[Histonet] Paraffin Tissue Crumbles

[Nails, Felton]

I agree most of your times are too long, but you can still get sections if you put water on your ice block and allow the tissue to rehydrate a bit or put a little ammonium hydroxide and water on your ice and sit your blocks to be section in this solution. 

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Merced M Leiker
Sent: Wednesday, November 10, 2010 9:17 AM
To: Michael Mashore; histonet <@t> lists.utsouthwestern.edu
Subject: Re: [Histonet] Paraffin Tissue Crumbles

Having some experience processing and embedding rodent tissues myself (by hand), I would say that you are over-dehydrating the tissues. Try cutting back the alcohol incubation times to 30 min or even 10 min each.

Regards,
Merced

--On Tuesday, November 09, 2010 2:56 PM -0800 Michael Mashore <mmashore <@t> vapop.ucsd.edu> wrote:

> Hello Histonet Users,
>
>
>
> I have just started using paraffin and am having many difficulties. 
> Most of the time my tissue crumbles when sectioning. I have no real 
> experience in paraffin histology and have been given the task of 
> becoming proficient by myself, so I am hoping for feedback as to why 
> my tissue keeps crumbling. The tissue in question has been:  skeletal 
> muscle, cardiac muscle, liver, and brain (all from rat).
>
>
>
> The tissue was fixed in 10% neutral buffered formalin for 7 days at 
> 4°C and then transferred to an automated tissue processor, with the 
> following
> schedule:
>
>
>
> 2 hours 70% dehydration alcohol
>
> 2 hours 80% dehydration alcohol
>
> 2 hours 95% dehydration alcohol
>
> 2.5 hours 95% dehydration alcohol
>
> 2 hours 100% dehydration alcohol
>
> 2 hours 100% dehydration alcohol
>
> 2 hours 100% dehydration alcohol
>
> .5 hour Hemo-De
>
> .5 hour Hemo-De
>
> .5 hour Hemo-De
>
> 1 hour paraffin
>
> 4 hours paraffin
>
>
>
> They were infiltrated for 1 hour without vacuum then embedded.
>
> The blocks were stored in the freezer before cutting.
>
> The knife angle was 5°.
>
> Sections were 5µm thick.
>
>
>
> I would appreciate any feedback whatsoever.
>
>
>
> Thank you very much.
>
>
>
> Michael
>
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>



Merced M Leiker
Research Technician III
Cardiovascular Medicine
348 Biomedical Research Building
State University of New York at Buffalo
3435 Main St, Buffalo, NY 14214  USA
leiker <@t> buffalo.edu
716-829-6118 (Ph)
716-829-2665 (Fx)

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