Aim: To obtain recombinant VP0 protein of enterovirus 71, and generate the corresponding VP0-specific polyclonal antibodies, for molecular detection and characterization of EV71.
Methods: The VP0 gene was amplified by PCR and cloned into vector pET26b to make pET-VP0 for the prokaryotic expression of VP0. The recombinant VP0 protein was expressed in E.coli BL21 harboring pET-VP0, purified from inclusion bodies, renatured, and subsequently used to immunize guinea pigs. The resultant antisera were evaluated for anti-VP0 titer, binding capacity and specificity by ELISA, immunofluorescence staining and Western blot assays.
Results: Recombinant protein VP0 was efficiently produced in E.coli. Immunization of guinea pigs with recombinant VP0 elicited high-titer (1:10(6)) VP0-specific antibodies. Western blot analysis showed the resultant anti-VP0 sera reacted with E.coli-expressed VP0 as well as EV71 propagated in Vero cells. Moreover, the antisera positively recognized EV71 infected cells by immunofluorescence staining.
Conclusion: The recombinant VP0 and the corresponding polyclonal antibodies can be used to identify and characterize EV71, and therefore represent useful agents and tools for the development of new diagnostic methods and vaccines for EV71.