Center for Relativistic Laser Science

124th Summer Symposium of KCS-Physical Chemistry Division

Single-molecule localization method is commonly used to construct a super-resolution image or to track a molecule of interest. The fitting accuracy of PSF to the image of fluorescence emission from a single molecule, which is the most frequently used method to find a molecule’s position, highly depends on the intensity of fluorescence and suffers from pixel-to-pixel variation of gain and noise of the camera. Thus the spatial resolution of single-molecule localization microscopy usually exceeds 10 nm. In this study, we developed a novel approach to localize a single molecule by illuminating it with a stripe-patterned light. When we modulated the phase of the illumination pattern, the total amount of emitted photons from a single molecule varied in proportion to the excitation intensity on the exact position of the molecule. Since the noisy features from pixel-to-pixel variation should be averaged during integrating the photons, our approach can provide better spatial resolution than conventional methods, especially under a low signal-to-noise ration condition.

References

[1] Sengupta, P.; Engelenburg, S. B.; L.-Schwartz, J. Superresolution imaging of biological systems using photoactivated localization microscopy. Chem. Rev. 2014. 114, 3189-3202.

[2] Shao, L.; Rego, E. H.; Gustafsson, M. G. Super-resolution 3D microscopy of live whole cells using structured illumination. Nat. Methods 2011. 8, 1044-1046.