scholarly journals THE LOCALIZATION OF CATECHOLAMINE FLUORESCENCE TO DOG HYPOTHALAMIC NEUROMELANIN-BEARING NEURONS

1973 ◽  
Vol 21 (2) ◽  
pp. 175-183 ◽  
Author(s):  
HERBERT BARDEN ◽  
ROBERT BARRETT
Keyword(s):  
Optical Properties ◽  
Fluorescence Microscopy ◽  
Ultraviolet Light ◽  
Dark Field ◽  
Fluorescence Method ◽  
Green Fluorescence ◽  
Bright Field ◽  
Neuronal Perikaryon ◽  
Histochemical Methods ◽  
Catecholamine Fluorescence

By use of histochemical methods, a survey has been made of the localization of catecholamine to neuronal perikarya and processes in relation to neuromelanin accumulation in neuronal perikarya in the hypothalamus of dogs of various ages. Catecholamine was visualized in sections as a green fluorescence by means of the formaldehyde gasinduced fluorescence method of Falck and Hillarp. Neuromelanin, if also present in these sections, was identified on the basis of its characteristic optical properties as visualized through bright field, dark field and fluorescence microscopy. The study demonstrated a sequential replacement of temporally diminishing catecholamine fluorescence by accumulating neuromelanin within the same neuronal perikaryon wherein they were found to overlap during part of the 1st year of life. Catecholamine in processes in the perifornical region of the hypothalamus, where the neuromelanin-accumulating neurons were situated, was also diminished with age. If neuromelanin was extensively exposed to ultraviolet light, as required for viewing and for photography, the initially nonfluorescent pigment was converted to one which appeared fluorescent yellow. The larger and older accumulations of neuromelanin were more resistant to this induced alteration than the smaller and younger accumulations. These results support the concept that catecholamine contributes to the formation of neuromelanin although they do not exclude the possibility that the presence of both these substances in the same perikaryon is entirely coincidental.

Use of Astronomy Filters in Fluorescence Microscopy

2012 ◽  
Vol 18 (1) ◽  
pp. 196-211
Author(s):  
Jörg Piper
Keyword(s):  
Fluorescence Microscopy ◽  
Incident Light ◽  
Optical Design ◽  
Dark Field ◽  
Light Sources ◽  
Light Path ◽  
Bright Field ◽  
Energy Excitation ◽  
Halogen Light ◽  
Background Brightness

AbstractMonochrome astronomy filters are well suited for use as excitation or suppression filters in fluorescence microscopy. Because of their particular optical design, such filters can be combined with standard halogen light sources for excitation in many fluorescent probes. In this “low energy excitation,” photobleaching (fading) or other irritations of native specimens are avoided. Photomicrographs can be taken from living motile fluorescent specimens also with a flash so that fluorescence images can be created free from indistinctness caused by movement. Special filter cubes or dichroic mirrors are not needed for our method. By use of suitable astronomy filters, fluorescence microscopy can be carried out with standard laboratory microscopes equipped with condensers for bright-field (BF) and dark-field (DF) illumination in transmitted light. In BF excitation, the background brightness can be modulated in tiny steps up to dark or black. Moreover, standard industry microscopes fitted with a vertical illuminator for examinations of opaque probes in DF or BF illumination based on incident light (wafer inspections, for instance) can also be used for excitation in epi-illumination when adequate astronomy filters are inserted as excitatory and suppression filters in the illuminating and imaging light path. In all variants, transmission bands can be modulated by transmission shift.

scholarly journals Dual-phone illumination-imaging system for high resolution and large field of view multi-modal microscopy

Lab on a Chip ◽  
2019 ◽  
Vol 19 (5) ◽  
pp. 825-836 ◽  
Author(s):  
Sara Kheireddine ◽  
Ayyappasamy Sudalaiyadum Perumal ◽  
Zachary J. Smith ◽  
Dan V. Nicolau ◽  
Sebastian Wachsmann-Hogiu
Keyword(s):  
High Resolution ◽  
Fluorescence Microscopy ◽  
Mobile Phone ◽  
Imaging System ◽  
Dark Field ◽  
Field Of View ◽  
Large Field ◽  
Bright Field

Bright-field, dark-field, Rheinberg, fluorescence microscopy on a mobile phone with phone screen illumination.

