Coherent Anti-Stokes Raman Scattering Imaging: A Basic Overview With An Emphasis On The Optics Required

Coherent anti-­Stokes Raman scattering (CARS) imaging is a nonlinear optical technique that captures intrinsic molecular vibrations to create optical contrast. It offers submicron level spatial resolution and video-­speed imaging rate without the needs for extrinsic labels (e.g. fluorochromes). In a CARS process, a pump laser, a Stokes laser and a probe laser (usually same as the Pump laser) are involved. When the frequency difference (beating frequency) between the pump and the Stokes lasers is in resonance with the frequency of a molecular vibration, an enhanced optical signal is generated at the anti-­Stokes frequency. The major advantage of CARS is that the signal yield is much higher, typically six orders of magnitude, than the signal yield obtained through the conventional spontaneous Raman scattering process. By tuning the beating frequency to a designated vibrational mode of a chemical bond (e.g. O-­P-­O, H-­C-­H), CARS enables chemically selective imaging of targeted molecules. Several optical designs have been reported for CARS imaging. These designs use different wavelength combinations for the pump and Stokes beams, resulting in variations of CARS emission wavelength for the same chemical bond. Consequently, different dichroic mirrors and emission filters are required. In this application note, we will primarily focus on filter sets for CARS excitation of symmetric vibration of the H-­C-­H bond, which is the most popular vibrational mode used in biomedical imaging applications.