Polarization-Modulated Second Harmonic Generation Ellipsometric Microscopy at Video Rate

Abstract

Fast 8 MHz polarization modulation coupled with analytical modeling, fast beam-scanning, and synchronous digitization (SD) have enabled simultaneous nonlinear optical Stokes ellipsometry (NOSE) and polarized laser transmittance imaging with image acquisition rates up to video rate. In contrast to polarimetry, in which the polarization state of the exiting beam is recorded, NOSE enables recovery of the complex-valued Jones tensor of the sample that describes all polarization-dependent observables of the measurement. Every video-rate scan produces a set of 30 images (10 for each detector with three detectors operating in parallel), each of which corresponds to a different polarization-dependent result. Linear fitting of this image set contracts it down to a set of five parameters for each detector in second harmonic generation (SHG) and three parameters for the transmittance of the incident beam. These parameters can in turn be used to recover the Jones tensor elements of the sample. Following validation of the approach using z-cut quartz, NOSE microscopy was performed for microcrystals of both naproxen and glucose isomerase. When weighted by the measurement time, NOSE microscopy was found to provide a substantial (>7 decades) improvement in the signal-to-noise ratio relative to our previous measurements based on the rotation of optical elements and a 3-fold improvement relative to previous single-point NOSE approaches.

Imaging the Nonlinear Susceptibility Tensor of Collagen by Nonlinear Optical Stokes Ellipsometry

“…4 and 5. Subsequent conversion to polynomial coefficients was accomplished through a simple matrix multiplication before solving for local tensor elements, as described previously in DeWalt et al (27). Contrast in the Fourier coefficient color maps of the collagenous tissues is largely dependent on the orientation of the fibers within the FOV.…”

“…The ability to perform polarized laser transmittance imaging simultaneously with SHG imaging illustrated in this and De Walt et al (27) served a dual purpose, both to quantify the polarization-state of the electrooptic modulator from global analysis across the image and assess on a per-pixel basis the potential significance of birefringence. In all the measurements described in this work, the measured effects from local birefringence were negligible relative to other sources of measurement uncertainty.…”

“…While the images presented in the preceding figures were signal-averaged with 0.3-1 ms per pixel measurement times, polarization images of collagen samples can be acquired at video rate (15 frames per s) (27) with as little as~150 ns per pixel total measurement time. The high achievable framerate coupled with image stabilization algorithms can allow for similar signal averaging advantages even in samples with positions changing in time (e.g., for in vivo analysis of skin).…”

“…NOSE is based on polarization-dependent SHG imaging, and utilizes rapid polarization modulation with EOM and analytical modeling to recover the unique polarizationdependent parameters of the sample. This work builds upon previous studies where NOSE imaging was carried out at video rate on a variety of crystalline samples, enabling recovery of parameters directly related to the laboratory-frame Jones tensor (27). Here, we developed an iterative approach to investigate the orientation angles of the collagen structure and extend the analysis to recovery of the local-frame tensor elements.…”

“…SHG and laser transmittance signals were collected using a custom microscope as described in detail in DeWalt et al (27) and shown schematically in Fig. 3.…”