Super-resolution Microscopy at Cryogenic Temperatures Using Solid Immersion Lenses
Benji Bateman,
Laura Zanetti-Domingues,
Amy Moores,
Sarah Needham,
Daniel Rolfe,
Lin Wang,
David Clarke,
Marisa Martin-Fernandez
Affiliations
Benji Bateman
Central Laser Facility, Research Complex at Harwellcility, Science and Technology Facilities Council, Rutherford Appleton Laboratory, DidcotHarwell, Didcot, Oxford, OX11 0QX, UK
Laura Zanetti-Domingues
Central Laser FaCentral Laser Facility, Research Complex at Harwellcilitycility, Science and Technology Facilities Council, Rutherford Appleton Laboratory, DidcotHarwell, Didcot, Oxford, OX11 0QX, UK
Amy Moores
Central Laser FaCentral Laser Facility, Research Complex at Harwellcilitycility, Science and Technology Facilities Council, Rutherford Appleton Laboratory, DidcotHarwell, Didcot, Oxford, OX11 0QX, UK
Sarah Needham
Central Laser FaCentral Laser Facility, Research Complex at Harwellcilitycility, Science and Technology Facilities Council, Rutherford Appleton Laboratory, DidcotHarwell, Didcot, Oxford, OX11 0QX, UK
Daniel Rolfe
Central Laser FaCentral Laser Facility, Research Complex at Harwellcilitycility, Science and Technology Facilities Council, Rutherford Appleton Laboratory, DidcotHarwell, Didcot, Oxford, OX11 0QX, UK
Lin Wang
Central Laser FaCentral Laser Facility, Research Complex at Harwellcilitycility, Science and Technology Facilities Council, Rutherford Appleton Laboratory, DidcotHarwell, Didcot, Oxford, OX11 0QX, UK
David Clarke
Central Laser FaCentral Laser Facility, Research Complex at Harwellcilitycility, Science and Technology Facilities Council, Rutherford Appleton Laboratory, DidcotHarwell, Didcot, Oxford, OX11 0QX, UK
Marisa Martin-Fernandez
Central Laser Facility, Research Complex at Harwell, Science and Technology Facilities Council, Rutherford Appleton Laboratory, Harwell, Didcot, Oxford, OX11 0QX, UK
Our mechanistic understanding of cell function depends on imaging biological processes in cells with molecular resolution. Super-resolution fluorescence microscopy plays a crucial role 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, which substantially increases the brightness of most fluorophores and preserves native ultrastructure much better than chemical fixatives. Cryogenic conditions are, however, underutilized because of the lack of compatible high numerical aperture (NA) objectives. Here we describe a protocol for the use of super-hemispherical solid immersion lenses (superSILs) to achieve super-resolution imaging at cryogenic temperatures with an effective NA of 2.17 and resolution of ~10 nm.
