Two-photon microscopy (TPM) has revolutionized the field of biology by enabling researchers to observe complex biological processes in living tissues at high resolution. In contrast to traditional fluorescence microscopy techniques, TPM makes use of low-energy photons to excite fluorescent molecules for observation. This, in turn, makes it possible to penetrate the tissue much more deeply, and ensures that the fluorescent molecules, or fluorophores, are not permanently damaged by the excitation laser.