A novel lens assembly enabling super resolution imaging of biological samples under cryogenic conditions

About

STFC are looking to develop relationships with: • Microscope manufacturers • Microscope component manufacturers • Microscope stage manufacturers • Research groups/companies interested in using SuperSIL for research Cell immoblisation is required to carry out structural studies using super-resolution fluorescence microscopy. This is currently achieved using chemical fixation at room temperature. This technique can present multiple problems, such as the introduction of artifacts as well as insufficient protein fixation. These problems can be completely avoided by cooling cells to cryogenic temperatures, causing immobilisation without the need for chemical fixation. Imaging under cryogenic conditions also significantly increases the brightness of most fluorophores and allows cells to be imaged in their native states. However, super-resolution microscopy currently requires the use of a state of the art oil or water immersion lenses, which are incompatible with cryogenic conditions. The Lasers for Science team at STFC have developed and patented SuperSIL, a novel ASIL (aplanatic solid immersion lens) which removes the need for an oil or water immersion lens, enabling super-resolution cryogenic fluorescence microscopy (SR cryo-FM). The SuperSIL lens assembly has been successfully deployed on a modified commercial microscope system with a low/medium NA air objective. Resolutions up to 12nm have been demonstrated using single molecule localisation microscopy (SMLM) under cryogenic conditions, which is considered to be a game changer in cryo-FM. In comparison, a high NA air objective lens used in typical cryo-FM gives a resolution of approximately 400nm.

Key Benefits

• Perform super-resolution cryo-FM to achieve up to 12nm resolution compared with 400nm offered by conventional cryo-FM • Achieve a 120nm theoretical resolution if not using super-resolution techniques • Avoid artefacts associated with conventional super-resolution FM images • Image cells in their native states • Significantly increase the brightness of most fluorophores compared to room temperature FM • Reduce photobleaching compared to room temperature FM • Potential enabler for future advances in cryogenic correlative light electron microscopy (cryo-CLEM)

Applications

Enabling novel studies for life sciences applications in: • Cell and developmental biology • Microbiology • Biomedical science • Molecular biology and biotech • Structural biology • Synthetic biology Other potential uses: • Semiconductor defect inspections

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