RE: [Histonet] Citrate Buffer pH

From:"Steven E. Slap"

RE: [Histonet] Citrate Buffer pH
Hi HistoNetters

As Dr. Shi points out in his book and articles about antigen retrieval, there are three parameters involved in achieving  consistent results:

1) temperature
2) time
3) pH

How you get there (steamer, water bath, microwave, pressure cooker) doesn't matter, so long as you can successfully monitor these three variables very accurately.

As Jeff Gordon correctly points out, some antigens need temperatures higher than 98C for optimal retrieval, however.  For this reason, pressure cookers, whether by themselves, or inside of a microwave have been used to reach temperatures as high as 120C as needed.  Using a combination microwave/pressure method, I have been able to use citrate buffer at pH 6.0 to retrieve most antigens at 110C in 3-5 minutes.

best regards,
Steven Slap

At 3:53 PM -0500 12/3/03, Houston, Ronnie wrote:
Patsy,
 
We routinely use Reveal and Borg, both from Biocare Medical, with great success in  a B&D steamer, 20 minutes, followed by 2 rinses in hot buffer, and 20 minute cool down. Very rarely use pressure cooker, and don't use micrwave at all.
 
Ronnie
 
Ronnie Houston
Regional Histology Operations Manager

Bon Secours HealthPartners Laboratories

5801 Bremo Road

Richmond, VA 23226

(804) 287 7972

ronnie_houston@bshsi.com
 
-----Original Message-----
From: Patsy Ruegg [mailto:pruegg@colobio.com]
Sent: Wednesday, December 03, 2003 1:19 PM
To: Jackie.O'Connor@abbott.com; Pat Flannery
Cc: Histonet@lists.utsouthwestern.edu; histonet-admin@lists.utsouthwestern.edu
Subject: RE: [Histonet] Citrate Buffer pH

Has anybody tried the AR reagents that deparaffinzed such as BioCare Decloaker in a steamer in place of the pressure cooker?  PC is too harsh for some of my stuff but I like using these retrieval/deparaffination reagents.
Patsy
 
-----Original Message-----
From: histonet-admin@lists.utsouthwestern.edu [mailto:histonet-admin@lists.utsouthwestern.edu]On Behalf Of Jackie.O'Connor@abbott.com
Sent: Tuesday, December 02, 2003 2:50 PM
To: Pat Flannery
Cc: Histonet@lists.utsouthwestern.edu; histonet-admin@lists.utsouthwestern.edu
Subject: Re: [Histonet] Citrate Buffer pH
 

I've been using a product from Biocare, a Universal Decloaker (6.0 citrate buffer) , which deparaffinizes during HIER in their pressure cooker (Decloaking Chamber) - they are a very Trekkie oriented company.  It works great - saves a bunch o' steps.  You might try contacting them for the specifics.  I don't think about the nature of these things if they work - I just follow along blindly . . . . . . . . .sometimes.

Jacqueline M. O'Connor HT(ASCP)
Abbott Laboratories
Global Pharmaceutical Research and Development
Discovery Chemotheraputics

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