This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. NK cells in rhesus macaques have been variably defined as CD3-negativeCD16-positive or CD3-negativeCD8-positive, although only limited efforts have been made to rigorously validate these definitions. To better understand the role of NK cells in macaque disease models, we undertook a multiparameter analysis of macaque NK cells employing four color flow cytometry and a panel of lineage-specific and non-lineage specific lymphocyte markers. Using this approach, we recently identified two distinct populations of candidate NK cells: a major CD8brightCD16-positive population and a minor CD8brightCD16-negative population. In addition, a CD56-positive subset of cells within the minor rhesus NK population was identified that expressed chemokine and lymph node homing receptors similar to those expressed by the CD56bright NK cell population identified in humans. This work has now been extended using 8 color flow cytometry and has verified the existence of this different populations of NK cells. In addition, we have developed a functional flow cytometric assay for the ability of NK cells to degranulate in response to stimulation with target cells. These improved immunophenotypic definitions of macaque NK cells should facilitate future analysis of innate immune responses in rhesus macaques and the role of NK cells in AIDS pathogenesis in SIV-infected macaques.