RE: FAM fixative and immunohistochemistry

From:Karen Larison <>

It's my bet that most commercial antibodies have been screened on 
formaldehyde-fixed material, and if you are using commercial 
antibodies, formaldehyde fixation is probably the best.  But my 
experience in screening antibodies made at our local monoclonal 
facility is that the optimal fixation method is wholly 
antibody-dependent.  Some antibodies work on aldehyde-fixed material; 
some work better with denaturing fixatives, and some won't work 
unless you use Carnoy's, a chloroform-containing fixative that 
apparently extracts components of the tissue.  This actually makes 
good sense.  If your epitope is on the surface of the protein or if 
you have a conformational epitope, formaldehyde-based fixatives are 
no doubt the best.  But if your epitope is buried in the protein, a 
denaturing fixative may be required to expose the epitope.

Karen in Oregon

>Yes in my experience 10% NBF is the best fixative for immunohistochemistry
>procedures, using different fixatives wil cause unsatisfactory results and
>overfixation will also cause a problem.
>-----Original Message-----
>From: []
>Sent: Wednesday, May 30, 2001 2:30 PM
>Subject: FAM fixative and immunohistochemistry
>Hi all,
>I have a question for all of the chemistry and IHC geniuses out there.  I
>have a request that specifies formaldehyde-acetic acid-methanol (FAM)
>fixation on brain tissue, which after routine processing and paraffin
>embedding requires immunohistochemistry.  Early IHC testing on this tissue
>has been less than satisfactory.  Does this fixative pose a problem?  Would
>10% neutral-buffered formalin be more appropriate?  (Okay, that's two
>David A. Reynolds, HT(ASCP)
>6605 Merrill Creek Parkway
>Everett, WA  98203
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