Abstract
Analysis of human monoclonal antibodies (mAbs) developed from influenza infected donors have enormously contributed to the identification of neutralization sensitive epitopes of influenza virus. The HA protein is a crucial target of neutralizing antibodies and at monoclonal level only Abs binding to HA have been able to neutralize the virus. In this study, eight A (H1N1)pdm 2009 seropositive patients within the age range of 20-50 years (median = 36 years) were recruited. Two anti-HA mAbs secreting stable clones, 2D8 and 2F12 were established under optimized conditions from the peripheral blood mononuclear cells (PBMCs) of the volunteers. These antibodies efficiently neutralized the homologous laboratory isolated strain of the pandemic virus as well as the reference strain. Our study suggests that the anti-HA antibodies derived from infected Indian patients display neutralization potential against the A(H1N1)pdm 2009 virus. This is the first ever study of generation of mAbs against the pandemic influenza virus involving the immune repertoire if Indian patients. Molecular characterization of the target regions will help in identifying potential immunogens in the Indian pandemic isolates and confer protective immunity against this virus.
Footnotes
Selection and peer-review under responsibility of the 8th Vaccine Conference Organizing Committee.
References
- 1.Friesen R.H.E., Koudstaal W. New Class of Monoclonal Antibodies against Severe Influenza: Prophylactic and Therapeutic Efficacy in Ferrets. PLoS ONE. 2010;5:e9106. doi: 10.1371/journal.pone.0009106. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2.Ge Q., McManus M.T. RNA interference of influenza virus production by directly targeting mRNA for degradation and indirectly inhibiting all viral RNA transcription. Proc Natl Acad Sci U S A. 2003;100:2718–2723. doi: 10.1073/pnas.0437841100. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Wang T.T., Tan G.S. Broadly Protective Monoclonal Antibodies against H3 Influenza Viruses following Sequential Immunization with Different Hemagglutinins. PLoSPathog. 2010;6:e1000796. doi: 10.1371/journal.ppat.1000796. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Fukuda K., Bridges C.B., Brammer T.L., Izurieta H.S., Cox N.J. Morbid Mortal Wkly Rep. 1999;48:1–28. [Google Scholar]
- 5.French H. Human serum in influenza. Br Med J. 1919:575. [Google Scholar]
- 6.Sawyer L.A. Antibodies for the prevention and treatment of viral diseases. Antiviral Res. 2000;47:57–77. doi: 10.1016/s0166-3542(00)00111-x. [DOI] [PubMed] [Google Scholar]
- 7.Weltzin R., Monath T.P. Intranasal antibody prophylaxis for protection against viral disease. Clin. Microbiol Rev. 1999;12:383–393. doi: 10.1128/cmr.12.3.383. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Gorny M.K. Production of human monoclonal antibodies via fusion of Epstein-Barr virus-transformed lymphocytes with heteromyeloma. In: Celis J.E., editor. Cell Biology: A Laboratory Handbook. Academic Press; Waltham: 1994. pp. 276–281. [Google Scholar]
- 9.Funaro A., Gribaudo G. Generation of potent neutralizing human monoclonal antibodies against cytomegalovirus infection from immune B cells. BMC Biotechnol. 2008;8:85. doi: 10.1186/1472-6750-8-85. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Traggiai E., Becker S., Subbarao K. An efficient method to make human monoclonal antibodies from memory B cells: potent neutralization of SARS coronavirus. Nat Med. 2004;10(8):871–875. doi: 10.1038/nm1080. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Rowe T., Abernathy R.A. Detection of antibody to avian influenza A (H5N1) virus in human serum by using a combination of serologic assays. J Clin Microbiol. 1999;37:937–943. doi: 10.1128/jcm.37.4.937-943.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Reed L.J., Muench H. A simple method of estimating fifty percent endpoints. The Am J Hyg. 1938;27:493–497. [Google Scholar]
- 13.Casali P., Inghirami G. Human monoclonals from antigen specific selection of B lymphocytes and transformation by EBV. Science. 1986;234:476–479. doi: 10.1126/science.3020687. [DOI] [PubMed] [Google Scholar]
- 14.Kohler G., Milstein C. Continuous cultures of fused cells secreting antibodies of pre defined specificity. Nature. 1975;7:495–497. doi: 10.1038/256495a0. [DOI] [PubMed] [Google Scholar]
- 15.Steinitz M., Klein G., Koskimies S., Makel O. EB virus induced B lymphocyte cells lines producing specific antibody. Nature. 1977;269:420–422. doi: 10.1038/269420a0. [DOI] [PubMed] [Google Scholar]
- 16.Kozbor D., Lagarde A.E., Rodr J.C. Proc Natl. acad. Sci. U.S.A. 1982;79:6651. doi: 10.1073/pnas.79.21.6651. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 17.Yu X., McGraw P.A. An optimized electrofusion-based protocol for generating virus-specific human monoclonal antibodies. J Immunol Methods. 2008;336:142–151. doi: 10.1016/j.jim.2008.04.008. [DOI] [PMC free article] [PubMed] [Google Scholar]
