Hello,

I have now  perplexed results; I saw good amount of protein when I did
immunoprecipitation, but I cannot detect the protein by confocal
micromicroscope.  I am wondering whether IP is more sensitive than
microscopic observation, in general.  Of course, my staining system matters
here.  I used biotinylated Ab as an primary, then used
streptoavidin-conjugated either Cy3, Red-X or FITC.  I assume that I may
need brighter staining, so that I am considering to enhance the signal
(using TSA most probably - Thank you very much for your input!  It was me
who was asking about triple staining a several days ago).  Then hopefully I
can detect the protein also microscopically.

Any information will be appreciated.  And thank you for your help/replies on
my last question.


Mari

P.S. To be more specifically speaking, I am using anti-MHC class antibody
(28-14-8).  It is known that RMA-S cells produce class I molecules and they
can be detected by IP using this particular Ab, but the molecules does not
come out to cell surface under normal condition.  I use this cells as a
control of experiment and stained it.  There was no cell surface staining,
of course, but there was no tracellular staining as well.  And my testing
cells behaved the same.  So that's why I am wondering whether IP is
super-sensitive and confocal detection cannot be that sensitive  (otherwise,
it does not makes sense...)