Recent findings underline the role of NK cell subsets in regulating adaptive immunity. To define characteristics of NK cell subpopulations, purified CD56(dim) and CD56(bright) NK cells were analyzed by using gene chip arrays covering more than 39,000 transcripts. Gene profiling revealed resting NK cells to differ in respect to 473 transcripts with 176 exclusively expressed in CD56(dim) and 130 solely in CD56(bright) NK cells. Results were compared with array analyses using mRNA obtained from activated CD56(dim) and CD56(bright) NK cells. In this approach, NK cell receptors, cytolytic molecules, adhesion structures, and chemokine ligands showed differential expression patterns in the two subpopulations. These data were validated using FACS, RT-qPCR, or cytokine bead array (CBA) techniques. Cytokines produced by CD56(dim) and CD56(bright) NK cells were determined using a protein array covering 79 different bioactive mediators. GDNF, IGFBP-1, EGF, and TIMP-2 were detected in both subsets. In contrast, IGFBP-3 and IGF-1 were mainly produced by CD56(dim), while GM-CSF, TARC, and TGFbeta3 were expressed by CD56(bright) NK cells. In summary, we report new characteristic features of CD56(dim) and CD56(bright) NK cells, further underscoring that they represent independent populations with functionally diverse capabilities. The information on NK cells generated in this study will help to define corresponding NK cell populations in other species that lack CD56 expression on NK cells, such as mice. This will subsequently lead to the establishment of suitable animal models for detailed analysis of NK cell populations in vivo.