Dynamics of single dye molecules observed by confocal imaging and spectroscopy

The fluorescence emission of single rhodamine dye molecules (rhodamine 6G and rhodamine 630) at room temperature was analyzed by using scanning confocal laser microscopy in conjunction with polarization analysis, fluorescence spectroscopy, time-resolved detection (minutes to microseconds), and excitation saturation. Results are presented and discussed 1) for samples with dye molecules at the glass-air interface and 2) covered with an additional thin protective polymer film (polyvinylbutyral). Under the polymer layer, the single-molecule fluorescence was more stable than the glass-air interface. This result may be explained by fewer spontaneous variations of the fluorescence rate, polarization changes, spectral shifts, and longer photochemical lifetimes.