Simultaneous forward and epi-CARS microscopy with a single detector by timecorrelated single photon counting

Abstract

We present a novel scheme to simultaneously detect coherent anti-Stokes Raman scattering (CARS) microscopy signals in the forward (F) and backward (epi - E) direction with a single avalanche photodiode (APD) detector using time-correlated single photon counting (TCSPC). By installing a mirror at a well-defined distance above the sample the forward-scattered F-CARS signal is reflected back into the microscope objective leading to spatial overlap of the F and E-CARS signals. Due to traveling an additional distance the F-CARS signal is time delayed relative to the E-CARS signal. TCSPC then allows for the two signals to be resolved in the time domain. This results in an efficient, simple, and compact method of CARS signal detection. We demonstrate this technique by analyzing forward and backward CARS signals obtained by imaging living adipocyte cells derived from human mesenchymal stem cells.