i am going to experiment with immunohistochemistry in relation to salmonella and macrophage.
by the way, i have no experience with IHC.
so, i write such a this short letter for you in order to give your hands.
first of all, the biggest problem is that i do not know weather macrophage inner proportion is possible to be stained by any other stain buffer or not.
if there are people who experience with IHC, then give me a good way and discussion.
i will wait for your help.
Macrophages are antigen presenting cells and getting a specific marker to an antigen presenting cell can be a difficult task as they express different markers. Nonetheless, several reports hint to using F4/80 as a specific marker for mouse macrophages and I am currently using this marker both to immunophenotype tumor infiltrating macrophages using FACS and to detect their presence in tumor tissues using IHC. To come back to your question of how to do the immunohistochemistry, I think that will depend on the source of your reagents particularly the primary antibody and the type of detection (biotin, HRP, ...) as the protocol can be different accordingly which can be learnt from the data sheet of the antibody.
I stain mouse tissue macrophages with A monoclonal antibody against F4/80, which is an extracellular cell surface marker, although I found this was not as specific as I liked. I've started using CD68, an intracellular marker which doesn't stain langerhans cells, one of my initial problems.
There are many protocols for IHC, depending on what you need to observe. If you're looking at a cultured cell line, you might want to try immunocytochemistry, where you spin the cells down onto a glass slide and stain using fluorescent probes instead of HRP. This will also allow more a more versatile staining profile where you can further characterise the polarisation state of the macs with other antibodies.
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