switched between the in-line and off-axis geometries and provides optimised reconstruction from holograms recorded underwater. As a videocamera is automatically stepped through the 200 by 200 by 1000mm sample volume, image processing and object tracking routines locate and extract particle images for further classification by a separate software module

Cross-Section Bead Image Prediction in Laser Keyhole Welding of AISI 1020 Steel Using Deep Learning Architectures

A deep learning model was applied for predicting a cross-sectional bead image from laser welding process parameters. The proposed model consists of two successive generators. The first generator produces a weld bead segmentation map from laser intensity and interaction time, which is subsequently translated into an optical microscopic (OM) image by the second generator. Both generators exhibit an encoder & x2013;decoder structure based on a convolutional neural network (CNN). In the second generator, a conditional generative adversarial network (cGAN) was additionally employed with multiscale discriminators and residual blocks, considering the size of the OM image. For a training dataset, laser welding experiments with AISI 1020 steel were conducted on a large process window using a 2 KW fiber laser, and a total of 39 process conditions were used for the training. High-resolution OM images were successfully generated, and the predicted bead shapes were reasonably accurate (R-Squared: 89.0 & x0025; for penetration depth, 93.6 & x0025; for weld bead area)

A holographic system for subsea recording and analysis of plankton and other marine particles

We report here details of the design, development, initial testing and field-deployment of the HOLOMAR system for in-situ subsea holography and analysis of marine plankton and nonliving particles. HOLOMAR comprises a submersible holographic camera ("HoloCam") able to record in-line and off-axis holograms at depths down to 100 m, together with specialised reconstruction hardware ("HoloScan") linked to custom image processing and classification software. The HoloCam consists of a laser and power supply, holographic recording optics and holographic plate holders, a water-tight housing and a support frame. It utilises two basic holographic geometries, in-line and off-axis such that a wide range of species, sizes and concentrations can be recorded. After holograms have been recorded and processed they are reconstructed in full three-dimensional detail in air in a dedicated replay facility. A computer-controlled microscope, using video cameras to record the image at a given depth, is used to digitise the scene. Specially written software extracts a binarised image of an object in its true focal plane and is classified using a neural network. The HoloCam was deployed on two separate cruises in a Scottish sea loch (Loch Etive) to a depth of 100 m and over 300 holograms were recorded