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"Photomicrography"
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Mosquito bite
As children play hide-and-seek on a hot summer night, a mosquito, Culex, hides and seeks as well.
Book
A Scalable and Modular Dome Illumination System for Scientific Microphotography on a Budget
A scalable and modular LED illumination dome for microscopic scientific photography is described and illustrated, and methods for constructing such a dome are detailed. Dome illumination for insect specimens has become standard practice across the field of insect systematics, but many dome designs remain expensive and inflexible with respect to new LED technology. Further, a one-size-fits-all dome cannot accommodate the large breadth of insect size encountered in nature, forcing the photographer to adapt, in some cases, to a less than ideal dome design. The dome described here is scalable, as it is based on a isodecahedron, and the template for the dome is available as a downloaded file from the internet that can be printed on any printer, on the photographer's choice of media. As a result, a photographer can afford, using this design, to produce a series of domes of various sizes and materials, and LED ring lights of various sizes and color temperatures, depending on the need.
Journal Article
Microscopic Image Photography Techniques of the Past, Present, and Future
The field of pathology is driven by microscopic images. Educational activities for trainees and practicing pathologists alike are conducted through exposure to images of a variety of pathologic entities in textbooks, publications, online tutorials, national and international conferences, and interdepartmental conferences. During the past century and a half, photographic technology has progressed from primitive and bulky, glass-lantern projector slides to static and/or whole slide digital-image formats that can now be transferred around the world in a matter of moments via the Internet.
To provide a historic and technologic overview of the evolution of microscopic-image photographic tools and techniques.
Primary historic methods of microscopic image capture were delineated through interviews conducted with senior staff members in the Emory University Department of Pathology. Searches for the historic image-capturing methods were conducted using the Google search engine. Google Scholar and PubMed databases were used to research methods of digital photography, whole slide scanning, and smart phone cameras for microscopic image capture in a pathology practice setting.
Although film-based cameras dominated for much of the time, the rise of digital cameras outside of pathology generated a shift toward digital-image capturing methods, including mounted digital cameras and whole slide digital-slide scanning. Digital image capture techniques have ushered in new applications for slide sharing and second-opinion consultations of unusual or difficult cases in pathology. With their recent surge in popularity, we suspect that smart phone cameras are poised to become a widespread, cost-effective method for pathology image acquisition.
Journal Article
Testing an Alternative Method for Estimating the Length of Fungal Hyphae Using Photomicrography and Image Processing
This study aimed to develop and test an unbiased and rapid methodology to estimate the length of external arbuscular mycorrhizal fungal (AMF) hyphae in soil. The traditional visual gridline intersection (VGI) method, which consists in a direct visual examination of the intersections of hyphae with gridlines on a microscope eyepiece after aqueous extraction, membrane-filtration, and staining (e.g., with trypan blue), was refined. For this, (i) images of the stained hyphae were taken by using a digital photomicrography technique to avoid the use of the microscope and the method was referred to as \"digital gridline intersection\" (DGI) method; and (ii), the images taken in (i) were processed and the hyphal length was measured by using ImageJ software, referred to as the \"photomicrography-ImageJ processing\" (PIP) method. The DGI and PIP methods were tested using known grade lengths of possum fur. Then they were applied to measure the hyphal lengths in soils with contrasting phosphorus (P) fertility status. Linear regressions were obtained between the known lengths (Lknown) of possum fur and the values determined by using either the DGI (LDGI) (LDGI = 0.37 + 0.97 × Lknown, r2 = 0.86) or PIP (LPIP) methods (LPIP = 0.33 + 1.01 × Lknown, r2 = 0.98). There were no significant (P > 0.05) differences between the LDGI and LPIP values. While both methods provided accurate estimation (slope of regression being 1.0), the PIP method was more precise, as reflected by a higher value of r2 and lower coefficients of variation. The average hyphal lengths (6.5-19.4 m g-1) obtained by the use of these methods were in the range of those typically reported in the literature (3-30 m g-1). Roots growing in P-deficient soil developed 2.5 times as many hyphae as roots growing in P-rich soil (17.4 vs 7.2 m g-1). These tests confirmed that the use of digital photomicrography in conjunction with either the grid-line intersection principle or image processing is a suitable method for the measurement of AMF hyphal lengths in soils for comparative investigations.
Journal Article
Evaluation of the safety of time-lapse observations for human embryos
Purpose
To assess the effects of light from an integrated optical microscope and evaluate the safety of time-lapse observations using a built-in microscope incubator.
Methods
We prospectively compared the fertilization rate and embryonic morphology after intracytoplasmic sperm injection between embryos cultured with time-lapse observations every 15 min in an incubator with an integrated optical microscope and embryos with intermittent observations (once a day) in conventional incubators.
