en 30090208 <p>Background: Hepatitis C (HCV) is known as a serious blood-borne disease that infects millions of people globally. NS3 is a conserved non-structural sequence of hepatitis C virus which has a major role in activating specific CTL responses. As known, there is no effective vaccine against HCV infection, thus it is required to design a specific regimen of vaccination. Recently, the strong immunological properties of Heat shock proteins (Hsps) led to their use as immunomodulators and an antigen carrier for subunit vaccine candidates. In the current study, the role of Hsp20 was evaluated as a HCV NS3 gene carrier in mammalian cell line.<br /> Methods: At first, the recombinant plasmids of pEGFP-Hsp20, pEGFP-NS3, and pEGFP-Hsp20-NS3 were constructed and their accuracy was confirmed by digestion and sequencing. Then, all recombinant plasmids were transfected into HEK293T cells by Lipofectamine and TurboFect gene delivery systems. Finally, the expression of proteins was assessed by fluorescent microscopy, western blotting, and flow cytometry.&nbsp;<br /> Results: In western blotting, the 47, 59, and 79 kDa bands were detected for pEGFP-Hsp20, pEGFP-NS3, and pEGFP-Hsp20-NS3, respectively. The percentage of NS3-Hsp20-GFP protein expression was ~67% by TurboFect and ~50% by Lipofectamine indicating high potency of TurboFect delivery system. Furthermore, the expression of Hsp20 (~83%) was higher than NS3 (~58%) in the cells transfected by TurboFect using flow cytometry analysis. This result was confirmed in the expression of Hsp20-NS3 fusion (~67%) in which Hsp20 increased the delivery of HCV NS3 <em>in vitro</em>. The same data were obtained by Lipofectamine transfection reagent.&nbsp;<br /> Conclusion: Briefly, our data confirmed the role of Hsp20 as a suitable antigen carrier for DNA vaccine design.</p> Hepatitis C virus, Small heat-shock proteins, Vaccines 152 157 https://www.ajmb.org/En/Article.aspx?id=320 https://www.ajmb.org/PDF/En/FullText/320.pdf MarziehBasirnejadDepartment of Molecular and Cellular Sciences, Faculty of Advanced Sciences & Technology, Pharmaceutical Sciences Branch, Islamic Azad University, Tehran, Iran1288AzamBolhassaniDepartment of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran50Seyed MehdiSadatDepartment of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran817