Polarization-controlled Photo-switching Resolves Dipole Directions with Super-spatial resolution

DoctorWe demonstrate an imaging method that can resolve two fluorophores within a diffraction limited spot and simultaneously determine their dipole orientations. The intensity of the fluorophore emission can be modulated by the angle between the molecular transition dipole and the excitation polarization. The rotation of the linearly polarized incident light alternately switches the two fluorophores on and off, which offers another distinct photo-switching way and the molecular orientation as well. The polarization-rotating total internal reflection fluorescence microscopy was successfully applied for the 2D structure of DNA Holliday junction that is revealed by measuring the distance between Cy3 dyes stacked to its two ends with a ~10 nm spatial resolution and determining their in-plain dipole orientations. There are several advantages on this photo-switching super resolution microscopy utilizing the rotating polarization: 1) The imaging process is fast and simple since this technique is based on a non-stochastic control and the imaging by wide-field TIRF microscopy. 2) A super spatial resolution and a molecular orientation can be achieved simultaneously without the combination of two techniques. 3) The instruments in this technique are inexpensive compared to other super resolution microscopy (STED, PALM, STORM etc) that requires complicated and expensive instruments