I have purchased a Anti-FLAG Mouse antibody conjugated with FITC from Sigma. I intend to use it for confocal microscopy and western blotting. They have provided a proceedure for my microscopy work, however when i contacted them about the western blotting (which the product information specifically recommends) they have informed me that it has never been used in a western blotting application (atleast in published work or in their testing). I do not have the correct equipment to detect the FITC directly. So i was thinking of using a secondary detection system (such as Lumi-Light Plus Western Blotting Kit from Roche) which detects the primary mouse anti-body.
I am worried that the conjugated FITC may inhibit the binding of the secondary anti-body. I have found examples where people have had success using a secondary detection system with primary anti-bodies conjugated with FITC and it did not appear to have any effect on the binding of the secondary anti-body.
I was wondering if anyone else had used a conjugated anti-body in the same mannor and had any success?
FLAG Antibody conjugated with FITC
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