Three-photon fluorescence microscopy is poised to initiate a wave of biomedical discoveries by allowing researchers to produce high-resolution images of living tissues that are beyond the depth limit of two-photon technology. But current three-photon technology has limitations: the signal is inherently weaker than the signal produced by two-photon technology, making it harder to capture rapid cellular processes; laser sources for three-photon microscopy are not yet optimized for reaching deep tissues; and the laser systems currently in use for three-photon microscopy are too complex and costly for a typical research lab.

An interdisciplinary team of Cornell engineers led by Chris Xu, Applied and Engineering Physics, is developing a new generation of three-photon fluorescence microscopy that can quickly capture dynamic cellular processes in living tissues using a single excitation wavelength and a low-cost fiber laser system. This research aims to make three-photon microscopy a powerful but routine instrument for deep-tissue imaging, just as two-photon microscopy is for shallower regions of biological tissues and organs.

The ability to create high-resolution optical images of living tissues deep within an organism has the potential to power major advances in neuroscience, immunology, and cancer biology. This research will enable visualization of dynamic processes at the subcellular level in intact organs and animal models that are beyond the reach of any existing imaging techniques. Furthermore, whereas applications of three-photon excitation fluorescence microscopy have been developed mostly in connection with brain tissue, this project will apply novel three-photon imaging techniques to a wide variety of tissues and biological systems, creating a valuable knowledge base for researchers.

NIH Award Number: 1R01EB033179-01