The skin is a valuable target for vaccine delivery because it contains many immune cell populations, notably an-tigen presenting cells. Skin immune cells have been extensively described in mice and humans but not in non-human primates, which are pertinent models for immunological research in vaccination. The aim of this work was to describe immune cell populations in the epidermis, dermis and skin draining lymph nodes in cynomolgus macaques by a single 12-parameter flow cytometry protocol. Given that skin cells share several markers, we defined a gating strategy to identify accurately immune cells and to limit contamination of one immune cell population by another. The epidermis contained CD1a + CD1c − Langerhans cells (LCs), CD3 + T cells and putative NK cells. The dermis contained CD1a + CD1c − cells, which were similar to LCs, CD1a + CD1c + dermal dendritic cells (DDCs), CD163 high CD11b + resident macrophages, CD3 + T cells and putative NK cells. The skin also contained CD66 + polymorphonuclear cells in some animals. Thus, immune cell populations in the macaque are similar to those in humans despite some differences in phenotype. In skin draining lymph nodes, we identified migratory LCs, CD1a + CD1c + DDCs and macrophages. The simultaneous identification of these different immune cells with one panel of markers avoids the use of large amounts of precious sample and may improve the understanding of immune mechanisms in the skin after treatment or vaccination.