Abstract
Here we report a simple way to enhance the resolution of a confocal scanning microscope under cryogenic conditions. Using a microscope objective (MO) with high numerical aperture (NA = 1.25) and 1-propanol as an immersion fluid with low freezing temperature we were able to reach an imaging resolution at 160 K comparable to ambient conditions. The MO and the sample were both placed inside the inner chamber of the cryostat to reduce distortions induced by temperature gradients. The image quality of our commercially available MO was further enhanced by scanning the sample (sample scanning) in contrast to beam scanning. The ease of the whole procedure marks an essential step towards the development of cryo high-resolution microscopy and correlative light and electron cryo microscopy (cryoCLEM).
Highlights
Light microscopy is an often used technique especially for investigating biological samples and in combination with spectroscopy for getting a deeper understanding of chemical and physical processes with high spatial resolution
A droplet (1 μl) of quantum dots (QD) solution diluted in H2O/glycerol with concentration 1 pmol/l to guaranty single particle microscopy was deposited between two 4 × 4 mm2 glass coverslips cleaned with chromosulfuric acid
We can claim that the lateral resolution of the immersed microscope objective (MO) under cryogenic conditions is enhanced by a factor of 1.3 with respect to our standard air objective with numerical aperture (NA) = 0.85
Summary
Light microscopy is an often used technique especially for investigating biological samples and in combination with spectroscopy for getting a deeper understanding of chemical and physical processes with high spatial resolution. 1957 [1] was a remarkable step in optical microscopy that increased the imaging contrast for gaining spatial information on luminescent samples. The principle of confocal microscopy relies on focusing a laser beam to a diffraction limited spot and recording light emerging mainly from that volume. In this technique out-of-focus light will be suppressed and increases the image contrast and enables to record 3D images of different structures such as biological cells by implementing a pinhole in front of the detector [2].
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
