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ISO 18337-2015 pdf free download
ISO 18337-2015 pdf free download.Surface chemical analysis Surface characterization Measurement of the lateral resolution of a confocal fluorescence microscope
Analyse chimique des surfaces — Caractérisation des surfaces— Mesurage de la resolution latérale d’un microscope confocal a fluorescence.
5.2 Selection of the sample and sample requirements
lithe size of the fluorescent nanoparticle is comparable to the size of the PSF in a CFM, a deconvolution process for deducing the actual PSF of the microscope is required. Deconvolution complicates the process of characterizing the PSF, The nanoparticles used here should be as small as possible; however, small particles produce a weaker fluorescence signal that can degrade the signal-to-noise ratio in a CFM image. Assuming that the PSF has a Gaussian profile, a particle less than one-fourth the size of the PSF will contribute less than I % to the resultant imagell 141. Considering the uncertainty of measuring the FWHM of a particle, this discrepancy may be taken as negligible. One-fourth the size of a typical PSF corresponds to 75 nm for a 300 nm PSF, indicating that one can use a 75 nm nanoparticle as a test specimen without worrying about deconvoluting the PSF from the obtained image. The apparent size of the nanoparticle in the image may then be taken as the lateral profile of the PSF The shape of the fluorescent nanoparticle does not matter as long as the size in any direction of the nanoparticle is less than one fourth of the expected lateral resolution. Note that specified sizes of the manufactured fluorescent nanoparticles are not always correct with some variation depending on the manufacturer. The confirmation of the size of the nanoparticles can be obtained by using other high-resolution microscopes, such as atomic force microscopes and electron microscopes.
Nanoparticies must be immobilized on a clean substrate with low roughness, such as glass or a silicon wafer. If the nanoparticle is purchased In the form of a colloid, the colloid must be appropriately diluted prior to dispersal on the substrate. Colloidal solutions may be spread or spin-coated onto a substrate to an optimum density of several isolated particles per 100 pm2. The cleanness of the substrate is very important. A substrate may be cleaned using a Piranha solutionl9l or by ultrasonicating the substrate in an alkaline 1 M KOH solution, followed by washing with H20. Plasma cleaning is effective as well11f1,lUl.
5.3 Setting the parameters prior to operating the instrument
An image obtained by CFM depends on the operator’s skill and the experimental parameters. Different CFM resolution values may be obtained for a given parameter set for different instrument operators. The extent to which a properly optimized experimental parameter set is achieved will depend on the operator. Therefore, this International Standard does not impose specific experimental parameters for use in operating a CFM instrument. Instead, the operator may choose the appropriate experimental parameters for optimized CFM imaging at his or her discretion.ISO 18337-2015 pdf free download.
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