Principles and Technique of Fluorescence Microscopy

1961 ◽  
Vol s3-102 (60) ◽  
pp. 419-449
Author(s):  
M. R. YOUNG
Keyword(s):  
Fluorescence Microscopy ◽  
Focal Length ◽  
Dark Field Image ◽  
Wide Band ◽  
Dark Field ◽  
Microscopical Study ◽  
Light Sources ◽  
Bright Field ◽  
Alternative Systems ◽  
Light Filters

Interest in fluorescence microscopy has greatly increased in recent years. Technical considerations have to some extent prevented even wider application of the various fluorescence techniques now available for microscopical study of biological specimens. This paper outlines the basic requirements for optimal image quality, for the benefit of biologists and others who may not be conversant with the optical principles involved. The central problem of illumination is reviewed in some detail, and an assessment given of the two methods in current use, namely the bright-field and dark-field systems. Ratios of fluorescent to activating light received by the objective aperture, given by the two systems, have been compared, and measurements have been made of their relative light-concentrating power. Available light sources and their suitability for the excitation of fluorescence are discussed, with the problems of selecting appropriate light filters for use with the alternative systems of illumination. It is concluded that the dark-field system has decided advantages in practice and in theory for the following reasons: (1) The dark-field condenser serves as an efficient primary filter, contributing to a black background and hence good contrast. (2) The equivalent focal length is less than that of the bright-field condenser and it concentrates energy in a smaller area; this compensates in part for the loss of energy inevitably caused by the central stop. (3) It permits the use of wide-band primary filters of maximum transmission because contrast in the fluorescent image is affected only by a weak superimposed dark-field image produced in the object-plane by scattered residual activating light passed by the primary filter. With blue-light activation the visible dark-field image is effectively eliminated by means of a weak blue-absorbing secondary filter. (4) The loss of contrast due to veiling glare is minimized. A rational layout for fluorescence microscopy and methods for accurate alignment of the microscope in the vertical and horizontal positions are described. Factors influencing the choice of suitable objectives and eyepieces and some details of methods for mounting specimens are given.

Two Investigations into Grain Boundary Structure

1977 ◽  
Vol 35 ◽  
pp. 688-691
Author(s):  
P. Humble
Keyword(s):  
Grain Boundary ◽  
Dark Field ◽  
Boundary Structure ◽  
Boundary Dislocation ◽  
Bright Field ◽  
Intrinsic Structure ◽  
Weak Beam ◽  
Grain Boundary Structure ◽  
Independent Information ◽  
Diffraction Patterns

There has been sustained interest over the last few years into both the intrinsic (primary and secondary) structure of grain boundaries and the extrinsic structure e.g. the interaction of matrix dislocations with the boundary. Most of the investigations carried out by electron microscopy have involved only the use of information contained in the transmitted image (bright field, dark field, weak beam etc.). Whilst these imaging modes are appropriate to the cases of relatively coarse intrinsic or extrinsic grain boundary dislocation structures, it is apparent that in principle (and indeed in practice, e.g. (1)-(3)) the diffraction patterns from the boundary can give extra independent information about the fine scale periodic intrinsic structure of the boundary.In this paper I shall describe one investigation into each type of structure using the appropriate method of obtaining the necessary information which has been carried out recently at Tribophysics.

