I have been trying for ages to block a surface protein on my cells through administering a specific antibody that binds to the protein and thus inhibits physical interactions with other proteins or the same protein on other cells. However, I never had success - I never see the expected morphologic effect. I've so far tried:
- different antibodies
- different antibody concentrations
- different scales (cell densities and well plates)
- different ways of administering the antibody (adding directly to cell culture vs. preincubation with subsequent culturing)
The one thing I have not yet tried is to preincubate, culture and add the antibody again every 24 hrs or so. But I don't think the antibody is degraded so quickly when the cells are in medium, for I have found that it is detectable by staining flow cytometry even days after administration. I should also add that I had proper controls (isotype and no antibody), and the antibody-treated cells look like the control cells.
I am at a complete loss why it is not working or what I could try next. Has anyone got an idea ?