I want to fix biological cells on metal substrate using Glutaraldehyde, so is it possible to store the samples for SEM imaging? Would this affect the samples? And what is the max. period of time and what is the optimum storage conditions?
Glut does not provide "immediate penetration", but for single layer cell culture it works pretty fast, but still I would not recommend to have fixation time shorter than 0.5 hrs (1.5 hrs for thicker specimens). Generally speaking, with some exceptions, specimens can be stored in glut fixative for as long as at least one month in refrigerator. But why would you want to do it? Dehydrate, mount and coat you specimens and you can store them for years (preferably in desiccator). If you cells produce mineral, be aware that in can be partially dissolved during storage in water solutions.
Gluteraldehye provides immediate penetration for fixation of samples prepared for electron microscopy. Yet, it causes over dehydration if samples emersed for more than 24 hours. Therefore, it is recommended to fix in gluteraldehyde overnight , then transfer to 4-formaldehyde -1- Gluteraldehyde fixation mixture for maximum 72 hours . This should be followed by applying further processing protocol for SEM.
Glut does not provide "immediate penetration", but for single layer cell culture it works pretty fast, but still I would not recommend to have fixation time shorter than 0.5 hrs (1.5 hrs for thicker specimens). Generally speaking, with some exceptions, specimens can be stored in glut fixative for as long as at least one month in refrigerator. But why would you want to do it? Dehydrate, mount and coat you specimens and you can store them for years (preferably in desiccator). If you cells produce mineral, be aware that in can be partially dissolved during storage in water solutions.
I agree with Vladimir Dusevich: " Dehydrate, mount and coat you specimens and you can store them for years (preferably in desiccator)." Go on.
Keep your sample in a container with Silica gel or any other moisture absorbent materials. Be careful to completely close the container to prevent gase/moisture transmission. In this condition, I could store the samples more than two months, then took some great SEM micrographs. I have not followed for more than two months. I've heard that in this condition, you may can store the samples for 1 year, not sure.
Good luck
Well, some researches suggest that you may keep the samples stored in the fixative for some time under certain conditions. Please refer to the following discussion:
https://www.google.com.eg/amp/s/www.researchgate.net/post/How_long_we_could_keep_a_glutaraldehyde_fixed_sample_for_sem/amp
The idea of keeping the samples in the fixative is based on the belief that this may prevent any bacterial contamination. However, as any immersion process, this may dissolve or influence surface deposits (this would be a problem if you are interested to see these deposits in the SEM.
However, my opinion is to finish the fixation process, rinse and dry the samples, sputter coat them with gold then you can store them in a dark air-tight container with silica gel for months before SEM examination
I do agree with Dr. Vladimir Dusevich. You can store for short periods in Glut. But it is a molecule that have a low penetration rate… captured to PFA, for example. If your focus is to storage your sample for long periods (i.e. years) follow Dr. Dusevich recommendation. Be careful to avoid loss information that is deposit on the surface of the biological samples. I am used to work on plant pathogens and long fixation periods wash the pathogen from the surfaces.
Good luck
João
Thanks a lot everyone!
For your information, the Glut fixed samples stored for like 12 hours, their cells were very clear and good. However when the storage exceeds this period, the cells looked like solid particles on the surface.
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