[Histonet] HELP!!! peroxidase blocking on HCC paraffin sections!!!!

[Rene J Buesa]

Victor: You should be able to quench endogenous peroxidase with 3% UNLESS the H2O2 you are using has "expired" meaning that is NOT really a 3% solution.
  My first reccommendation would be to get really fresh H2O2 and try, again, Just 5 minutes at 3%, and that ought to be enough.
  The rationale is that peroxidase is an omnipresent enzime in all cells and if not oxidized (with H2O2) will produce a "background noise" (=unspecific staining) during the IHC procedure.
  Similarly DAB to react needs the addition of H2O2.
  Try a new fresh batch of hydrogen peroxide.
  René J.

Victor Wong <vhlwong <@t> yahoo.com> wrote:
  Hi all,

Last time I tried to block endogenous peroxidase with 0.3% hydrogen peroxide for 15 minutes but it came out a lot of background. Then I tried to block with 3% in PBS for 10 and 30 minutes. The background staining was greatly reduced esp. in that treated for 30 minutes. But the positive staining was also muchly reduced. Since I have no spared slides left, I cannot make others trial to massage the blocking conditions. I felt very frustrated and now I am seeking help from you folks. I use the DAKO's Envision+ kit therefore it should not be the endogenous biotin's problem. I also want to know the rationale behind the H2O2 on blocking the peroxide and the visualization of chromogen. Tons of thanks.

Victor

__________________________________________________
Do You Yahoo!?
Tired of spam? Yahoo! Mail has the best spam protection around 
http://mail.yahoo.com 
_______________________________________________
Histonet mailing list
Histonet <@t> lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet


 __________________________________________________
Do You Yahoo!?
Tired of spam?  Yahoo! Mail has the best spam protection around 
http://mail.yahoo.com