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10.3791/50988 http://scihub22266oqcxt.onion/10.3791/50988 C4162510!4162510 !24747818     free     free     free Warning: file_get_contents(https://eutils.ncbi.nlm.nih.gov/entrez/eutils/elink.fcgi?dbfrom=pubmed&id=24747818 &cmd=llinks): Failed to open stream: HTTP request failed! 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Quantitative optical microscopy: measurement of cellular biophysical features with a standard optical microscope |pdf=|usr=}}{{24747818 }} gab.com Text Quantitative optical microscopy: measurement of cellular biophysical features with a standard optical microscope JO: J Vis Exp http://www.kidney.de/pget.php?&tw=1&pmid=24747818 #FOAvip We describe the use of a standard optical microscope to perform quantitative measurements of mass, volume, and density on cellular specimens through a combination of bright field and differential interference contrast imagery. Two primary approaches are presented: noninterferometric quantitative phase microscopy (NIQPM), to perform measurements of total cell mass and subcellular density distribution, and Hilbert transform differential interference contrast microscopy (HTDIC) to determine volume. NIQPM is based on a simplified model of wave propagation, termed the paraxial approximation, with three underlying assumptions: low numerical aperture (NA) illumination, weak scattering, and weak absorption of light by the specimen. Fortunately, unstained cellular specimens satisfy these assumptions and low NA illumination is easily achieved on commercial microscopes. HTDIC is used to obtain volumetric information from through-focus DIC imagery under high NA illumination conditions. High NA illumination enables enhanced sectioning of the specimen along the optical axis. Hilbert transform processing on the DIC image stacks greatly enhances edge detection algorithms for localization of the specimen borders in three dimensions by separating the gray values of the specimen intensity from those of the background. The primary advantages of NIQPM and HTDIC lay in their technological accessibility using "off-the-shelf" microscopes. There are two basic limitations of these methods: slow z-stack acquisition time on commercial scopes currently abrogates the investigation of phenomena faster than 1 frame/minute, and secondly, diffraction effects restrict the utility of NIQPM and HTDIC to objects from 0.2 up to 10 (NIQPM) and 20 (HTDIC) ?m in diameter, respectively. Hence, the specimen and its associated time dynamics of interest must meet certain size and temporal constraints to enable the use of these methods. Excitingly, most fixed cellular specimens are readily investigated with these methods. Twit Text FOAVip Quantitative optical microscopy: measurement of cellular biophysical features with a standard optical microscope ????J Vis Exp http://www.kidney.de/pget.php?&tw=1&pmid=24747818 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=24747818 #FOAvip English Wikipedia <ref>{{Cite pmid|24747818 }}</ref> Quantitative optical microscopy: measurement of cellular biophysical features with a standard optical microscope #MMPMID24747818 Phillips KG ; Baker-Groberg SM ; McCarty OJ J Vis Exp 2014[Apr]; ä (86 ): ä PMID24747818 show ga We describe the use of a standard optical microscope to perform quantitative measurements of mass, volume, and density on cellular specimens through a combination of bright field and differential interference contrast imagery. Two primary approaches are presented: noninterferometric quantitative phase microscopy (NIQPM), to perform measurements of total cell mass and subcellular density distribution, and Hilbert transform differential interference contrast microscopy (HTDIC) to determine volume. NIQPM is based on a simplified model of wave propagation, termed the paraxial approximation, with three underlying assumptions: low numerical aperture (NA) illumination, weak scattering, and weak absorption of light by the specimen. Fortunately, unstained cellular specimens satisfy these assumptions and low NA illumination is easily achieved on commercial microscopes. HTDIC is used to obtain volumetric information from through-focus DIC imagery under high NA illumination conditions. High NA illumination enables enhanced sectioning of the specimen along the optical axis. Hilbert transform processing on the DIC image stacks greatly enhances edge detection algorithms for localization of the specimen borders in three dimensions by separating the gray values of the specimen intensity from those of the background. The primary advantages of NIQPM and HTDIC lay in their technological accessibility using "off-the-shelf" microscopes. There are two basic limitations of these methods: slow z-stack acquisition time on commercial scopes currently abrogates the investigation of phenomena faster than 1 frame/minute, and secondly, diffraction effects restrict the utility of NIQPM and HTDIC to objects from 0.2 up to 10 (NIQPM) and 20 (HTDIC) ?m in diameter, respectively. Hence, the specimen and its associated time dynamics of interest must meet certain size and temporal constraints to enable the use of these methods. Excitingly, most fixed cellular specimens are readily investigated with these methods. |Algorithms [MESH] |Biophysical Phenomena [MESH] |Cell Line, Tumor [MESH] |Colorectal Neoplasms/pathology [MESH] |Fourier Analysis [MESH] |Humans [MESH] |Microscopy/*methods [MESH]Quantitative optical microscopy: measurement of cellular biophysical f(epmc).pdf DeepDyve Pubget Overpricing