[HISTONET] embedding cell cultures

[Geoff McAuliffe]

Hi Nick:

You can use the Epon substitutes such as EmBed 812. Fix, osmicate, 
dehydrate as usual, but omit the proplylene oxide as it will react with 
the plastic dish. Epon substitutes will mix with ethanol. I used 2:1 
then 1:1 then 1:2 ratios of absolute ethanol to "Epon" with catalyst 
added with agitation. Then several changes of pure Epon and polymerize. 
Yes, you will have to saw away the plastic, if you try to section the 
Epon-plastic combo the two will separate.
How much easier do you want it to be?

Geoff

Nicholas David Evans wrote:
> Dear all,
>
> I was hoping someone might be able to offer me some advice on embedding and sectioning cell cultures.
>
> In short we are growing cells which form 3D dome-like structures on tissue culture plastic. Does anyone have any experience or advice to offer on embedding the cultures in situ before sectioning? I have seen various methods in the literature, which often use Epon to embed the material followed by sawing away the plastic, but if anyone can offer some tips on other possible (easier) ways of doing it, or can refer me to some useful literature, I'd be very grateful.
>
> I would like to have simple 10um sections at 90 degrees to the substrate, which I can use for IHC.
>
> Best wishes
> Nick
>
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Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
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mcauliff <@t> umdnj.edu
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