Single-molecule orientation localization microscopy I: fundamental limits

Precisely measuring the three-dimensional position and orientation of individual fluorophores is challenging due to the substantial photon shot noise in single-molecule experiments. Facing this limited photon budget, numerous techniques have been developed to encode 2D and 3D position and 2D and 3D orientation information into fluorescence images. In this work, we adapt classical and quantum estimation theory and propose a mathematical framework to derive the best possible precision for measuring the position and orientation of dipole-like emitters for any fixed imaging system. We find that it is impossible to design an instrument that achieves the maximum sensitivity limit for measuring all possible rotational motions. Further, our vectorial dipole imaging model shows that the best quantum-limited localization precision is 4%-8% worse than that suggested by a scalar monopole model. Overall, we conclude that no single instrument can be optimized for maximum precision across all possible 2D and 3D localization and orientation measurement tasks. (C) 2021 Optical Society of America

Automated summary

This study proposes a mathematical framework to derive the best possible precision for measuring the position and orientation of dipole-like emitters in any fixed imaging system. It is found that designing an instrument to achieve the maximum sensitivity limit for measuring all possible rotational motions is impossible. Additionally, the best quantum-limited localization precision is shown to be 4%-8% worse than suggested by a scalar monopole model through a vectorial dipole imaging model.