Re: Ag retreival vs. Permeabilizing

From:Carrie Kyle-Byrne

hold on....i've worked with novel antibodies (monoclonal) in FFPE that have
responded to saponin treatment better than harsher digestions (ie pepsin and
trypsin) while giving no staining in sections that received no pretreatment.
i haven't got a clue what it does.....just that it works better than the
myriad of other things i tried.

Carrie Kyle-Byrne, BHS, HT(ASCP)
Assoc. Research Scientist II
Molecular Target Research

Exelixis, Inc.
170 Harbor Way
P.O. Box 511
South San Francisco
CA 94083-0511 USA

Phone: (1 650) 837-8023
Fax: (1 650) 837-7240
Email: ckbyrne@exelixis.com

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----- Original Message -----
From: "Greg Dobbin" 
To: "Morken, Tim" 
Cc: 
Sent: Tuesday, August 06, 2002 11:04 AM
Subject: RE: Ag retreival vs. Permeabilizing


> Hi Tim,
> I'm with you, that's exactly what I thought as well. But these guys
> were working on formalin-fixed, paraffin-embedded tissues!?
>
> So I'm thinking the permeabilizing step is not necessary. I'm going
> to try it without any treatment and perhaps with a heat retreival
> method and compare results. Thanks for the reply.
> Greg
>
> Date sent:      Tue, 06 Aug 2002 13:41:45 -0400
> From:           "Morken, Tim" 
> Subject:        RE: Ag retreival vs. Permeabilizing
> To:             'Greg Dobbin' ,
histonet@pathology.swmed.edu
>
> > Detergents such as Triton and saponin "permeabilize" cell membranes by
> > making holes in them. This is a physical process that simply makes a
"door"
> > for the antibody to enter the cell. Usually this is done on whole-cell
preps
> > such as cell smears and cytospins in which the cell membrane is usually
> > intact. It can be done on frozen sections, however it is usually not
done on
> > paraffin-embedded tissue since that is normally fixed. The process is
very
> > different than the "antigen retreival" techniques we use on paraffin
> > sections which are designed to break formalin/protein bonds.
> >
> > Tim Morken
> > CDC, Atlanta
> >
> >
> > -----Original Message-----
> > From: Greg Dobbin [mailto:dobbin@Upei.CA]
> > Sent: Tuesday, August 06, 2002 12:55 PM
> > To: histonet@pathology.swmed.edu
> > Subject: Ag retreival vs. Permeabilizing
> >
> >
> > Hello All,
> >
> > I found a "Brief Communication" in Vet Pathology (38:116-119;
> > 2001) that discusses expression of COX-2 in canine renal
> > carcinomas. The authors do not mention antigen retreival, rather,
> > they speak of permeabilizing the tissues using triton and
> > saponin. Is permeabilizing (as described) a form of antigen
> > retreival or does it serve another purpose?
> >
> > Has anyone else heard of this before? Explanation??
> > Thanks,
> > Greg
> > ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
> > Greg Dobbin
> > Pathology Lab
> > Atlantic Veterinary College, U.P.E.I.
> > 550 University Ave.
> > Charlottetown, P.E.I.
> > Canada,  C1A 4P3
> > Phone: (902)566-0744
> > Fax: (902)566-0851
> > ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
> > "A farmer is a person outstanding in their field."
>
>
> ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
> Greg Dobbin
> Pathology Lab
> Atlantic Veterinary College, U.P.E.I.
> 550 University Ave.
> Charlottetown, P.E.I.
> Canada,  C1A 4P3
> Phone: (902)566-0744
> Fax: (902)566-0851
> ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
> "A farmer is a person outstanding in their field."
>





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