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From:"Andrea Kaj" <andreakaj@hotmail.com>
Date:Tue, 27 Jul 1999 23:19:48 PDT
Content-Type:text/plain; format=flowed

Hi Marija
Kappa/lambda is very demanding when it comes to fixation. You really need to 
be sure that the tissue is sliced appropriately and fixed immediately. That 
can be hard to achieve in routine pathology, -even lymphnodes are better off 
being sliced in halves.

Also it is very important to do antigen retrieval in a large volume of 
citratebuffer. I use 600 mL of buffer for each 20 slides (can be re-used, 
this ensures that the solution is not boiled. Youshould experiment a bit 
with the timing, but usually 22 minutes works the best for me. Afterwards 
flush with plenty of cold tap water

I use DAKO primaries - A0191 Kappa 1:2000 and A0193 Lambda 1:4000 and use 
their Chemmate DAB kit. Gives excelent results - then again, only when the 
fixation is proper. You will almost always see some diffusion of light chain 
IgG's in connective tissue and epithelium, but that is normal.
Hope this helps - Andrea
PS: I do this on a Techmate instrument, so adjustments might be needed.

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