Kappa/lambda staining
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From: | Dave Tacha <dtacha@ncal.verio.com> (by way of histonet) |
To: | histonet <histonet@magicnet.net> |
Reply-To: | |
Content-Type: | text/plain; charset="us-ascii" |
> Kappa/lambda technique
There is quite an art in getting good kappa/lambda staining, especially
if tissues have been decalcified. Here are some tipsI've used and
learned over the past 10 years.
I use an Ionic Exchange Decal Unit for bone marrow biopsies. This works
very fast, but is much more gentle than HCL or RDO. After,
decalcification, I do not use antigen retrieval. I not sure about
DECLERE. It may not work well with certain types of antigens or
antibodies. I've heard a lot of mixed reports over the Histonet. I
prefer trypsin or pepsin digestion. I also block with 10% goat serum.
This seems to slightly improve staining. I prefer to use monoclonals
versus polyclonals. Sometimes, I dilute my antibodies in a mixture of
goat serum and casein. I also develop the slides under the microscope
to try and determine end-point staining. I prefer to use AEC as a
chromogen. You can differentiate AEC (before hematoxylin staining) in
95% alcohol. Then wash in water and observe the staining under the
microscope. I repeat the steps, if necessary. This technique really
helps improve contrast and reduces some of the background staining
caused by circulating antibodies.
I hope this has helped you.
If you want more information about the Ion Exchange Unit, e-mail me at
dave@biocare.net
David Tacha HTL (ASCP)
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