Multiphoton imaging relies on the simultaneous absorption of two photons of longer wavelength (and thus lower energy) to excite a fluorophore. This process reduces photodamage and allows for deeper tissue penetration. A pulsed laser, typically a titanium-sapphire laser, is used to deliver the photons. The emission from the excited fluorophores is then collected to form an image. The nonlinear nature of the excitation confines the fluorescence to the focal plane, providing inherent optical sectioning and reducing background noise.
