Rhipicephalus microplus, hard tick causes babesiosis and anaplasmosis in cattle. For decades, ticks control has been depending on synthetic acaricide use, however, acaricide-resistant ticks led to alternative effective and eco-friendly tick control approach such as vaccination. Currently, vaccine design using immuno-informatics is a promising method in the vaccination field. Here, we proposed a multi-epitope-based (ME) vaccine against R. microplus tick, comprising potential immunodominant B-cells, Helper-T Lymphocytes and Cytotoxic-T Lymphocytes epitopes. Then, designing of ME vaccine construct, containing sequences of Bm86, Subolesin and Bm95, was cloned into pET28a a prokaryotic expression vector. The recombinant ME protein was expressed in the Escherichia coli BL21 strain. After that protein purification was done by Nickle-NTA affinity chromatography. Evaluation of expression of recombinant ME based protein by SDS-PAGE analysis showed that optimal expression in LB medium for 8 h at 37 °C post-induction by 0.7 mM IPTG. The quality of purified recombinant protein was validated by Western blot analysis. 100 µg/2ml of purified ME protein with adjuvant Montanide ISA 50V was used in rabbits to evaluate the antigenicity of ME protein. Our study finding revealed that the expressed recombinant ME protein's molecular weight was 42kDa. The anti-recombinant protein antibodies raised in immunized rabbit’s sera was detected through western blotting and ELISA. This study results showed that recombinant ME protein is a potential vaccine candidate against Rhipicephalus microplus tick infestations.