Cryo-Structured Illumination Microscopic Data Collection from Cryogenically Preserved Cells | Zendy

Nina Vyas | Zendy; Nina Perry | Zendy; Chidinma Okolo | Zendy; Ilias Kounatidis | Zendy; Thomas M. Fish | Zendy; Kamal L. Nahas | Zendy; Archana Jadhav | Zendy; Mohamed A. Koronfel | Zendy; J. Groen | Zendy; Eva Pereiro | Zendy; Ian M. Dobbie | Zendy; Maria Harkiolaki | Zendy

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Cryo-Structured Illumination Microscopic Data Collection from Cryogenically Preserved Cells

Author(s) -

Nina Vyas,

Nina Perry,

Chidinma Okolo,

Ilias Kounatidis,

Thomas M. Fish,

Kamal L. Nahas,

Archana Jadhav,

Mohamed A. Koronfel,

J. Groen,

Eva Pereiro,

Ian M. Dobbie,

Maria Harkiolaki

Publication year - 2021

Publication title -

journal of visualized experiments

Language(s) - Uncategorized

Resource type - Journals

SCImago Journal Rank - 0.596

H-Index - 91

ISSN - 1940-087X

DOI - 10.3791/62274

Subject(s) - microscopy , fluorescence microscope , resolution (logic) , synchrotron , microscope , beamline , electron microscope , biological specimen , materials science , fluorescence lifetime imaging microscopy , fluorescence , optics , computer science , physics , artificial intelligence , beam (structure)

Three-dimensional (3D) structured illumination microscopy (SIM) allows imaging of fluorescently labelled cellular structures at higher resolution than conventional fluorescence microscopy. This super-resolution (SR) technique enables visualization of molecular processes in whole cells and has the potential to be used in conjunction with electron microscopy and X-ray tomography to correlate structural and functional information. A SIM microscope for cryogenically preserved samples (cryoSIM) has recently been commissioned at the correlative cryo-imaging beamline B24 at the UK synchrotron. It was designed specifically for 3D imaging of biological samples at cryogenic temperatures in a manner compatible with subsequent imaging of the same samples by X-ray microscopy methods such as cryo-soft X-ray tomography. This video article provides detailed methods and protocols for successful imaging using the cryoSIM. In addition to instructions on the operation of the cryoSIM microscope, recommendations have been included regarding the choice of samples, fluorophores, and parameter settings. The protocol is demonstrated in U2OS cell samples whose mitochondria and tubulin have been fluorescently labelled.

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