期刊
CHEMPHYSCHEM
卷 15, 期 4, 页码 587-599出版社
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cphc.201300880
关键词
fluorescence microscopy; molecular orientation; rotational mobility; single-molecule studies; super-resolution imaging
资金
- Robert and Marvel Kirby Stanford Graduate Fellowship
- National Science Foundation
- 3Com Corporation Stanford
- National Defense Science and Engineering Graduate Fellowship
- National Institute of General Medical Sciences [R01GM085437]
Numerous methods for determining the orientation of single-molecule transition dipole moments from microscopic images of the molecular fluorescence have been developed in recent years. At the same time, techniques that rely on nanometer-level accuracy in the determination of molecular position, such as single-molecule super-resolution imaging, have proven immensely successful in their ability to access unprecedented levels of detail and resolution previously hidden by the optical diffraction limit. However, the level of accuracy in the determination of position is threatened by insufficient treatment of molecular orientation. Here we review a number of methods for measuring molecular orientation using fluorescence microscopy, focusing on approaches that are most compatible with position estimation and single-molecule super-resolution imaging. We highlight recent methods based on quadrated pupil imaging and on double-helix point spread function microscopy and apply them to the study of fluorophore mobility on immunolabeled microtubules.
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