[HISTONET] embedding cell cultures

[Peggy Bisher]

I have done just what you are talking about using Aclar. It is a plastic
embedding film (purchased from EMS). It works great for us.

Margaret E. Bisher
Electron Microscopy & Histology Core Facility Manager
Department of Molecular Biology
Princeton University
Moffett Laboratory, Room 113
Princeton, New Jersey
Office: (609) 258-7026
Fax: (609) 258-8468
mbisher <@t> princeton.edu





On 1/27/10 3:39 PM, "Sherwood, Margaret" <MSHERWOOD <@t> PARTNERS.ORG> wrote:

> Nick,
> 
> I am assuming that your 3D cells only grow on plastic.  They make plastic
> cover
> slips which, if your cells attach to them and grow, might make the embedding
> much easier.  You would follow the same method as stated, but then you could
> invert the coverslips on a beem capsule and separate the coverslip from
> capsule
> with liquid nitrogen.  However, I have never done it with plastic coverslips
> (only glass), so not sure if they would easily separate from capsule with
> liquid
> nitrogen. If anyone else has done so, please weigh in.
> 
> Peggy   
> 
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu
> [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Geoff
> McAuliffe
> Sent: Wednesday, January 27, 2010 3:20 PM
> To: Nicholas David Evans
> Cc: histonet <@t> lists.utsouthwestern.edu
> Subject: Re: [HISTONET] embedding cell cultures
> 
> 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
>> 
>> _______________________________________________
>> Histonet mailing list
>> Histonet <@t> lists.utsouthwestern.edu
>> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>> 
>> 
>>   
>