Evaluation of Antibodies Induced by the Injection of Single Capsid Protein or Purified Virus Particle of Coxsackievirus B3 in Mice


Four capsid proteins (VP1, VP2, VP3, and VP4) of coxsackievirus B3 (CVB3) were expressed as recombinant proteins in an Escherichia coli expression system and used as antigens for subunit vaccines against CVB3 in ICR mice. Antigens were expressed as thioredoxin-histidine (TrxHis)-tagged protein and purified before immunization. Although all VPs other than VP4 induced anti-CVB3 specific antibodies in mice (detected by ELISA and western blotting), they did not neutralize the infectious CVB3 in a virus neutralization assay. Meanwhile, 2 virus strains were purified from CVB3 virus stock on the basis of their plaque size on HeLa cells. ICR mice were infected with the 2 purified virus strains (S-strain and L-strain) and unpurified virus stock (wild type) to analyze the difference in antibody responses against infections of purified and unpurified virus strains. The reactivity of antisera against each virus strain was tested by ELISA, and the results showed that the inoculation of purified virus strain induced a strong antibody response against the inoculated strain. As a result, the antibody response against wild-type and other virus strains was suppressed. These results suggest using unpurified virus stock as an antigen is advantageous for inducing a broad antibody response in inoculated animals.

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Shimoyama, T. , Kubota, T. , Shirai, J. and Watanabe, R. (2014) Evaluation of Antibodies Induced by the Injection of Single Capsid Protein or Purified Virus Particle of Coxsackievirus B3 in Mice. World Journal of Vaccines, 4, 164-174. doi: 10.4236/wjv.2014.44019.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] Domingo, E., Martin, V., Perales, C. and Escarmis, C. (2008) Coxsackieviruses and Quasispecies Theory: Evolution of Enteroviruses. Current Topics in Microbiology and Immunology, 323, 3-32.
[2] Chung, C.Y., Chen, C.Y., Lin, S.Y., Chung, Y.C., Chiu, H.Y., Chi, W.K., et al. (2010) Enterovirus 71 Virus-Like Particle Vaccine: Improved Production Conditions for Enhanced Yield. Vaccine, 28, 6951-6957.
[3] McMinn, P.C. (2012) Recent Advances in the Molecular Epidemiology and Control of Human Enterovirus 71 Infection. Current Opinion in Virology, 2, 199-205.
[4] Edlmayr, J., Niespodziana, K., Popow-Kraupp, T., Krzyzanek, V., Focke-Tejkl, M., Blaas, D., et al. (2011) Antibodies Induced with Recombinant VP1 from Human Rhinovirus Exhibit Cross-Neutralisation. European Respiratory Journal, 37, 44-52.
[5] Wu, C.N., Lin, Y.C., Fann, C., Liao, N.S., Shih, S.R. and Ho, M.S. (2001) Protection against Lethal Enterovirus 71 Infection in Newborn Mice by Passive Immunization with Subunit VP1 Vaccines and Inactivated Virus. Vaccine, 20, 895-904.
[6] DiMarchi, R., Brooke, G., Gale, C., Cracknell, V., Doel, T. and Mowat, N. (1986) Protection of Cattle against Foot-and-Mouth Disease by a Synthetic Peptide. Science, 232, 639-641.
[7] Pfaff, E., Mussgay, M., Bohm, H.O., Schulz, G.E. and Schaller, H. (1982) Antibodies against a Preselected Peptide Recognize and Neutralize Foot and Mouth Disease Virus. EMBO Journal, 1, 869-874.
[8] Wang, J.H., Liang, C.M., Peng, J.M., Shieh, J.J., Jong, M.H., Lin, Y.L., et al. (2003) Induction of Immunity in Swine by Purified Recombinant VP1 of Foot-and-Mouth Disease Virus. Vaccine, 21, 3721-3729.
[9] Domingo, E., Escarmis, C., Lazaro, E. and Manrubia, S.C. (2005) Quasispecies Dynamics and RNA Virus Extinction. Virus Research, 107, 129-139.
