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Abstract In the present study, for the first time, we released and assembled the particles of three major structural proteins of velogenic NDV (M, HN, and F glycoproteins) as a NDV-VLPs. The ElISA result of the cytokines of splenocyte suspension cells showed that IL2, IL10, TNF-α, and IFN- ˠ titers were significantly higher (p ≤ 0.05) in mice that were immunized only with NDV-VLPs three times with a 10-day interval, in comparison to those that were immunized with NDV-VLPs twice in a 10-day interval and received a B1 live vaccine boost on the third interval. Flow cytometry results showed that CD8 + titers in the group that only received NDV-VLP was higher than other group. However, serum ELISA results did not show a significantly (p ≥ 0.05) higher NDV antibody titer in NDV-VLPs immunized mice compared to the boosted group. Besides, HI results of SPF chickens vaccinated with NDV-VLPs and boosted with B1 live vaccine were significantly (p ≤ 0.05) higher than those that only received NDV-VLPs. Interestingly, after challenging with NDV sub-genotype VII, all the chickens that were solely vaccinated with NDV-VLPs remained alive (six out of six), whereas two out of six chickens that were vaccinated with NDV-VLPs and also received the B1 live vaccine boost died. In conclusion, our results strongly indicated that the T-cell immune response in an NDV host is more important than the B-cell response. Also, the results of the present study revealed that to completely protect chickens against velogenic NDV strains, a vaccine comprising specific epitopes of velogenic strain is needed.