[Histonet] Kappa and Lambda

[Hugh Luk]

Ellen,
Kappa and Lambda staining has always been hard.  It also depends on what 
tissues you are staining (ie liver has a lot of endogenous Biotin).  What 
are you using as a control?  Tonsil?  Is your tissue fixation and processing 
the same as the control?  How are you fixing your bone marrows?  Decal 
solutions often changes or "kills" IHC staining parameters.

Dako's Envision Plus is great (yeah, this is a very subjective statement; 
but tough! I'm getting good, repeatable results), but are your antibodies 
"Optimized" for Envision Plus?  Your incubation may be too long or (if you 
buy concentrated antibodies) your dilution too strong.  One of the hospitals 
I work at uses Proteinase K followed by 8 min primary, then LSAB2.  My other 
job uses HEIR, 10 min primary made with diluent WITH Background Reducing 
Components (S3022), then Envision plus.

Decal, because the timing changes with the pathologists/grossers, may need 
to be standardized.  This may be the hardest part (as it was with mine, I 
had 23 pathologists and 9 sets of grossers).

Three suggestions then:
1) Reoptimize titers to determine best time/concentration for Bouin's 
tissues with a Bouin's Fixed and/or decal tonsil.  This may include that you 
"Dictate" grossing situations (fixatives, timing of fixatives, and 
decalcification).

2)  Try a Protein block.  And/or a Avidin/Biotin block.

3) Check with Dako.  I have found their technical support to be awesome.

As we say in Hawaii; "Eh! No be shame!"  We all have had some problems with 
these antibodies.  If Dako's Ab's are not workable with Bouin's or whatever 
other variables there are, ask other vendors/techs which company's 
antibodies work best.

Good luck.  Hope this helps.

Hugh Luk, HTL(ASCP)
hluk <@t> crch.hawaii.edu
Epidemiology Section
Cancer Research Center of Hawaii
1236 Lauhala Street, Room 316
Honolulu, HI  96813
Tel: (808) 440-5238




>From: Chewy71874 <@t> aol.com
>To: histonet <@t> lists.utsouthwestern.edu
>Subject: [Histonet] Kappa and Lambda
>Date: Fri, 17 Dec 2004 01:38:58 EST
>
>We are having a problem with background staining on Kappa and Lambda 
>stains.
>We use DAKO rabbit polyclonal antibodies, DAKO TRS for antigen retrieval in
>the steamer, Envision+ HRP, DAB.  The background staining is especially bad 
>on
>Bouin's fixed, decaled bone marrows.  Does anyone else have this problem?   
>Do
>you get cleaner staining with monoclonal antibodies?  We are open to
>suggestions.
>
>Thanks in advance,
>
>Ellen Yee, HT
>Lead Tech, Immuno Dept.
>Central Histology Facility
>Diagnostic Pathology Medical Group
>2420 J St.
>Sacramento, CA 95816
>ph: 916-447-2718
>fax: 916-447-0620
>Sacramento, CA
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