Leica SP8 Multiphoton Microscope

Multiphoton microscopy (MPM) is an advanced fluorescence imaging technique for generating high-resolution optical sections from thick biological samples. While laser scanning confocal microscopy (LSCM) utilises a pinhole to remove out-of-focus lights from the image, MPM restricts the excitation volume to the laser focus using a pulsed long-wavelength laser. As the focus of the laser is the only location where photons are dense enough to excite the fluorophores, optical sectioning can be achieved without a pinhole. On the other hand, the use of excitation laser light in the infrared range is particularly suitable for live fluorescence imaging, for it would only cause minimal photocytotoxicity to live tissues and cells, as well as photobleaching to fluorophores when compared to the use of visible lasers in LSCM. Furthermore, infrared laser light can penetrate animal tissues with much less light scattering, making deep tissue imaging possible. The laser with continuously tuneable wavelengths also allows researchers to examine a wider range of fluorescent probes with diverse excitation spectra.