two-photon microscopy
(rus. микроскопия, двухфотонная)
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a technique used for detecting fluorescent microscale objects using a light microscope, where the object's fluorescence is induced as a result of simultaneous absorption of two photons of comparable lower energy than needed for fluorescence excitation.
Description
Photon emission by a molecule (fluorescence) occurs in molecules at a high-energy state . A molecule is raised into an excited high energy state when it absorbs a photon of energy equal to the difference between the ground state and the excited state. However, transition into a high-energy state is also possible when a molecule absorbs several low-energy photons at once. While the probability of this event is proportional to the squared value of the intensity of light focused on a sample, such simultaneous absorption and, correspondingly, simultaneous emission (two-photon fluorescence) may only be observed in the focal plane of a microscope system.
The high degree of localisisation of excitation and the use of lower-energy photons compared to conventional fluorescence-based techniques (see fluorescence microscopy) make it possible to image samples at a high focal depth (up to 1 mm) with a high contrast image, protecting the specimens from fading or decomposition due to light effects. Confocal microscopy offers similar opportunities.
The high degree of localisisation of excitation and the use of lower-energy photons compared to conventional fluorescence-based techniques (see fluorescence microscopy) make it possible to image samples at a high focal depth (up to 1 mm) with a high contrast image, protecting the specimens from fading or decomposition due to light effects. Confocal microscopy offers similar opportunities.
Author
- Grigory G. Borisenko
Source
- Denk W., Strickler J.H., Webb W.W. Two-photon laser scanning fluorescence microscopy // Science. 1990. V. 248. P. 73–76.