= Two-photon excitation microscopy
= Two-photon microscopy
{synonym}
<Two-photon excitation microscopy> excites <fluorescence> through the nearly simultaneous absorption of two <photons>. In the weak unsaturated regime, the excitation rate is proportional to the square of the local illumination <intensity>. Focusing a pulsed <laser> therefore strongly favors excitation near its focus rather than throughout the illuminated path.
Scanning that focus provides <optical sectioning> without relying on a detection pinhole to reject all out-of-focus emission. The <photons> can have lower individual <energies> than a single excitation <photon>, but their combined <energy> must match an allowed transition. High peak <intensity>, bleaching and <phototoxicity> remain relevant; longer excitation <wavelength> alone does not guarantee less damage.<Two-photon laser scanning fluorescence microscopy by Winfried Denk and others>[Abstract: intrinsic three-dimensional localization through simultaneous two-photon absorption]{ref}
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