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Multiphoton Excitation

Left: A single input photon (blue) excites a single photon (green).
Right: Multiphoton Excitation. Two input photons (red) combine to excite a single photon (green).
MPE is the most simple form of Multiphoton Microscopy (MPM) which is used to excite fluorescence. A pulsed laser is used to provide simultaneous excitation with two (or three) photons of low energy, exciting the fluorophore to the same level as one high energy photon. MPE has many advantages over normal laser scanning microscopy. The use of lower energy photons (infrared) reduces damage to the sample and also permits deeper penetration into scattering samples. Multiphoton excitation occurs only at the focal plane, removing the need for pinhole apertures.

As well as excitation of extraneous fluorophores, MPE can be used to excite indigenous tissue components such as NAD(P)H, flavins, and porphyrins. The figure below shows an MPE image of artery wall using elastin as the intrinsic fluorophore.


Elastin in an artery wall viewed with Multiphoton Excitation

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