[Histonet] Signal amplification

[Kim Merriam]

We use Perkin Elmer for biotinyl tyramide and invitrogen for fluroescent tyramide amp (they sell several kits with differnt flourochromes, including streptavidin TSA).


Kim Merriam, MA, HT(ASCP)QIHC Cambridge, MA


________________________________
From: Andrea Hooper <andreahooper <@t> rocketmail.com>
To: "histonet <@t> lists.utsouthwestern.edu" <histonet <@t> lists.utsouthwestern.edu> 
Sent: Wednesday, January 25, 2012 5:26 PM
Subject: [Histonet] Signal amplification

What kits/vendors are people using for their CSA/tyramide amplification protocols? 

Thanks in advance!
Andrea

On Jan 25, 2012, at 3:45 PM, "Mayer,Toysha N" <TNMayer <@t> mdanderson.org> wrote:

> Also, to help cut the tissues, I face at room temperature (very slowly), then I place my uterus/prostate blocks on ice with lots of water, then I cut them last. This helps to allow the blocks to soak up the water and soften some, plus allows nice thin sections.
> 
> I do agree with Andi that the processing procedure is too long with too many 100% alcohols. 
> 
> Toysha N. Mayer, MBA, HT (ASCP)
> Instructor, Education Coordinator
> Program in Histotechnology
> School of Health Professions
> MD Anderson Cancer Center
> (713) 563-3481
> tnmayer <@t> mdanderson.org
> 
> 
> 
> ------------------------------
> 
> Message: 16
> Date: Wed, 25 Jan 2012 07:22:07 -0800
> From: "Grantham, Andrea L - (algranth)" <algranth <@t> email.arizona.edu>
> Subject: Re: [Histonet] Reg: Tissue is hard to cut after processing
> Cc: "histonet <@t> lists.utsouthwestern.edu"
>    <histonet <@t> lists.utsouthwestern.edu>
> Message-ID: <D5CBD3E4-5447-4F24-964C-19BF8144EDF9 <@t> email.arizona.edu>
> Content-Type: text/plain; charset="us-ascii"
> 
> Arun,
> It looks like you are overprocessing your tissues. Uterus and prostate are tissues that are often hard to cut and your schedule for processing may be adding to your difficulties.
> Unless the pieces are very large and thick (and you can't do anything about that), cut down on the time, eliminate the heat and vacuum in your alcohols and xylenes but not the paraffins. You might also want to eliminate one of the 100% alcohols and one of the xylenes or try a xylene substitute like Clear-Rite 3, which is a more gentle clearing agent. If you eliminated one of these you could add another 70% or 80% alcohol before the 95% to help wash out the formalin. Right now you are really drying out your tissues and making them harder.
> If your tissues aren't totally fixed you may need the time in Formalin but if they are well fixed you could cut these times also.
> You probably don't need 4 hrs in paraffin at 63 degrees either. I'd cut the time down and the temp. to at most 60 degrees. I usually go 3-4 degrees above the melting point of the paraffin.
> 
> Andi Grantham
> 
> On Jan 25, 2012, at 6:25 AM, Arun Jyothi S.P wrote:
> 
>> Dear all,
>> 
>> We are having a hard time in cutting blocks. All our blocks especially
>> uterus, prostate chips etc became very hard after processing, we cant even
>> trim the blocks its very hard.
>> we use SLEE MTM automated tissue processor
>> and our schedule is
>> 1, Formalin                            - 1hr          - ambient  - 40 degree
>> 2, Formalin                            - 2 hr          - vacuum - 40 degree
>> 3, 70% ethanol                      - 1hr          - vacuum  - Room temp
>> 4, 95% ethanol                      - 1hr          - vacuum  - Room temp
>> 5 and 6- 100% ethanol            - 1hr each  - vacuum - room temp
>> 7, 100% ethanol                    - 1.30 hr      - vacuum  -  room temp
>> 8, 9 and 10 - Xylene                - 1 hr each  - vacuum  - 30, 35, 40
>> degree respectively
>> 11,12,13 and 14 - paraffin wax  - 1 hr each  - vacuum -  63 degree for all.
>> 
>> We use Merck paraffin wax of 56 degree M.P
>> 
>> Is there any problem with tissue processing schedule or something else is
>> wrong?
>> 
>> We are in a big problem really expecting some valuable suggestion and
>> advices.
>> 
>> with regards
>> 
>> 
>> 
>> ARUN JYOTHI S.P.
>> Histotechnologist
>> United Laboratories Co. <http://www.unitedlabs.com.kw/>
>> Kuwait
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> 
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