Phenotypic diversity of antigen-presenting cells in ovine-afferent intestinal lymph

The phenotypic and functional repertoire of antigen-presenting cells (APCs) remains incompletely characterized, particularly during the migratory phase of their life history, when these cells leave peripheral tissues and travel via afferent lymphatic vessels to regional lymph nodes. Lymphatic cannulation procedures were used to collect ovine APCs as they migrated from the mucosa of the small intestine to regional lymph nodes. A panel of 19 new monoclonal antibodies (mAbs) was produced to characterize surface molecules expressed on APCs by means of two-color flow cytometry and microscopy. Two broad patterns of mAb reactivity were evident. Twelve mAbs reacted almost exclusively with cells in the APC-gated region, because all these mAbs stained < 3% of cells in the lymphocyte-gated region. Within this group, some mAbs identified distinct subsets of dendritic cells (DCs). The second group of seven mAbs displayed high-intensity staining on cells in the APC-gated region but also reacted with variable numbers (4-26%) of cells in the lymphocyte-gated region. This indicates that molecules recognized by these mAbs are highly expressed on APCs but also occur on other lineages. When new mAbs were analyzed by two-color flow cytometry of cells in afferent intestinal lymph, a wide range of differences in reactivity were observed, especially on CD11b(+), CD11c(+), CD4(+), MHCII(+), and gammasigmaTCR(+) cells. Although molecular specificities of mAbs reported here remain undefined, marked heterogeneity of staining patterns indicates considerable phenotypic, and probably functional, diversity within APC population in afferent intestinal lymph. MAbs reported here will provide useful tools to explore these features further.