Imaging of platinum-shadowed macromolecular complexes in dark field

1984 ◽  
Vol 42 ◽  
pp. 146-147
Author(s):  
D.W. Andrews ◽  
F.P. Ottensmeyer
Keyword(s):  
Thin Film ◽  
Three Dimensional ◽  
Average Diameter ◽  
Dark Field ◽  
Bright Field ◽  
Field Mode ◽  
Macromolecular Complexes ◽  
Average Mass ◽  
Topological Features ◽  
Projection Images

Shadowing with heavy metals has been used for many years to enhance the topological features of biological macromolecular complexes. The three dimensional features present in directionaly shadowed specimens often simplifies interpretation of projection images provided by other techniques. One difficulty with the method is the relatively large amount of metal used to achieve sufficient contrast in bright field images. Thick shadow films are undesirable because they decrease resolution due to an increased tendency for microcrystalline aggregates to form, because decoration artefacts become more severe and increased cap thickness makes estimation of dimensions more uncertain.The large increase in contrast provided by the dark field mode of imaging allows the use of shadow replicas with a much lower average mass thickness. To form the images in Fig. 1, latex spheres of 0.087 μ average diameter were unidirectionally shadowed with platinum carbon (Pt-C) and a thin film of carbon was indirectly evaporated on the specimen as a support.

A Many-Beam Technique for Enhanced Resolution of Partial Dislocations

1972 ◽  
Vol 30 ◽  
pp. 646-647
Author(s):  
J. M. Oblak ◽  
B. H. Kear
Keyword(s):  
Phase Shift ◽  
Field Method ◽  
Dark Field ◽  
Bright Field ◽  
Field Technique ◽  
Weak Beam ◽  
Partial Dislocations ◽  
Enhanced Resolution ◽  
Nickel Base ◽  
Beam Technique

The “weak-beam” and systematic many-beam techniques are the currently available methods for resolution of closely spaced dislocations or other inhomogeneities imaged through strain contrast. The former is a dark field technique and image intensities are usually very weak. The latter is a bright field technique, but generally use of a high voltage instrument is required. In what follows a bright field method for obtaining enhanced resolution of partial dislocations at 100 KV accelerating potential will be described.A brief discussion of an application will first be given. A study of intermediate temperature creep processes in commercial nickel-base alloys strengthened by the Ll2 Ni3 Al γ precipitate has suggested that partial dislocations such as those labelled 1 and 2 in Fig. 1(a) are in reality composed of two closely spaced a/6 <112> Shockley partials. Stacking fault contrast, when present, tends to obscure resolution of the partials; thus, conditions for resolution must be chosen such that the phase shift at the fault is 0 or a multiple of 2π.

Removal of inelastically scattered electrons substantially increases phase contrast on frozen-hydrated molecules

1987 ◽  
Vol 45 ◽  
pp. 652-653
Author(s):  
John P. Langmore ◽  
Brian D. Athey
Keyword(s):  
Electron Diffraction ◽  
Phase Contrast ◽  
Low Dose ◽  
Dark Field ◽  
Biological Structure ◽  
Bright Field ◽  
Low Contrast ◽  
Negative Stain ◽  
The Difference ◽  
Better Than

Although electron diffraction indicates better than 0.3nm preservation of biological structure in vitreous ice, the imaging of molecules in ice is limited by low contrast. Thus, low-dose images of frozen-hydrated molecules have significantly more noise than images of air-dried or negatively-stained molecules. We have addressed the question of the origins of this loss of contrast. One unavoidable effect is the reduction in scattering contrast between a molecule and the background. In effect, the difference in scattering power between a molecule and its background is 2-5 times less in a layer of ice than in vacuum or negative stain. A second, previously unrecognized, effect is the large, incoherent background of inelastic scattering from the ice. This background reduces both scattering and phase contrast by an additional factor of about 3, as shown in this paper. We have used energy filtration on the Zeiss EM902 in order to eliminate this second effect, and also increase scattering contrast in bright-field and dark-field.

