Stimulated Raman scattering (SRS) microscopy that is capable of both high speed imaging and rapid spectroscopy will be advantageous for detailed chemical analysis of heterogeneous biological specimens. We have developed a system based on spectral focusing SRS technology, with the integration of a rapid scanning optical delay line (RSODL), which allows continuous tuning of SRS spectra by scanning a galvo mirror. We demonstrated SRS spectral measurements of dimethyl sulfoxide solution at low concentrations, and multi-color imaging of rice pollens and HeLa cells with line-by-line delay tuning to reduce motion artifacts, as well as fast acquisition of SRS spectra at specific regions of interest.
Two-color Stimulated Raman scattering (SRS) microscopy has shown great potential in label-free digital histology with diagnostic results similar to H&E stain. However, achieving real-time two-color SRS imaging is challenging. We have precisely engineered the pulse profiles of the Stokes beams, and fully utilized the in-phase (X) and quadrature (Y) outputs of a phase sensitive lock-in amplifier to realize simultaneous two-color SRS imaging. We have demonstrated its robustness and advantages in rapid histology, as well as real-time in vivo imaging of live animals, both in transmission and epi modes. Moreover, we have also adapted this method to other pump-probe based microscopes, such as transient absorption (TA) microscopy.
A compact, alignment-free, and inexpensive fiber source for coherent Raman spectroscopy would benefit the field considerably. We present a fiber optical parametric oscillator offering the best performance from a fiber-source to date. Pumping the oscillator with amplified pulses from a 1 μm fiber laser, we achieve widely spaced, narrowband pulses suitable for coherent anti-Stokes Raman scattering microscopy. The nearly transform limited, 2 ps signal pulses are generated through the use of normal dispersion four wave mixing in photonic crystal fiber, and can be tuned from 779-808 nm, limited by the tuning range of the seed laser. The average signal power can reach 180 mW (pulse energies up to 4 nJ). The long-wavelength idler field is resonant in the oscillator, and the use of a narrow bandpass filter in the feedback loop is critical for stable operation, as seen in both simulation and experiment. Due to the self-consistent nature of the oscillator, this source provides lower relative intensity noise on its output pulses than parametric amplifiers based on the same frequency conversion process. We present high quality images of mouse tissues taken with this source that exhibit an outstanding signal to noise ratio at top imaging speeds.