[Histonet] PSA_NCAM immunohistochemistry

[tf]

Dear All:

Just wonder anyone have worked on PSA_NCAM IHC of frozen sections?
How about 1:1000 mouse anti-PSA NCAM from millipore to stain 30 um brain sections?

And, do you need to add triton into the PBS, to "pore" the membrane? 
I think PSA-NCAM is distributed on the cell membrane, thus PBS may work?

Protocol:
1. Rehydration. PBS
2. Mouse-anti PSA-NCAM 1:1000  overnight + Blocking simultaneously.
3. 3* PBS
4. Goat-anti mouse conjugated with 488 fluroscein
5. 3 * PBS
6. DAPI
7. PBS wash
8. mount

Any other comments on this? 
especially the time & concentration for first antibody?
thx,

2008-07-31 



tf