Density-based discrimination of protein and RNA in the ribosome

1992 ◽  
Vol 50 (1) ◽  
pp. 464-465
Author(s):  
Joachim Frank
Keyword(s):  
Transfer Function ◽  
Spatial Frequency ◽  
Three Dimensional ◽  
Wiener Filter ◽  
Dark Field Image ◽  
Dark Field ◽  
Frequency Range ◽  
Bright Field ◽  
Contrast Transfer Function ◽  
Pass Filter

Cryo-electron microscopy combined with single-particle reconstruction techniques has allowed us to form a three-dimensional image of the Escherichia coli ribosome.In the interior, we observe strong density variations which may be attributed to the difference in scattering density between ribosomal RNA (rRNA) and protein. This identification can only be tentative, and lacks quantitation at this stage, because of the nature of image formation by bright field phase contrast. Apart from limiting the resolution, the contrast transfer function acts as a high-pass filter which produces edge enhancement effects that can explain at least part of the observed variations. As a step toward a more quantitative analysis, it is necessary to correct the transfer function in the low-spatial-frequency range. Unfortunately, it is in that range where Fourier components unrelated to elastic bright-field imaging are found, and a Wiener-filter type restoration would lead to incorrect results. Depending upon the thickness of the ice layer, a varying contribution to the Fourier components in the low-spatial-frequency range originates from an “inelastic dark field” image. The only prospect to obtain quantitatively interpretable images (i.e., which would allow discrimination between rRNA and protein by application of a density threshold set to the average RNA scattering density may therefore lie in the use of energy-filtering microscopes.

A New Detector System For The HB5 Stem Instrument

1985 ◽  
Vol 43 ◽  
pp. 134-135
Author(s):  
J.M. Cowley
Keyword(s):  
Dark Field ◽  
Detector System ◽  
Electron Holography ◽  
Small Specimen ◽  
Bright Field ◽  
Multiple Images ◽  
Practical Applications ◽  
Wide Range ◽  
Diffraction Patterns ◽  
Operational Modes

The HB5 STEM instrument at ASU has been modified previously to include an efficient two-dimensional detector incorporating an optical analyser device and also a digital system for the recording of multiple images. The detector system was built to explore a wide range of possibilities including in-line electron holography, the observation and recording of diffraction patterns from very small specimen regions (having diameters as small as 3Å) and the formation of both bright field and dark field images by detection of various portions of the diffraction pattern. Experience in the use of this system has shown that sane of its capabilities are unique and valuable. For other purposes it appears that, while the principles of the operational modes may be verified, the practical applications are limited by the details of the initial design.

Systematic zone-Axis orientation of NM-scale particles for microdiffraction

1990 ◽  
Vol 48 (4) ◽  
pp. 262-263
Author(s):  
E D Boyes ◽  
L Hanna
Keyword(s):  
Real Space ◽  
Dark Field ◽  
Zone Axis ◽  
High Symmetry ◽  
Lattice Imaging ◽  
Cell Level ◽  
Bright Field ◽  
Axis Orientation ◽  
Annular Dark Field ◽  
Target Designation

A VG HB501 FEG STEM has been modified to provide track whilst tilt [TWIT] facilities for controllably tilting selected and initially randomly aligned nanometer-sized particles into the high symmetry zone-axis orientations required for microdiffraction, lattice imaging and chemical microanalysis at the unit cell level. New electronics display in alternate TV fields and effectively in parallel on split [+VTR] or adjacent externally synchronized screens, the micro-diffraction pattern from a selected area down to <1nm2 in size, together with the bright field and high angle annular dark field [HADF] STEM images of a much wider [˜1μm] area centered on the same spot. The new system makes it possible to tilt each selected and initially randomly aligned small particle into a zone axis orientation for microdiffraction, or away from it to minimize orientation effects in chemical microanalysis. Tracking of the inevitable specimen movement with tilt is controlled by the operator, with realtime [60Hz] update of the target designation in real space and the diffraction data in reciprocal space. The spot mode micro-DP and images of the surrounding area are displayed continuously. The regular motorized goniometer stage for the HB501STEM is a top entry design but the new control facilities are almost equivalent to having a stage which is eucentric with nanometric precision about both tilt axes.

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