Refractive effects in coherent anti-Stokes Raman scattering microscopy.

Coherent anti-Stokes Raman scattering (CARS) microscopy with high sensitivity and high three- dimensional resolution has been developed for the vibrational imaging of chemical species. Due to the coherent nature of the CARS emission, it has been reported that the detection of epi-CARS and forward-CARS (F-CARS) signals depends on the size and shape of the sample. We investigate theoretically and experimentally the effects on the CARS signal of refractive index mismatches between the sample and its surroundings. Backward-CARS and F-CARS signals are measured for different polystyrene bead diameters embedded in different refractive index solvents. We show that index mismatches result in a backward-reflected F-CARS signal that generally dominates the experimentally backward-detected signal. Simulations based on geometrical and wave optics comparing forward- and backward-detected signals for polystyrene beads embedded in different index solvents confirm our findings. Furthermore, we demonstrate that the maxima of forward- and backward-detected signals are generated at different positions along the optical axis in the sample if refractive index mismatches are present between the sample and its surroundings.