Autofluorescence
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Autofluorescence is the fluorescence of other substances than the fluorophore of interest. It increases the background signal.
Autofluorescence can be problematic in fluorescence microscopy. In most fluorescence microscopy, fluorescent stains (such as fluorescently-labeled antibodies) are applied to samples to stain specific structures. Autofluorescence interferes with detection of the resulting specific fluorescent signals, especially when the signals of interest are very dim — it causes structures other than those of interest to become visible. Depending upon the shape of the structures of interest and the other structures, it may not be obvious that this has occurred. In some microscopes (mainly confocal microscopes), it is possible to make use of different lifetime of the excited states of the added fluorescent markers and the endogenous molecules to exclude most of the autofluorescence.
In a few cases, autofluorescence may actually illuminate the structures of interest, or serve as a useful diagnostic indicator.
Without labelling, these substances show fluorescence. Because of scattering, it is better to use the nonlinear two photon excited fluorescence microscopy.
- chlorophyll (in plant cells)
- collagen
- elastin
- fibrillin
- flavin
- indolamine
- indolamine dimer
- indolamine trimer
- lipofuscin
- NADH (reduced form only)
- polyphenols (in plant cells)
- tryptophan
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