Mounting is a critical step in microscopy preparation procedures, aimed at preserving the sample and enhancing the image quality during observation. Proper mounting maintains the structural integrity of your specimen, reduces image distortions (like spherical aberration) caused by refractive index mismatch, and allows long-term storage.
The protocol
The protocol outlined below describes a common mounting process, ideal for most adherent monolayer cell types. As always, adjustments may be necessary depending on your specific experimental requirements.
1. Start by thoroughly rinsing your sample to remove excess salts and other residues.
2. Allow the specimen to completely dry, which is highly important for minimizing refractive index mismatch and spherical aberration. This typically takes at least 30 minutes. Please note that this is just one of many approaches, others may include procedures like a glycerol gradient transfer from water to the mounting medium, etc. Make sure to use an approach that best suits your specific experiment and sample type.
3. Prepare a clean microscope slide, and if using an adhesive mounting spacer, affix it onto the slide.
4. Apply an appropriate volume of mounting medium onto the slide. If using viscous mounting media, consider cutting the pipette tip for easier application.
5. Carefully remove the liner from the spacer, revealing its adhesive side.
6. Place the dried cover glass with your sample onto the spacer, making sure the cell side is facing down.
7. Press gently to secure the spacer and cover glass onto the slide.
8. The sample is now ready for microscopy. It’s resistant to typical cleaning methods and doesn’t require additional sealing.
For storage, keep the slide in a cool, dry, and dark place to prevent sample degradation.
Remember, these steps are part of a general protocol, so be sure to adapt as necessary for your specific needs and conditions.
In the demonstration video protocol, we employed the use of microscopic spacers AD-SEALS and the mounting medium AD-MOUNT S, both products of ADVI, Ricany, CZ.
