Assessment of pDCs functional capacity upon exposure to tumor-derived soluble factors.

Plasmacytoid dendritic cells (pDCs) are a minority subset of dendritic cells that despite their tiny quantity play an important role in the immune system, especially in antiviral immunity. They are known mostly as the major producers of type I IFN, which they secrete upon stimulation of endosomal Toll-like receptors 7 and 9 with viral RNA and DNA. However, the functionality of pDCs is more complex, as they were shown to be also involved in autoimmunity, inflammation, and cancer. In the context of the tumor microenvironment, pDCs mostly show substantial functional defects and thus contribute to establishing immunosuppressive micromilieu. Indeed, tumor-infiltrating pDCs were shown to be predominantly pro-tumorigenic, with reduced ability to produce IFNα and capacity to prime regulatory T cells via the ICOS/ICOS-L pathway. Here we describe in detail a method to assess the functional capacity of pDCs upon exposure to tumor-derived cell culture supernatants. The same technique can be implemented with minimal variations to test any soluble factor's impact on pDC phenotype and function.