Methods and Techniques Discussion => Antigen Retrieval => Topic started by: Leonard Cheung on November 28, 2005, 03:57:57 AM

Title: Enzyme Digestion
Post by: Leonard Cheung on November 28, 2005, 03:57:57 AM
Which enzyme digestions are good for VWF & PECAM?
Title: Enzyme Digestion
Post by: richard03 on November 28, 2005, 07:10:36 AM
Proteinase K digestion method is good for both VWF & PECAM. You can use the following protocol and it works well.

http://www.ihcworld.com/_protocols/epitope_retrieval/proteinase-k.htm

Richard
Title: Enzyme Digestion
Post by: Leonard Cheung on November 28, 2005, 12:58:56 PM
Thx..Richard

Are there any other antigen retrieval methods good for VWF & PECAM?
Title: Enzyme Digestion
Post by: richard03 on November 28, 2005, 08:39:08 PM
Leonard,

Other enzyme methods (pronase, trypsin, pepsin) should work fine too. In general, they are similar.

http://www.ihcworld.com/epitope%20retrieval.htm

Richard
Title: Enzyme Digestion
Post by: Leonard Cheung on November 28, 2005, 09:47:43 PM
Richard,

How about the heating methods? such as citrate buffer, etc..

Thx
Leonard
Title: Enzyme Digestion
Post by: richard03 on November 29, 2005, 07:19:09 AM
Leonard,

The key factor for HIER is pH. For example, citrate buffer (pH 6) and Tris-EDTA (pH 9) work for different antibodies. Usually, the higher pH, the more intense staining.

HIER and PIER are usually not interchangable.  An antibody working for PIER is usually not working for HIER.

Richard
Title: Enzyme Digestion
Post by: Leonard Cheung on November 29, 2005, 11:23:19 AM
ic....thx a lot ...Richard