Time-domain measurements reveal spatial aberrations in a sub-surface two-photon microscope

We show that in a nonlinear microscopy system the effects of chromatic and spherical aberrations are revealed by a difference in the focal positions corresponding to the shortest pulse duration and the minimum lateral resolution. By interpreting experimental results from a high-numerical-aperture two-photon microscope using a previously reported spatio-temporal model, we conclude that the two-photon autocorrelation of the pulses at the focal plane can be used to minimize both the chromatic and spherical aberrations of the system. Based on these results, a possible optimization strategy is proposed whereby the objective lens is first adjusted for minimum autocorrelation duration, and then the wavefront before the objective is modified to maximize the autocorrelation intensity.