**6. Conclusions**

The successful development of an effective vaccine needs exact data regarding the physicochemical characteristics of the antigen and adjuvant, as well as expertise in their combination to generate a safe, steady, and immunogenic vaccine.

Our study described the immunogenicity and protective capacity of H2B/PLA as a particulate vaccine in an anti-leishmanial therapeutic setting. The outcomes clearly demonstrate that the PLA nanoparticles serve as an adjuvant for the recombinant protein H2B since BALB/c mice were protected against experimental infection by the *L. major* parasite, a standard virulent strain GLC94. In contrast, mice immunized with a soluble protein were unable to stop the parasite's spread, which led the paw to completely ulcerate. The H2B/PLA's added value refers to its colloidal stability, simple manufacturing process, and established adjuvant capacity.

It will be interesting to extend these data by incorporating immunostimulatory molecules (such as imiquimod, 3 M 052) in the NP PLA core to increase its protective effect.

**Author Contributions:** Conceptualization, M.E.; Formal analysis, S.A.-R. and N.B.k.; Investigation, S.A.-R.; Methodology, T.T. and M.C.; Resources, B.V., Z.B., M.C. and M.E.; Supervision, B.B.-Z., B.V., T.T. and M.E.; Validation, B.V. and T.T.; Writing—original draft, S.A.-R.; Writing—review & editing, S.A.-R., N.B.k., B.B.-Z., B.V. and T.T. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Institutional Review Board Statement:** The animal study protocol was approved by the Biomedical Ethics Committee of Institut Pasteur de Tunis, Tunisia (CEBM: 2015/13/I/LR11IPT08 V1) and according to the 2010/63/EU Directive for animal experiments.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** Data available from the corresponding author.

**Acknowledgments:** In this section, special thanks are due to Hechmi Louzir, from the Institut Pasteur de Tunis, for his scientific comments and support. Thanks are addressed to Marouani A, Ellefi A, and Rammeh S, for technical assistance on laboratory animals.

**Conflicts of Interest:** The authors declare no conflict of interest.
