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Schematic of a fluorescence microscope. The majority of fluorescence microscopes, especially those used in the life sciences, are of the epifluorescence design shown in the diagram. Light of the excitation wavelength illuminates the specimen through the objective lens.
A fluorophore (or fluorochrome, similarly to a chromophore) is a fluorescent chemical compound that can re-emit light upon light excitation. Fluorophores typically contain several combined aromatic groups, or planar or cyclic molecules with several π bonds .
Toxicity: Aome fluorochromes can be toxic to cells, to tissues, in vivo, or by producing mutations. [10] Limited resolving power: Fluorescence microscopes are limited in their ability to distinguish close objects at the macroscopic level. In comparison, electron microscopes for example, have the capacity to resolve at a much smaller range.
Most light sheet fluorescence microscopes are used to produce 3D images of the sample by moving the sample through the image plane. If the sample is larger than the field of view of the image sensor, the sample also has to be shifted laterally. An alternative approach is to move the image plane through the sample to create the image stack. [32]
IF can additionally be used in combination with other, non-antibody methods of fluorescent staining, e.g., the use of DAPI to label DNA. [10] [11] Examination of immunofluorescence specimens can be conducted utilizing various microscope configurations, including the epifluorescence microscope, confocal microscope, and widefield microscope. [12]
The DyLight Fluor family of fluorescent dyes are produced by Dyomics in collaboration with Thermo Fisher Scientific. [4] DyLight dyes are typically used in biotechnology and research applications as biomolecule , cell and tissue labels for fluorescence microscopy , cell biology or molecular biology .
Distribution of fluorescent proteins in animals. The hippocampus of a mouse imaged via fluorescence microscopy. Biofluorescent emission spectra from amphibians Example uses of fluorescent proteins for imaging in the life sciences. Fluorescence is used in the life sciences generally as a non-destructive way of tracking or analysing biological ...
Strong fluorescence when bound to DNA led to the rapid adoption of DAPI for fluorescent staining of DNA for fluorescence microscopy. Its use for detecting DNA in plant , metazoa and bacteria cells and virus particles was demonstrated in the late 1970s, and quantitative staining of DNA inside cells was demonstrated in 1977.