AimsWe evaluated the potential of a nanoparticle (NP) delivery system to improve methods of delivery of candidate peptide-based vaccines for Paratuberculosis in cattle. Methods and ResultsPeptides derived from Mycobacterium avium subsp. paratuberculosis (Map), and the pro-inflammatory monophosphoryl lipid A (MPLA) were incorporated in polymeric NPs based on poly (d,l-lactide-co-glycolide) (PLGA). The PLGA/MPLA NPs carriers were incubated with macrophages to examine their effects on survival and function. PLGA/MPLA NPs, with and without Map antigens, are efficiently phagocytized by macrophages with no evidence of toxicity. PLGA/MPLA NP formulations did not alter the level of expression of MHC I or II molecules. Expression of TNF and IL12p40 was increased in Map-loaded NPs. T-cell proliferation studies using a model peptide from Anaplasma marginale demonstrated that a CD4 T-cell recall response could be elicited with macrophages pulsed with the peptide encapsulated in the PLGA/MPLA NP. ConclusionsThese findings indicate PLGA/MPLA NPs can be used as a vehicle for delivery and testing of candidate peptide-based vaccines. Significance and Impact of the StudyThese results will assist on more in depth studies on PLGA NP delivery systems that may lead to the development of a peptide-based vaccine for cattle.