Frequency-modulation Stimulated Raman Scattering microscopy with an Acousto-Optic Tunable Filter

Abstract

Stimulated Raman Scattering (SRS) is a Coherent Raman microscopy method that has been increasingly employed in recent years for highly-specific, label-free, and high-speed bioimaging. Compared to a similar Coherent Raman method, the so-called Coherent Anti-Stokes Scattering (CARS) microscopy, it exhibits advantages such as the absence of nonresonant background (NRB) and the linearity of the signal intensity on the concentration of molecules of interest. However, SRS can be affected by unwanted background signals that hinder the acquisition of an accurate Raman information. These unwanted signals are generated by parasitic effects that are difficult to suppress in standard SRS setups. Here, I present a frequency-modulation (FM) SRS technique via an Acousto-Optic Tunable ioilter (AOTF), describing its implementation on Vibra Lab setup and assessing its efficiency with imaging results. The FM technique provides a cancellation of the unwanted background signals, maintaining intact the SRS information. It is based on the weak spectral dependence of the parasitic effects as compared to the high spectral specificity of the SRS signal.

The proposed scheme presents a few advantages when compared with other solutions presented in the literature. In particular, it doesn't require a complex setup configuration, and it can be used seamlessly in a very broad range of the vibrational spectrum.