[Histonet] Cytospin - misshapen cells

[Kemlo Rogerson]

Hi All,

 

I'm having some trouble with cytospins - the cells/nuclei look very
misshapen after spinning. I have used a number of different cell lines
and have the same problem with them all. I have tried fixing the cells
before spinning with 4% PFA in PBS (for this I centrifge the cells,
resuspend in PBS, centrifuge, resuspend in PFA for 10min on ice,
centrifuge, resuspend in PBS and cytospin). Unfixed cells were cytopsun
in media. The cytospin is set to 500rpm for 5min with low acceleration.
After spinning I let them air dry (~5min), wash PBS, fix 4% PFA (if not
post-fixed), wash PBS and continue with staining.

 

When staining the nuceli just with Dapi I get some very strange looking
nuclei, please see some pics on www.histonet.org the file is called
Cytospin Dapi.jpg.

 

Is this normal for cytospins? Any ideas?

 

Thanks

Sonya

 

Did they look 'normal' before? You mentioned 'cell lines' and from my
experience these look very abnormal when 'normal'. I remember carrying
out cytospins on 'grown' cells and the longer you 'grew' them the more
abnormal they looked.

 

Maybe you're looking at normal.

 

Kemlo Rogerson

Pathology Manager

DD   01934 647057 or extension 3311

Mob 07749 754194; Pager 07659 597107;

 

If the world seems cold to you, kindle fires to warm it. --Lucy Larcom 

 

 

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