Multiphoton fluorescence microscopy is a powerful research tool that combines the advanced optical techniques of laser scanning microscopy with long wavelength multiphoton fluorescence excitation to capture high-resolution, three-dimensional images of specimens tagged with highly specific fluorophores. The technique features attractive advantages over confocal microscopy for imaging living cells and tissues with three-dimensionally resolved fluorescence imaging. Multiphoton excitation, which occurs only at the focal point of the microscope, minimises the photobleaching and photodamage that are the ultimate limiting factors in imaging live cells. This advantage allows investigations on thick living tissue specimens that would not otherwise be possible with conventional imaging techniques.