Nonlinear microscopy, where two or three photons are used to generate an optical signal, is intrinsically confocal and therefore provides direct depth-sectioning and large penetration depths in tissue, enabling the 3D characterization of tissue slices up to 600 µm thickness. In second and third harmonic generation microscopy the nonlinear optical properties of the tissue itself generate the signal, so no fluorescent dyes are required. Therefore, SHG and THG are label-free techniques, which is attractive because dyes may be toxic to tissue or induce photodamage, limiting the duration of an experiment.