Solid immersion microscopy images cells under cryogenic conditions with 12 nm resolution

Commun Biol. 2019 Feb 21:2:74. doi: 10.1038/s42003-019-0317-6. eCollection 2019.

Abstract

Super-resolution fluorescence microscopy plays a crucial role in our understanding of cell structure and function by reporting cellular ultrastructure with 20-30 nm resolution. However, this resolution is insufficient to image macro-molecular machinery at work. A path to improve resolution is to image under cryogenic conditions. This substantially increases the brightness of most fluorophores and preserves native ultrastructure much better than chemical fixation. Cryogenic conditions are, however, underutilised because of the lack of compatible high numerical aperture objectives. Here, using a low-cost super-hemispherical solid immersion lens (superSIL) and a basic set-up we achieve 12 nm resolution under cryogenic conditions, to our knowledge the best yet attained in cells using simple set-ups and/or commercial systems. By also allowing multicolour imaging, and by paving the way to total-internal-reflection fluorescence imaging of mammalian cells under cryogenic conditions, superSIL microscopy opens a straightforward route to achieve unmatched resolution on bacterial and mammalian cell samples.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATP-Binding Cassette Transporters / genetics
  • ATP-Binding Cassette Transporters / metabolism
  • Animals
  • CHO Cells
  • Cricetinae
  • Cricetulus
  • Cryoelectron Microscopy / methods*
  • Cytological Techniques / instrumentation*
  • Cytological Techniques / methods*
  • ErbB Receptors / chemistry
  • ErbB Receptors / genetics
  • ErbB Receptors / metabolism
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism
  • Fluorescent Dyes / chemistry
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Maleimides / chemistry
  • Microscopy, Fluorescence / methods*
  • Reproducibility of Results

Substances

  • ATP-Binding Cassette Transporters
  • Alexa Fluor 488 C5-maleimide
  • Escherichia coli Proteins
  • Fluorescent Dyes
  • Maleimides
  • McjD protein, E coli
  • Green Fluorescent Proteins
  • ErbB Receptors