Microscopes - Multiphoton microscope

A multiphoton laser scanning microscope (MPLSM, also multi-photon microscopy, MPM) is a special light microscope from the group of laser scanning microscopes. Images are produced by exploiting one of two different physical phenomena: multiphoton fluorescence (usually two-photon fluorescence) or higher harmonic generation (doubling (Second Harmonic Generation, SHG) or tripling (Third Harmonic Generation, THG) the oscillation frequency of the irradiated light).   A strong, focused laser beam is used to generate nonlinear optical effects based on the interaction of several photons (light particles) arriving simultaneously in a molecule. The strength of the generated signal therefore does not increase linearly with the number of irradiated photons, but with the square (for two-photon effects) or the third power (for three-photon effects).   The operation of a multiphoton microscope is similar to that of a confocal laser scanning microscope. However, while confocal laser scanning microscopy has a penetration depth of 50-80 µm, depending on the specimen, multiphoton microscopy can reach deeper areas, e.g., of 200 µm, in very favorable cases even up to 1000 µm (=1 mm). This allows imaging of living tissues that are otherwise inaccessible for imaging.