Dear Histonet,

The following recommendation is intended to clarify questions regarding
proper use of HercepTest Antigen Retrieval in the water bath.  Use a water
bath that can maintain a temperature between 95º and 99ºC.  For expediency,
preheat the Antigen Retrieval solution provided with HercepTest in Coplin
jars. This may be done in the water bath or by using a microwave as long the
slides are NOT in the solution at this time.  Place the Coplin jars
containing the preheated Antigen Retrieval solution in the water bath and
allow it to equilibrate to at least 95ºC, but not above 99ºC.  You must
measure the temperature of the antigen retrieval solution itself to ensure
it has reached 95-99ºC.  At this point place the deparaffinized and
rehydrated slides in the antigen retrieval solution.  Once the antigen
retrieval solution in the water bath has reached 95-99ºC with the slides,
start the timer for 40 minutes of retrieval.  Set the waterbath temperature
to 97º or 98ºC so you have a little leeway with temperature fluctuations
that occur while opening and closing the waterbath lid.

If you have any questions please contact DAKO Technical Services at 800 424
0021.
Sincerely,
Danielle Mach
DAKO Corporation
Technical Service Specialist
(800)424-0021

-----Original Message-----
From: terij@prlnet.com [mailto:terij@prlnet.com]
Sent: Wednesday, September 06, 2000 6:11 AM
To: 'Histonet'
Subject: Re: Hercep help


Abizar,

You are correct about this.  However, I too was having trouble with the
plastic jars reaching the proper temperature and called Dako's technical
support.  They were the ones who instructed me to bring it up to temperature
using a microwave and then allow the slides to incubate further in the
waterbath. So if this isn't correct, we need to hear it from Dako Corp.

-Teri Johnson
Supervisor, Physicians Reference Laboratory
Overland Park, KS

----- Original Message -----
From: Abizar Lakdawalla <abizarl@innogenex.com>
To: Sebree Linda A. <la.sebree@hosp.wisc.edu>
Cc: 'Louise Taylor' <louiset@mail.saimr.wits.ac.za>; 'Histonet'
<histonet@pathology.swmed.edu>
Sent: Tuesday, September 05, 2000 2:11 PM
Subject: Re: Hercep help


> Just a comment: Changes in protocol for FDA tests are usually not allowed.
You
> have to follow the intructions to the dot.
> abizar
>
> "Sebree Linda A." wrote:
>
> > Hi Louise,
> >
> > When we use the waterbath for HIER, we first bring the buffer and slides
(in
> > a plastic coplin jar) up to a near boil in a microwave oven and then
> > transfer it to the waterbath and start our timing.  This works very well
for
> > us and is our most gentle form of HIER that we use.
> >
> > Linda A. Sebree, HT
> > University of Wisconsin Hospital & Clinics
> > Immunohistochemistry/In Situ Hybridization Laboratory
> > D4/218-2472
> > 600 Highland Avenue
> > Madison, WI  53792-2472
> >
> > (608)265-6596
> > FAX: (608)263-1568
> >
> > > -----Original Message-----
> > > From: Louise Taylor [SMTP:louiset@mail.saimr.wits.ac.za]
> > > Sent: Monday, September 04, 2000 10:08 AM
> > > To:   histonet
> > > Subject:      Hercep help
> > >
> > > HI histonetters
> > >
> > > I need some help urgently!  I am embarking on  a  hercep trial and
have
> > > run
> > > into an unexpected problem. What do you use to put your slides in to
> > > "boil"
> > > them for antigen retrieval? I tried using a glass coplin jar in my
> > > waterbath, but it shattered, and when I used a plastic one, I noticed
that
> > > the internal solution temperature did not go above 82 degrees. (I have
> > > never
> > > been required to use a waterbath for Ag retrieval, having quite
happily
> > > used
> > > a slide rack and a beaker in the microwave - but this sytem won't
really
> > > work, as it uses quite a lot of buffer).
> > >
> > > Any suggestions will be welcome, I have about 100 slides to do in the
next
> > > few days before I go on honeymoon!!
> > >
> > > Many thanks
> > >
> > > Louise Taylor
> > >
> > >
> > > Research Laboratory
> > > Department of Anatomical Pathology
> > > South African Institute for Medical Research
> > > Johannesburg
> > > South Africa
> > >
> > >
> > >
>
>