Results
No significant differences were observed in the fertilization rate (57.5% vs. 57.5%) or the rate of excellent-good cleavage embryos (36.0% vs. 36.0%).
Conclusions
These results suggest that time-lapse observations using an incubator with an integrated optical microscope may therefore be safely utilized in clinical practice.
Journal Article
Guide to light-sheet microscopy for adventurous biologists
Ten years of development in light-sheet microscopy have led to spectacular demonstrations of its capabilities. The technology is ready to assist biologists in tackling scientific problems, but are biologists ready for it? Here we discuss the interdisciplinary challenges light-sheet microscopy presents for biologists and highlight available resources.
Journal Article
Optimized localization analysis for single-molecule tracking and super-resolution microscopy
A theoretical and experimental treatment of fitting methods for localizing the centers of diffraction-limited spots is presented. Use of an analytical point spread function shows that maximum likelihood fitting is superior to both unweighted and weighted least squares Gaussian fitting.
We optimally localized isolated fluorescent beads and molecules imaged as diffraction-limited spots, determined the orientation of molecules and present reliable formulas for the precision of various localization methods. Both theory and experimental data showed that unweighted least-squares fitting of a Gaussian squanders one-third of the available information, a popular formula for its precision exaggerates beyond Fisher's information limit, and weighted least-squares may do worse, whereas maximum-likelihood fitting is practically optimal.
Journal Article
Optogenetic self-stimulation in the nucleus accumbens: D1 reward versus D2 ambivalence
The nucleus accumbens (NAc) contains multiple subpopulations of medium spiny neurons (MSNs). One subpopulation expresses D1-type dopamine receptors, another expresses D2-type receptors, and a third expresses both. The relative roles in NAc of D1 neurons versus D2 neurons in appetitive motivation were assessed here. Specifically, we asked whether D1-Cre mice would instrumentally seek optogenetic self-stimulation specifically targeted at D1 MSNs in NAc, and similarly if D2-Cre mice would self-stimulate D2 neurons in NAc. Mice were implanted with Cre-targeted channelrhodopsin (ChR2) virus and optic fibers in NAc. Subsequently, mice could earn brief NAc laser illuminations by actively touching a metal spout in one task, or by going to a particular location in a separate task. Results indicated that D1 neuronal excitation in NAc supported intense self-stimulation in both tasks. D1-Cre mice earned hundreds to thousands of spout-touches per half-hour session, and also sought out locations that delivered NAc laser to excite D1 MSNs. By comparison, D2 ChR2 mice showed lower but still positive levels of self-stimulation in the spout-touch task, earning dozens to hundreds of NAc laser illuminations. However, in the location task, D2 mice failed to show positive self-stimulation. If anything, a few D2 individuals gradually avoided the laser location. Brain-wide measures indicated that D1 and D2 stimulations in NAc recruited heavily overlapping patterns of Fos activation in distant limbic structures. These results confirm that excitation of D1 MSNs in NAc supports strong incentive motivation to self-stimulate. They also suggest that excitation of D2 neurons in NAc supports self-stimulation under some conditions, but fails under others and possibly may even shift to negative avoidance.
Journal Article
Deep learning for Alzheimer's disease: Mapping large-scale histological tau protein for neuroimaging biomarker validation
Abnormal tau inclusions are hallmarks of Alzheimer's disease and predictors of clinical decline. Several tau PET tracers are available for neurodegenerative disease research, opening avenues for molecular diagnosis in vivo. However, few have been approved for clinical use. Understanding the neurobiological basis of PET signal validation remains problematic because it requires a large-scale, voxel-to-voxel correlation between PET and (immuno) histological signals. Large dimensionality of whole human brains, tissue deformation impacting co-registration, and computing requirements to process terabytes of information preclude proper validation. We developed a computational pipeline to identify and segment particles of interest in billion-pixel digital pathology images to generate quantitative, 3D density maps. The proposed convolutional neural network for immunohistochemistry samples, IHCNet, is at the pipeline's core. We have successfully processed and immunostained over 500 slides from two whole human brains with three phospho-tau antibodies (AT100, AT8, and MC1), spanning several terabytes of images. Our artificial neural network estimated tau inclusion from brain images, which performs with ROC AUC of 0.87, 0.85, and 0.91 for AT100, AT8, and MC1, respectively. Introspection studies further assessed the ability of our trained model to learn tau-related features. We present an end-to-end pipeline to create terabytes-large 3D tau inclusion density maps co-registered to MRI as a means to facilitate validation of PET tracers.
Journal Article