Expression of the Recombinant Hsp20-Nef Protein for Evaluation of its Immunogenicity against HIV-1 Nef using Indirect ELISA
نویسندگان
1 Biology Department, Sciences & Research Branch, Islamic Azad University, Tehran, Iran
2 Pilot Nano-Biotechnology Department, Pasteur Institute of Iran, Tehran, Iran
3 Biology Department, Sciences & Research Branch, Islamic Azad University, Tehran, Iran
4 Hepatitis & AIDS Department, Pasteur Institute of Iran, Tehran, Iran
doi
چکیده
Aims: Nef protein has been considered as an attractive target for the development of therapeutic HIV-1 vaccine. Furthermore, strong immunological properties of heat shock proteins (HSPs) led to their use as for subunit vaccine candidates. In the current study, the generation of Hsp20-Nef fusion protein was performed in E. , and in BALB/c mice. Materials and Methods: At first, of Hsp20-Nef recombinant protein E. BL21 and Rosetta strains by SDS-PAGE and western blotting using anti-Nef monoclonal antibody. Then, the recombinant protein was purified by a reverse staining method. Finally, its potency was evaluated to elicit antibody response against HIV-1 Nef antigen using indirect ELISA in mice. Findings: Our data showed a clear band of ~1230bp related to Hsp20-Nef fusion on agarose gel indicating the correct gene cloning in pET28a vector. The expression of Hsp20-Nef protein was confirmed as a clear band of ~47 SDS-PAGE and western blotting. In the immunological assay, the Hsp20-Nef protein and also the Nef protein emulsified with Freund’s adjuvant significantly enhanced the level of total compared to other groups. Moreover, of Hsp20-Nef was higher than Freund’s adjuvant/Nef in protein regimens (p<0.05). Conclusion: The Hsp20-Nef fusion protein was effectively expressed in E. and significantly induced antibody response against HIV-1 Nef antigen.
