Fine phenotyping of swine immune cells by multicolor flow cytometry

National audienceImmunological research in a species such as the pig is of great importance for its direct impact on the veterinary control of the pig husbandry and thus the economy of the pig industry as well as for the development of biomedical model. In order to monitor all cell types simultaneously from peripheral whole blood cells from MeLiM pigs (melanoma-bearing minipigs), we compiled the available knowledge to develop gating strategies for multicolor flow cytometry. Lineage, exclusion and activation markers allowed us to identify lymphoid (αβ and δ T cells, B cells, NK and NKT cells) and myeloid cells (dendritic cells, monocytes, granulocytes). The subsets of T lymphocytes were also distinguished according to their helper, cytotoxic or regulatory functions. In contrast with published data, we found that the anti-human CD19 antibody (clone BD3) did not stain B cells efficiently, and we provided a new strategy to detect different B cell subsets according to their maturation status. In particular, their loss of MHCII and CD21 expression correlated with the increase of circulating total IgG concentration. Myeloid cells display a very heterogeneous expression of CD172a, CD14, CD163, PG68A and MHCII. Whereas PG68A were considered as a pan-granulocyte marker, we found that eosinophils were PG68A-/low using May-Grünwald-Giemsa stainings. We also characterized different monocyte subsets, corresponding to differential activation status. As a direct application of this strategy, we could monitor pigs from 3 to 21 weeks of age to gain knowledge about the maturation of pig immune system from weaning to puberty. This immuno-phenotyping has also been extended to a high-throughput approach, which provides new perspectives to qualify the immune capacity of pigs in production. Finally, our fine strategy based on multicolor flow cytometry will be valuable to monitor the regulation of the immune response notably in case of inflammation, disease progression or treatment in swine