Stimulated Raman Scattering Polarizing Microscopy
In SRS-PM, two beams, pump (at frequency 蠅p) and Stokes (at frequency 蠅s < 蠅p), combine to amplify the Stokes Raman signal when the difference between 蠅p and 蠅s equals the vibrational frequency (蠅vib) of a certain chemical bond in the molecules [Figure (a)]. As a result of the amplification of the Raman signal, the pump beam experiences a loss in its intensity, dubbed 鈥渟timulated Raman loss鈥 (SRL), while the Stokes beam experiences a gain in its intensity, dubbed 鈥渟timulated Raman gain鈥 (SRG). Since the gain/loss mechanism occurs only when the difference between the frequencies of the pump and Stokes beam equals the molecular vibration frequency, the non-resonant background is significantly reduced compared to that of CARS. The SRL signal is strongly dependent on the orientation of the linear polarization direction of the excitation light with respect to average orientation of chemical bonds that follow for example the director n(r) of liquid crystals.
The schematic diagram [Figure (b)] shows that the Stokes beam is modulated with an electro-optic modulator (EOM, M350-160 KD*P, Conoptics) at a couple of MHz by using a square-wave function-generator (DS345, Stanford Research Systems). The modulation amplitude and DC offset of RF-driver (model 275, Conoptics) is adjusted to maximize the SRL signal. A large-area biased Si photodiode (75.4 mm2, DET100A, Thorlabs) is used for detection of the pump beam. The output from the photodiode passes through a low-pass filter to suppress the strong laser pulsing signal, before sending it to a high-frequency RF lock-in amplifier (SR844, Stanford Research Systems). The analog R (i.e. modulus) or x (i.e. in-phase component)-output of the lock-in amplifier is fed into a modified input of A/D converter.