Automated correction of optical attenuation in stimulated Raman scattering imaging

Transmission losses in stimulated Raman scattering (SRS) microscopy degrade quantitative chemical imaging by introducing artifacts. We present a real-time correction scheme based on active electronic gain control to independently compensate for pump and Stokes transmission losses. In addition to correcting Stokes losses in transmission mode, our method uses the pump light that is backscattered from the focal volume to measure the pump transmission factors affecting the SRS process and exploits the logarithmic gain interface of a variable gain amplifier to implement the required square root operation for compensation. Validation experiments on both homogeneous and heterogeneous samples demonstrated a chemically specific contrast that is almost independent of transmission losses, leading to a significant improvement in image uniformity and representing a step towards quantitative SRS imaging.