Innate immunity activation largely depends on recognition of microorganism structures by Pattern Recognition Receptors (PRRs). PRR downstream signaling results in production of pro- and anti-inflammatory cytokines and other mediators. Moreover, PRR engagement in antigen-presenting cells initiates the activation of adaptive immunity. Recent reports suggest that for the activation of innate immune responses and initiation of adaptive immunity, synergistic effects between two or more PRRs are necessary. No systematic analysis of the interaction between the major PRR pathways were performed to date. In this study, a systematical analysis of the interactions between PRR signaling pathways was performed. PBMCs derived from 10 healthy volunteers were stimulated with either a single PRR ligand or a combination of two PRR ligands. Known ligands for the major PRR families were used: Toll-like receptors (TLRs), C-type lectin receptors (CLRs), NOD-like receptors (NLRs), and RigI-helicases. After 24 h of incubation, production of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1ß), IL-6, and IL-10 was measured in supernatants by enzyme-linked immunosorbent assay (ELISA). The consistency of the PRR interactions (both inhibitory and synergistic) between the various individuals was assessed. A number of PRR-dependent signaling interactions were found to be consistent, both between individuals and with regard to multiple cytokines. The combinations of TLR2 and NOD2, TLR5 and NOD2, TLR5 and TLR3, and TLR5 and TLR9 acted as synergistic combinations. Surprisingly, inhibitory interactions between TLR4 and TLR2, TLR4 and Dectin-1, and TLR2 and TLR9 as well as TLR3 and TLR2 were observed. These consistent signaling interactions between PRR combinations may represent promising targets for immunomodulation and vaccine adjuvant development.