AbstractMicroparticles have been intensively explored as next generation vaccine carriers in the last decade. They were mostly loaded with toll-like receptor ligands as adjuvants, which are not finally rated for their safety profile. Here, ligands of cytosolic receptors sensing nucleotide and oligomerization domains (NOD) were explored as alternative adjuvants in combination with immunologically inert particulate carriers from poly[(rac-lactide)-co-glycolide], which were shown to be capable to coencapsulate protein antigens as well as γ-d-glutamyl-meso-diaminopimelic acid (iE-DAP) and N-acetylmuramyl‐l-alanyl‐d-isoglutamine (MDP) as NOD receptor agonists. For selectively studying the immunomodulatory potency of the adjuvants rather than of potential immunogenic impurities and for providing a strategy for quality control of future particulate vaccines, the purity of the microparticles was confirmed with different reporter cell lines selectively expressing specific pathogen recognition receptors. Microencapsulated NOD ligands but not blank microparticles were shown to induce a dose-dependent maturation of human monocyte-derived dendritic cells to a proinflammatory phenotype with high levels of released cytokines particularly for iE-DAP. These data suggest their further exploration in vivo.