Objective(s): Brucellosis is a common infectious disease among animals and humans. While subunit vaccines could be used as an efficient strategy against pathogens, they usually seem to be less immunogenic than live or killed vaccines. However, the use of a suitable adjuvant accompanied by subunit vaccines can be a good alternative to enhance the immune response. Materials and Methods: To find a proper adjuvant against Brucellosis, the immune response of induced mice by Aluminum Hydroxide (AH), Incomplete Freund (IFA), and Chitosan Nanoparticle (CS) adjuvants in individuals and in combination with CS were assessed. Results: Immunization with CS stimulated higher interferon gamma (IFN-γ) immunity, while there were no significant differences between rOMP25 (IFA), rOMP25 (AH), rOMP25 (AH-CS) and rOMP25 (IFA-CS) recombinant proteins. Tumor necrosis factor alpha (TNF-α) analysis revealed there were no significant differences between immunized groups and the positive control group, except for the treatment formulated in single IFA. Furthermore, unlike IFN-γ, there was a reverse interleukin-4 (IL- 4) immune response trend for treatments, as rOMP25 (CS) displayed the lowest response. rOMP25 (CS) induced higher titer of total antibody than the other ones. Although the recombinant proteins emulsified in different adjuvants induced similar titer of IgG1 antibody, the ones that were formulated in CS, IFA and IFA-CS showed a higher titer of IgG2a. The cell proliferation assay demonstrating the antigen-specific cell proliferative response could be promoted after immunization with CS. Conclusion: CS whether single or in combination with IF adjuvants has potential to improve Th1-Th2 responses.

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