Re: Mart-1 IHC- I need your expertise please!!

From:Joseph Lee

We use Zymed's monoclonal MART-1 (Melan-A) cocktail
(clones: A103/M2-7C10/M2-9E3) antibody in our lab with
10 min HIER Citrate buffer, and DAB detection.

joe
SFSU


--- Vinnie Della Speranza  wrote:
> I'd appreciate any insight anyone may offer with the
> following.
> 
> we frequently perform Mart-1 ihc staining to
> demonstrate melanocytes.
> We use a monoclonal antibody from Dako (clone A103
> working dilution
> 1:50)  for this marker and an alkaline phosphatase
> detection kit from
> Biogenex utilizing a red chromogen. we perform high
> temperature antigen
> retrieval in citrate buffer pH 6.0
> 
> our positive control is a nevus selected by one of
> our
> dermatopathologists. we generally prepare a working
> dilution of this
> antibody once each week
> 
> from time to time, we've received feedback that "not
> enough melanocytes
> are staining" which causes the dermpathologist to
> conclude that the
> antibody is not performing to expectations. Slides
> are always checked
> prior to being turned in to verify that we have
> achieved the expected
> pattern of staining, however we do not attempt to
> quantitate the # of
> slides appearing positively stained with this
> marker.
> 
> I reviewed a case today that had been rejected by
> the pathologist. In
> evaluating the epidermis of four pieces of excised
> skin (FFPE), I found
> evidence of appropriate staining in each, although
> some areas of the
> epidermis contained many positive cells, some areas
> only a few, some
> areas none at all. the melanocytes in our control
> nevus stained with the
> expected pattern and intensity.
> 
> My questions to you are as follows:
> 
> has anyone found variable Mart-1 staining in tissues
> such as I've
> described and if so, can you elaborate as to it's
> suspected cause?
> 
> is it possible that handling of the specimen prior
> to fixation and
> processing could cause the result we are
> experiencing?
> 
> is it possible that pretreatment of the surgical
> site in the patient,
> including the use of topical or injected anesthetics
> or skin
> disinfectants could cause variable Mart-1 staining?
> 
> is there something about the stability of this
> antibody that we haven't
> considered that may be contributing to this
> situation?
> 
> is it possible that the pathologist expected a
> different outcome and
> thus concluded that the stain results were
> incorrect?
> your thoughts are most appreciated.
> 
> 
> 
> 
> 
> 
> Vinnie Della Speranza
> Manager for Anatomic Pathology Services
> Medical University of South Carolina
> 165 Ashley Avenue  Suite 309
> Charleston, SC 29425
> Ph: 843-792-6353
> fax: 843-792-8974
> 


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