Hello, I have stem cells in culture in a plastic flask and I want to observe it in a Scanning Electron Microscope (SEM), i want to know if there is a special preparation to the culture whitout using glass slides or scaffolds?
thanks
Hello Javi,
There is no special preparation. Cells grown on glass coverslips are perfect to use for SEM. The fixation is in 4-5% gluteraldehyde (the EM fixative), fresh or stored for one week maximum. However, you cannot observe them directly from flask.
Hope it helps!
Ajay
As Ajay mentioned, you have to cross linked them with glutaraldehyde. I normally do it with 2% (w/v) in pbs.
remove the medium, add the glutaraldehyde on the cells and leave them like that for 2 hours.
Remove the glutaraldehyde and wash gently with pbs. Then add alcohol 100% and leave it for one 1hr.
Then remove the alcohol and add HMDS (hexamethyldisilazane) to dry the samples overnight until the chemical dries out completely and once it is dried, and the samples are finally ready and completely dried.
Now, before SEM, you have to Sputer coat them with 5nm of platinium or gold for conductivity. And now, you can finally observe them under SEM
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