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Spatial-spectral coupling in hyperspectral CARS microscopy image formation

Hyperspectral coherent anti-Stokes Raman scattering (CARS) microscopy has provided an imaging tool for extraction of 3-dimensional volumetric information, as well as chemically-sensitive spectral information. These techniques have been used in a variety of different domains including biophysics, geology, and material science. The measured CARS spectrum results from interference between the Raman response of the sample and a non-resonant background. We have collected four dimensional data sets (three spatial dimensions, plus spectra) and extracted Raman response from the CARS spectrum using a Kramers-Kronig transformation. However, the three dimensional images formed by a CARS microscope are distorted by interference, some of which arises because of the Gouy phase shift. This type of interference comes from the axial position of the Raman resonant object in the laser focus. We studied how the Gouy phase manifests itself in the spectral domain by investigating microscopic diamonds and nitrobenzene droplets in a CARS microscope. Through experimental results and numerical calculation using finite-diference time-domain (FDTD) methods, we were able to demonstrate the relationship between the spatial configuration of the sample and the CARS spectral response in three dimensional space.