Single-molecule imaging and spectroscopy of adenine and an analog of adenine using surface-enhanced Raman scattering and fluorescence

We implemented Raman imaging and spectroscopy of adenine molecules adsorbed on Ag colloidal nanoparticles to find feasibility of detecting and identifying nucleic-acid bases at the single-molecule level. Surface enhanced Raman scattering of adenine molecules showed an intermittent on-and-off behavior called “blinking” and fluctuation of the center of spectra called “spectral diffusion”. The observation of blinking and spectral diffusion provides substantial evidence for detecting single adenine molecules. Also, we extended single molecule fluorescence imaging and time-resolved fluorometry from green to violet-excitation regime to detect a fluorescent analog of nucleic-acid bases at the single-molecule level. Using violet excitation we observed fluorescence spectra and lifetimes from single complexes of an analog of adenine and the Klenow fragment of DNA polymerase I.