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protocol for immunostaining primery cells

confocal microscopy immunostaining

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5 replies to this topic

#1 hanna12345

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Posted 07 February 2012 - 05:05 AM

I want to stain primery cells in suspension for cofocal microscopy.
(until today I had no problems with immunostaining cell which were grown on coverslips, but now it's more complicated situation because the cells are much more sensitive)

help?

thanks
Hanna

#2 scolix

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Posted 07 February 2012 - 09:23 AM

did you try coating coverslips and then plating the cells and staining them.

#3 hanna12345

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Posted 07 February 2012 - 11:12 PM

did you try coating coverslips and then plating the cells and staining them.


No.
can you explain how can I do that?
I do tried to force the cells to be stuck on slides which have some kind of coat, but it didn't work.
anyway, I want to emphasize that I need to stuck them but not to let them grow on the coverslip. does your advice to coat the coverslip anable to stuck the cells strong enough that when I wash them, the cells will not washed away?

thanks

#4 scolix

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Posted 07 February 2012 - 11:41 PM

You can coat coverslips with poly-lysine or collagen or something else. This helps cells to be stuck to the coverslips. But it will not stop them from growing.
Now, if you stain them, they should stick better. Also use PBS with calcium and magnesium, it also helps to keep cells attached.

#5 hanna12345

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Posted 09 February 2012 - 01:45 AM

So if I understand you, I can do all this process of sticking and after that to stain the cells?

#6 scolix

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Posted 09 February 2012 - 10:26 AM

yes. If the cells stick better, then they should be able to withstand the staining process. Else you will lose them.

Good Luck !!!





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