The Morphology And Immunogenicity Of Two EV71Virion Fractions

Hand, foot and mouth disease broke out frequently in China and around the Asia-Pacific region in recent years. Hand, foot and mouth disease which has become a serious public health problem has been reported popularly in most parts of the world. Enterovirus71(EV71) is a member of the Enterovirus genus of the Picornaviridae family and is one of the major pathogens of Hand, foot, and mouth disease (HFMD). At present, immunological characteristics and vaccine research of EV71has been receiving increased attention because of no effective drag.Available information has confirmed that the EV71has typical20-sided shell structure, positive RNA genome codes for a polymeric proteins which enzymatic hydrolyzed to form4structural proteins VP1,VP2,VP3and VP4through different stages。 This structure is quite analogous with the structure features of other virus of the Picornaviridae family, such as poliovirus, what may lead to form two EV71virion fractions (Pseudo-and Real-particles) which are similar with the two types of poliovirus structures (D-and C-antigens) in cells. Therefore, the related work in this paper is based on the requirement of the reseach about virus structure and immunological characteristics in development of inactivated vaccine. Gained the infected KMB17cell by EV71at5time points (10h,16h,22h,28h and34h), and obtained the two EV71virion fractions by density gradient centrifugation combined with ultrafiltration. The morphology of virion fractions was analyzed by electron microscopy, then the amount of antigen was determined by Biotin-Avidin Enzyme Linked Immunosorbent Assay (BA-ELISA). Made use of Virus Micro-titration to determine infective titer of the virus, while the antigenic characteristics and immunogenicity of two virion fractions were determined by SDS-PAGE, Western blot. To study the immunogenicity of two EV71virion fractions by immunizing ICR mice with two EV71virion fractions in5time points and suckling mice protective experiment.Experiment results found that the directly observational distinctions of two virion fractions were that the upper layer was mainly consisted with empty particles, while the lower layer was consisted with solid particles and was accompanied by empty particles in the later time point. Dynamic analysis of infection indicated that solid particles maintained obviously upward tendency, whose infective titer was2-3log higher than empty particles. ELISA displayed that the amount of antigen of two EV71virion fractions showed an increasing trend with the process of virus infection. Detection of SDS-PAGE that the VPO had clove to VP2and VP4in solid particles, but empty particles still was primarily VPO. Compared to the upper layer, the lower layer has the similar antigenic characteristics but lower pathogenicity. Both two virion fractions had the capability to primarily induce an effective immune response in mice, regardless of their different neutralizing antibody titers and some distinctions have been observed.Analysis of the structure characteristics of EV71prompted that the directly observational distinctions of two virion fractions was mainly between solid particles and empty particles. Simultaneously, the essential distinction was the cleavage of VPO to VP2and VP4. Both two EV71virion fractions can be used as the vaccine antigen for immunization.