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Digital Image Processing and Three-Dimensional Reconstruction in the Basic Neurosciences
Published in Rangachar Kasturi, Mohan M. Trivedi, Image Analysis Applications, 2020
Autoradiography is the generation of an image in a photographic or radiographic emulsion by the radiation emitted from a radioactive substance. In studies of metabolism, transport, and blood flow, the experiment involves the injection of a radiolabeled substance into the circulation, followed by sacrifice of the animal (most commonly the rat), sectioning of the brain, and incubation of the tissue sections with a radiographic film. After a period of time (a few days to a few months, depending on the isotope and the nature of the experiment), the film is developed, revealing life-size images of the tissue sections in which light and dark areas represent brain regions of low and high concentrations, respectively, of the tracer in the tissues. Depending on the nature of the molecule containing the radiolabel and on the design of the experiment, the observation of rates of glucose metabolism, blood flow, or transport across the blood-brain barrier may be quantitatively measured (Fig.5.2). (The regional mapping of receptors will be described later.)
Thin-Layer Chromatography
Published in James P. Lodge, Methods of Air Sampling and Analysis, 2017
Separations involving dyes and pigments can be viewed directly. However, when compounds are not detectable by their visible color, one must search for a method of locating these materials on the chromatogram. The immediate tendency is to spray the plate with a non-selective color forming reagent (see Figure 23:3). This practice should be resisted at least until some thought is given to the many nondestructive visualization methods available. Often, compounds can be located by their fluorescence under either long (366 nm) or short (254 nm) wavelength ultraviolet light, or by their ability to quench the fluorescence of an inorganic phosphor. Plates can be purchased with the fluorescent phosphor impregnated into the plate coating, or it can be added as a spray reagent after migration. A light spray of water is often repelled by hydrophobic compounds on the chromatogram so that white spots appear on a translucent background. Iodine vapor is reversibly absorbed by many organic compounds and thus may be used as a nondestructive visualization technique. Autoradiography is a highly sensitive method for monitoring of separations of radioactive or radiolabelled compounds.
Laboratory tutorials
Published in Firdos Alam Khan, Biotechnology Fundamentals, 2018
Histology samples have often been examined by radioactive techniques. In historadiography, a slide (sometimes stained histochemically) is x-rayed. More commonly, autoradiography is used to visualize the locations to which a radioactive substance has been transported within the body, such as cells in the S phase (undergoing DNA replication) that incorporate tritiated thymidine or sites to which radiolabeled nucleic acid probes bind in in situ hybridization. For autoradiography on a microscopic level, the slide is typically dipped into liquid nuclear tract emulsion, which dries to form the exposure film. Individual silver grains in the film are visualized with dark field microscopy. Recently, antibodies have been used to specifically visualize proteins, carbohydrates, and lipids, which is called immunohistochemistry; when the stain is a fluorescent molecule, it is called immunofluorescence. This technique has greatly increased the ability to identify categories of cells under a microscope. Other advanced techniques, such as non-radioactive in situ hybridization, can be combined with immunochemistry to identify specific DNA or RNA molecules with fluorescent probes or tags that can be used for immunofluorescence and enzyme-linked fluorescence amplification (especially alkaline phosphatase and tyramide signal amplification). Fluorescence microscopy and confocal microscopy are used to detect fluorescent signals with good intracellular detail. Digital cameras are increasingly used to capture histological and his-topathological images. The Nissl method and Golgi’s method are useful in identifying neurons.
Bioimaging of metals in environmental toxicological studies: Linking localization and functionality
Published in Critical Reviews in Environmental Science and Technology, 2022
Radioisotopes can be further employed for sectioning in autoradiographic analysis, although there have been only a few reports on the tissue distribution of metals using this technique. Autoradiography basically images the radioisotopes in a biological specimen, and there is both macroautoradiography (e.g., whole body imaging) and microautoradiography. Imaging may take weeks or even months, but it can generally provide images with reasonable resolution. Phosphor imager now provides much quicker imaging and it is now possible to quantify the radiotracer concentration with this technology (Rouleau & Kohli, 2008). Microautoradiography may enable the spatial differentiation of radioactivity at the tissue or even cellular level, although there are very few applications of this technique in metal environmental toxicological studies. Among the limited applications, Cresswell et al. (2015) radiolabeled 109Cd and 65Zn in water used as an exposure medium for a freshwater prawn Macrobrachium australiense for 3 weeks, followed by 2 weeks depuration in clean unlabeled water. The spatial distribution of the radiotracers was mapped using autoradiography after cryo-sectioning. Images showed that there were clear differences between Cd and Zn, e.g., most Cd was found in the gills and hepatopancreas whereas Zn was concentrated in the antennal gland and was transported much faster than Cd. A follow-up study examined the 109Cd in the same species of prawn, which appeared to be homogenously distributed in the hepatopancreas (Cresswell et al., 2017). The limit of autoradiographic detection of 109Cd was around 0.01 ng/cm2, which was sufficient for measuring the concentrations of accumulated Cd in the tissues. Pinho et al. (2011) exposed the copepod Calanus hyperboreus to radioactive 64Cu for 2 h, at a rather high concentration (2.3 mg/L), followed by 2 h depuration with radioactive Cu and the distribution of 64Cu in different tissues of Cu was determined by autoradiography. It was obvious that 64Cu was attached on the surface of copepods, especially in the ventral parts, likely due to the short period (2 h) of exposure. Much earlier, Rouleau et al. (1999) employed autoradiography to examine the distribution of 203Hg(II) in two freshwater fish (Salmo trutta, and rainbow trout Oncorhynchus mykiss). Different methods of dosing (waterborne exposure or intravenous injection) resulted in different Hg distributions in the fish bodies, especially in the brain. Apparently waterborne Hg(II) was distributed in the brains in different areas (olfactory system, spinal cord, and others). In general, application of autoradiography is metal ecotoxicological research is still limited nowadays. Most of these studies mainly focused on tissue distribution imaging, but with limited spatial resolution. Given the requirements of long-term film exposure (some of which may even take 10 months), it is impractical to use such technique for radioisotopes which have a very short half life.