[Histonet] How to do an EBER on cytological material?

Hoekert, W.E.J. W.E.J.Hoekert <@t> olvg.nl
Thu Mar 4 09:52:01 CST 2010

Dear histonetters,
I was wondering if it is possible to do an EBER ISH on a cell-smear. What pretreatment should I use?
1: Fix the cells in formalin (let's say overnight, or else one or two hours), and than use the proteinase K digestion step.
2: Or maybe it is enough to fix the cells in cold acetone, and skip the proteinase K step? Or will the RNA leak out of the cells?
Does anybody has experience whit this? We only have two slides left, and we don't have (cytological) control material, only FFPE control material. Also, I think that we should bake the slides prior to the asessment because we might lose a lot of cells during the (stringent) washing steps. 
Maybe it is not possible at all because the RNA is broken down allready (the cell smears were taken 3 days ago, they were stored at room temperature).
I hope you folks can help me out.
Willem Hoekert
OLVG, The Netherlands


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