How to build a two-photon microscope with a confocal scan head

This article provides practical guidelines for the conversion of a standard confocal microscope into a two-photon microscope. This conversion enables the investigator to have access to two-photon microscopy without the large budget necessary to purchase a commercial instrument. Two-photon fluorescence microscopy allows deep-tissue imaging in highly scattering preparations and long-term imaging of live tissue without the photodamage that is caused by out-of-focus light. It is, therefore, an essential tool for imaging cells under physiologically relevant conditions such as acute or cultured brain slices or in vivo.