Optical Microscopy

Optical Microscopy (often known as Light Microscopy) refers to observations under visible and ultraviolet light, although in this case a distinction is made between Visible Light Microscopy (VLM) and Fluorescent Light Microscopy (FLM).
With regard to standard instrumentation, a wealth of monographs and other literature is available. In addition to the wavelength of the radiation under which the observation is performed, the way the illumination is incident on the specimen or object under examination further distinguishes microscopic observation techniques, as images are obtained under reflected or transmitted light, polarised light, etc. Observation under polarised light, for example, is so widely applied in the study of artwork materials that one can speak of a separate technique, referred to as ‘Polarised Light Microscopy (PLM)’ (Kouloumpi et al. 2012, p. 373). At the same time, in the examination of paintings the most widely used imaging mode is dark field illumination, in which the light path is directed from the source outside the objective lenses, then reflected by the surface under study and finally returned through the objective back to the eyepiece lenses and the digital camera.

With respect to Fluorescence Microscopy, the purpose of the method is to offer the possibility of separating elements of the sample based on the fluorescence radiation in the visible produced by the excitation with UV radiation of specific wavelengths. Ultraviolet radiation of a specific wavelength or combination of wavelengths produced by a high energy mercury lamp is passed through the excitation filter and incident on the sample. Wavelengths close to the visible region are usually used, as most fluorescent groups found in plant tissues fluoresce strongly under stimulation in this region. The filtered UV radiation illuminates the sample, which emits fluorescent light of longer wavelengths, i.e. visible radiation (Stokes’ law). The emitted visible radiation passes through the cut-off filter which does not allow the reflected UV radiation to pass through. The resulting fluorescent areas of the sample radiate against a dark background with sufficient contrast to allow detection. And the darker the background of the non-fluorescent material, the more effective the imaging.

“Classical” optical microscopy is an invasive method and is applied to a microsample of the material under study after appropriate processing. However, despite all the technical limitations, the method can also be used to study the micromorphology of the surface in a non-invasive way, i.e. without taking a microsample, if the artwork can be placed on the microscope bench as it is.


(Handbook_of_Physical_Testing_of_Paper, n.d.; Pinna et al., 2009; Terlixi & Kouloumpi, 2012)

Translated with DeepL.com (free version)

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