Cascaded domain multiphoton spatial frequency modulation imaging.

Abstract

Multiphoton microscopy is a powerful imaging tool for biomedical applications. A variety of techniques and respective benefits exist for multiphoton microscopy, but an enhanced resolution is especially desired. Additionally multiphoton microscopy requires ultrafast pulses for excitation, so optimization of the pulse duration at the sample is critical for strong signals. We aim to perform enhanced resolution imaging that is robust to scattering using a structured illumination technique while also providing a rapid and easily repeatable means to optimize group delay dispersion (GDD) compensation through to the sample. Spatial frequency modulation imaging (SPIFI) is used in two domains: the spatial domain (SD) and the wavelength domain (WD). The WD-SPIFI system is an in-line tool enabling GDD optimization that considers all material through to the sample. The SD-SPIFI system follows and enables enhanced resolution imaging. The WD-SPIFI dispersion optimization performance is confirmed with independent pulse characterization, enabling rapid optimization of pulses for imaging with the SD-SPIFI system. The SD-SPIFI system demonstrates enhanced resolution imaging without the use of photon counting enabled by signal to noise improvements due to the WD-SPIFI system. Implementing SPIFI in-line in two domains enables full-path dispersion compensation optimization through to the sample for enhanced resolution multiphoton microscopy.