Choosing and Preparing Antigen‐Presenting Cells

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Abstract
The first issue in many antigen-processing experiments is to choose and obtain appropriate antigen-presenting cells (APCs). Macrophages have characteristics that dictate their use in certain antigen-processing experiments, so this unit describes preparation of activated macrophages, which involves eliciting peritoneal macrophages from mice by infection with Listeria monocytogenes. This method is often preferred because the resulting macrophages are more consistently activated to a high degree, but macrophages can also be obtained (as described here) by sequential injection of L. moncytogenes and proteose peptone, by induction with concanavalin A (Con A), or by isolation from bone marrow. Lipopolysaccharide-stimulated B lymphoblasts can also be used as APCs. Protocols are included for preparation of L. moncytogenes stocks and preparation of L929-conditioned medium which is used for culturing bone marrow-derived macrophages.