Autofluorescence
Definition
Native fluorescence of endogenous biological molecules (collagen, elastin, NADH, flavins) that produces background signal in fluorescence microscopy, potentially mimicking specific probe signals. Controlled by unstained substrate controls and multicolour spectral discrimination.
- Source
- Collagen, elastin, NADH, flavins
- Risk
- Mimics a genuine probe signal
- Control
- Unstained substrate control
- Mitigation
- Multicolour spectral discrimination
Common questions
How do examiners tell autofluorescence apart from a real stain signal?+
They run an unstained control of the same substrate to see what it fluoresces on its own, and compare the emission spectrum of the questioned signal against the known spectral profile of the specific probe used.
Which forensic materials are most prone to distracting autofluorescence?+
Natural and some synthetic fibers, paper, certain plastics, and biological tissue commonly show background autofluorescence that must be distinguished from a targeted stain or dye signal.
Related terms
- Birefringence
- The difference between the highest and lowest refractive indices of an anisotropic mineral, seen as a characteristic interference colour between crossed polars....
- Conoscopic Illumination
- A convergent polarised beam mode in which a range of angles is passed through the specimen simultaneously, producing an interference figure (optical...
- Crossed Nicols
- The configuration in which the polariser (below the condenser) and the analyser (above the objective) are oriented at 90 degrees to each...
- DAPI
- 4',6-diamidino-2-phenylindole; a fluorescent dye that binds double-stranded DNA and emits blue (~460 nm) under UV excitation (~360 nm). Standard nuclear counterstain; used...
- Dichroic Mirror
- A spectrally selective beam splitter that reflects shorter (excitation) wavelengths at 45 degrees and transmits longer (emission) wavelengths, forming the core element...
- Epi-Fluorescence Illumination
- Fluorescence excitation through the objective lens from above the specimen, with the same objective collecting the emission. A dichroic mirror directs excitation...
- Extinction Angle
- The angle, measured in degrees on the rotating stage of a petrographic microscope, between a crystallographic reference direction (cleavage trace or crystal...
- Michel-Lévy Chart
- A birefringence-thickness diagram correlating retardation values with the interference colour seen at that retardation under white-light crossed polars. Standard reference for identifying...
- Photobleaching
- The irreversible photochemical destruction of a fluorophore by repeated excitation. Limits the observation time for low-copy forensic fluorescence samples. Mitigated by antifade...
- Retardation
- The path difference (in nm) introduced between the fast and slow ray components as they traverse an anisotropic specimen of given thickness...
- Stokes Shift
- The wavelength difference between the excitation peak and the emission peak of a fluorophore. A positive Stokes shift (emission at longer wavelength...