Construction of a chimeric secretory IgA and its neutralization activity against avian influenza virus H5N1

J Immunol Res. 2014:2014:394127. doi: 10.1155/2014/394127. Epub 2014 Feb 13.

Abstract

Secretory immunoglobulin A (SIgA) acts as the first line of defense against respiratory pathogens. In this assay, the variable regions of heavy chain (VH) and Light chain (VL) genes from a mouse monoclonal antibody against H5N1 were cloned and fused with human IgA constant regions. The full-length chimeric light and heavy chains were inserted into a eukaryotic expressing vector and then transfected into CHO/dhfr-cells. The chimeric monomeric IgA antibody expression was confirmed by using ELISA, SDS-PAGE, and Western blot. In order to obtain a dimeric secretory IgA, another two expressing plasmids, namely, pcDNA4/His A-IgJ and pcDNA4/His A-SC, were cotransfected into the CHO/dhfr-cells. The expression of dimeric SIgA was confirmed by using ELISA assay and native gel electrophoresis. In microneutralization assay on 96-well immunoplate, the chimeric SIgA showed neutralization activity against H5N1 virus on MDCK cells and the titer was determined to be 1 : 64. On preadministrating intranasally, the chimeric SIgA could prevent mice from lethal attack by using A/Vietnam/1194/04 H5N1 with a survival rate of 80%. So we concluded that the constructed recombinant chimeric SIgA has a neutralization capability targeting avian influenza virus H5N1 infection in vitro and in vivo.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Neutralizing*
  • Antigens / immunology
  • Antigens / metabolism
  • CHO Cells
  • Cricetulus
  • Gene Expression
  • Gene Order
  • Genetic Vectors / genetics
  • Humans
  • Immunoglobulin A, Secretory / genetics*
  • Immunoglobulin A, Secretory / immunology*
  • Immunoglobulin A, Secretory / isolation & purification
  • Immunoglobulin A, Secretory / metabolism
  • Influenza A Virus, H5N1 Subtype / immunology*
  • Mice
  • Neutralization Tests
  • Orthomyxoviridae Infections / immunology
  • Orthomyxoviridae Infections / mortality
  • Orthomyxoviridae Infections / virology
  • Protein Binding / immunology
  • Recombinant Fusion Proteins*

Substances

  • Antibodies, Neutralizing
  • Antigens
  • Immunoglobulin A, Secretory
  • Recombinant Fusion Proteins