Backward multiplex coherent anti-Stokes Raman (CARS) spectroscopic imaging with electron-multiplying CCD (EM-CCD) camera

A multiplex CARS imaging system, equipped with an EM-CCD camera, was developed to improve the sensitivity of backward CARS imaging in biological analysis using an inverted microscope. The signal-to-noise ratio was improved by a factor of ca. 3 compared to a conventional CCD mode through the use of EM gain. When imaging cultured epithelial cells in the backward CARS configuration, intracellular organelles such as lipid droplets and nuclei were spectroscopically identified with an exposure time of only 100 ms/pixel.