Open AccessThree-dimensional adaptive optical nanoscopy for thick specimen imaging at sub-50-nm resolution
Author(s) -
Xiang Hao,
Edward S. Allgeyer,
DongRyoung Lee,
Jacopo Antonello,
Katherine Watters,
Julianne A. Gerdes,
Lena K. Schroeder,
Francesca Bottanelli,
Jiaxi Zhao,
P. Kidd,
Mark D. Lessard,
James E. Rothman,
Lynn Cooley,
Thomas Biederer,
Martin J. Booth,
Joerg Bewersdorf
Publication year - 2021
Publication title -
nature methods
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 19.469
H-Index - 318
eISSN - 1548-7105
pISSN - 1548-7091
DOI - 10.1038/s41592-021-01149-9
Subject(s) - resolution (logic) , optics , adaptive optics , microscopy , image resolution , diffraction , optical microscope , fluorescence , microscope , materials science , isotropy , sted microscopy , fluorescence microscope , super resolution microscopy , physics , stimulated emission , computer science , scanning electron microscope , laser , artificial intelligence
Understanding cellular organization demands the best possible spatial resolution in all three dimensions. In fluorescence microscopy, this is achieved by 4Pi nanoscopy methods that combine the concepts of using two opposing objectives for optimal diffraction-limited 3D resolution with switching fluorescent molecules between bright and dark states to break the diffraction limit. However, optical aberrations have limited these nanoscopes to thin samples and prevented their application in thick specimens. Here we have developed an improved iso-stimulated emission depletion nanoscope, which uses an advanced adaptive optics strategy to achieve sub-50-nm isotropic resolution of structures such as neuronal synapses and ring canals previously inaccessible in tissue. The adaptive optics scheme presented in this work is generally applicable to any microscope with a similar beam path geometry involving two opposing objectives to optimize resolution when imaging deep in aberrating specimens.