[10] Kawai, C. (1999) From Myocarditis to Cardiomyopathy: Mechanisms of Inflammation and Cell Death: Learning from the Past for the Future. Circulation, 99, 1091-1100.
[11] Zhang, L., Parham, N.J., Zhang, F., Aasa-Chapman, M., Gould, E.A. and Zhang, H. (2012) Vaccination with Coxsackievirus B3 Virus-Like Particles Elicits Humoral Immune Response and Protects Mice against Myocarditis. Vaccine, 30, 2301-2308.
[12] Henke, A., Wagner, E., Whitton, J.L., Zell, R. and Stelzner, A. (1998) Protection of Mice against Lethal Coxsackievirus B3 Infection by Using DNA Immunization. Journal of Virology, 72, 8327-8331.
[13] Henke, A., Zell, R. and Stelzner, A. (2001) DNA Vaccine-Mediated Immune Responses in Coxsackie Virus B3-In- fected Mice. Antiviral Research, 49, 49-54.
[14] Kim, J.Y., Jeon, E.S., Lim, B.K., Kim, S.M., Chung, S.K., Kim, J.M., et al. (2005) Immunogenicity of a DNA Vaccine for Coxsackievirus B3 in Mice: Protective Effects of Capsid Proteins against Viral Challenge. Vaccine, 23, 1672-1679.
[15] Fechner, H., Sipo, I., Westermann, D., Pinkert, S., Wang, X., Suckau, L., et al. (2008) Cardiac-Targeted RNA Interference Mediated by an AAV9 Vector Improves Cardiac Function in Coxsackievirus B3 Cardiomyopathy. Journal of Molecular Medicine, 86, 987-997.
[16] Watanabe, R., Hasegawa, A., Miyazawa, T., Kato, H. and Iwata, H. (2004) Generation of Monoclonal Antibodies to Porcine Interleukin 6 (PIL-6) Using the Recombinant PIL-6 Expressed in Escherichia coli. Journal of Veterinary Medical Science, 66, 1053-1057.
[17] Lindberg, A.M., Stalhandske, P.O. and Pettersson, U. (1987) Genome of Coxsackievirus B3. Virology, 156, 50-63.
[18] Lindberg, A.M., Crowell, R.L., Zell, R., Kandolf, R. and Pettersson, U. (1992) Mapping of the RD Phenotype of the Nancy Strain of Coxsackievirus B3. Virus Research, 24, 187-196.
[19] Pfaff, E., Thiel, H.J., Beck, E., Strohmaier, K. and Schaller, H. (1988) Analysis of Neutralizing Epitopes on Foot-and-Mouth Disease Virus. Journal of Virology, 62, 2033-2040.
[20] Beatrice, S.T., Katze, M.G., Zajac, B.A. and Crowell, R.L. (1980) Induction of Neutralizing Antibodies by the Coxsackievirus B3 Virion Polypeptide, VP2. Virology, 104, 426-438.
[21] Strebel, K., Beck, E., Strohmaier, K. and Schaller, H. (1986) Characterization of Foot-and-Mouth Disease Virus Gene Products with Antisera against Bacterially Synthesized Fusion Proteins. Journal of Virology, 57, 983-991.
[22] Strohmaier, K., Franze, R. and Adam, K.H. (1982) Location and Characterization of the Antigenic Portion of the FMDV Immunizing Protein. Journal of General Virology, 59, 295-306.
[23] Lee, C.D., Yan, Y.P., Liang, S.M. and Wang, T.F. (2009) Production of FMDV Virus-Like Particles by a SUMO Fusion Protein Approach in Escherichia coli. Journal of Biomedical Science, 16, 69.
[24] Page, G.S., Mosser, A.G., Hogle, J.M., Filman, D.J., Rueckert, R.R. and Chow, M. (1988) Three-Dimensional Structure of Poliovirus Serotype 1 Neutralizing Determinants. Journal of Virology, 62, 1781-1794.
[25] Liew, M.W., Rajendran, A. and Middelberg, A.P. (2010) Microbial Production of Virus-Like Particle Vaccine Protein at Gram-Per-Litre Levels. Journal of Biotechnology, 150, 224-231.
[26] Getts, M.T., Richards, M.H. and Miller, S.D. (2010) A Critical Role for Virus-Specific CD8+ CTLs in Protection from Theiler’s Virus-Induced Demyelination in Disease-Susceptible SJL Mice. Virology, 402, 102-111.

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