Fluorescence background suppression in Raman spectroscopy using combined Kerr gated and shifted excitation Raman difference techniques

An exceptionally high level of fluorescence rejection from resonance Raman spectra was achieved using a combination of two techniques, namely Kerr gated temporal rejection with shifted excitation Raman difference spectroscopy. The method was able to recover the resonance Raman spectrum from the intense fluorescence background with a signal-to-noise ratio at least 10 times higher than that achievable with either of the two approaches used individually. To demonstrate the effectiveness of the technique we obtained the resonance Raman spectrum of the laser dye rhodamine 6G (1 × 10−3 mol dm−3) in methanol by excitation at 532 nm and measuring under the maximum of fluorescence emission at 560–590 nm. The method reached the photon shot noise limit of the residual fluorescence providing a detection limit for Raman spectra 106 times lower than the original fluorescence intensity in an accumulation time of 800 s. A unique feature of the experiment was the way in which the optical parametric amplifier light source was configured to alternate automatically between the two excitation wavelengths using an optogalvanic mirror arrangement. The ultra-high sensitivity of the combined approach holds great promise for selective probing of complex biological systems using resonance Raman spectroscopy. Copyright © 2002 John Wiley & Sons, Ltd.