Publikationen des Max-Planck-Instituts für die Physik des Lichts

Two decades ago it was commonly believed that single molecules were too small to be seen using an optical microscope. Today, single-molecule fluorescence microscopy has become a standard tool in many biology laboratories. For example, the diffusion or directed motion of proteins labeled with a dye molecule can be detected and tracked by the fluorescent emission of the dye. An advantage of this approach is its specificity; each dye molecule emits at a particular wavelength, enabling the excitation light to be filtered out spectrally. Moreover, one can even count individual photons of fluorescence on a zeroed background. However, organic dye molecules typically used in these processes are prone to photobleaching after ∼1min of illumination. The technique also relies on the high fluorescent quantum yield of the target. Recently, there has been increasing interest in detecting single molecules that do not fluoresce. Here, we image single molecules directly in transmission mode using optics with very high numerical apertures.