Immunostaining - Amplification of signal - (Oct/09/2008 )
Hi all,
we´re having problems with the immunodetection of a transmembrane protein in murine tissue (several commercial ab´s failed so far in WB and IF) and one of the reasons might be the low expression level.
I´d like to go again for IHC and I wonder if there is any possibility to enhance the signal intensity by any amplification step?
Would it be worth trying HRP or AP staining instead of immunofluorescence to obtain a stronger signal?
I would be grateful for any general suggestions, hints or protocols!
-luna77-
QUOTE (luna77 @ Oct 9 2008, 02:16 AM)
Hi all,
we´re having problems with the immunodetection of a transmembrane protein in murine tissue (several commercial ab´s failed so far in WB and IF) and one of the reasons might be the low expression level.
I´d like to go again for IHC and I wonder if there is any possibility to enhance the signal intensity by any amplification step?
Would it be worth trying HRP or AP staining instead of immunofluorescence to obtain a stronger signal?
I would be grateful for any general suggestions, hints or protocols!
we´re having problems with the immunodetection of a transmembrane protein in murine tissue (several commercial ab´s failed so far in WB and IF) and one of the reasons might be the low expression level.
I´d like to go again for IHC and I wonder if there is any possibility to enhance the signal intensity by any amplification step?
Would it be worth trying HRP or AP staining instead of immunofluorescence to obtain a stronger signal?
I would be grateful for any general suggestions, hints or protocols!
I think HRP or AP or ABC methods all amplify signal very well,
but you doing tissue sample in paraffin? if so, and if possible, cryo-section is much easy to reveal antigen and stained for IF.
but I also think that you might need to change antibody itself, because you could not detect with WB means antibody might not be appropriate, or need to change antibody which recognize different epitope of your protein of interest.
-Rnotk-