[Histonet] Cytospin - misshapen cells

[Martin S.]

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