Direct visualization of a small-molecule drug by phase-modulated stimulated Raman scattering microscopy

Coherent Raman scattering microscopy, an emerging imaging technology using nonlinear-optical effects such as coherent anti-Stokes Raman scattering (CARS) or stimulated Raman scattering (SRS), has gained increasing attention because of its highly-sensitive, chemically-specific imaging capability with an intrinsic molecular-vibrational contrast. Despite the recent technical advancements, specific detection of a small-molecule drug in living tissue or cells is still challenging. This is because the Raman signal of the targeted drug is easily buried in Raman scattering of abundant macromolecules such as lipids, proteins or water. The time-resolved CARS method is known as a promising solution to reduce such a Raman-induced background [1]. With this approach, we have developed a compact single-beam heterodyne CARS micro-spectroscopy system [2] that efficiently removes the short-lived Raman background and non-resonant background, as well as two-photon fluorescence background. However, as the conventional single-beam CARS requires a narrowband probe pulse to obtain high spectral resolution, the detection sensitivity has been limited by the weak probe power.