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Background: In this study, the pilot production of aerobic bioreactor tropical theileriosis vaccine was optimized with the aim of immunological assays for further mass production. Methods: We have shown earlier the delayed type hypersensitivity (DTH) assay could be used for evaluating the immunity and memory cells against specific Theileria antigen in vaccinated animals. In addition, TNF-α is the principle cytokine in modulating the cytotoxic activity of cytotoxic T-lymphocytes (CTL). Immunological analysis of the vaccine was performed by using two cell mediated immunity (CMI) in vitro and in vivo DTH test (Theilerin) and TNF-α assay. Results: The results of immune responses of susceptible immunized cattle by bioreactor vaccine in comparison with conventional flask vaccine revealed a significant stimulation of immune cells by transcription of high level of TNF-α and positive reaction against Theileria antigen in Theilerin skin test (DTH). Conclusion: The equal immunological results achieved in both above mentioned vaccines verified the satisfactory immunity for aerobic bioreactor theileriosis vaccine for advance mass vaccination in the field on a large scale. [ABSTRACT FROM AUTHOR]