Expression and determination of immunization dose of recombinant tropomyosin protein of Hyalomma anatolicum for the development of anti-tick vaccine

Abstract

Present investigation was carried out to standardize the immunization dose of one recombinant antigen in rat model before conducting large animal experimentation. Tropomyosin (TPM), a muscle associated and highly conserved protein found in all species of invertebrates, was cloned, expressed in prokaryotic system and the downstream process was standardized. SDS-PAGE analysis showed a distinct band of approximately 51 kDa. Western blot analysis using specific sera gave a strong reaction of approximately the same size as that of SDSPAGE. For standardization of immunization dose, the rTPM at three different dosages viz., 150, 300, 450 μg was used to immunize wister rats and the antibody response was titrated by ELISA. Applying ANOVA, highly significant difference in anti-rTPM titre was recorded between the animals injected with 300 μg total dose (TD) and other dosages selected for the study. The significantly high antibody tire at 1:25600 dilution observed in animals immunized with 300 μg TD was selected for further study on in vivo immunization of calves and experimental challenge by the tick stages.