Characterization of monoclonal antibodies against Japanese encephalitis virus

Abstract

Hybridoma cells were produced by fusing P3X63Ag 8.653 mouse myeloma cell line with spleen cells from BALB/c mice immunized with Japanese encephalitis 0E) virus, Nakayama-RFVL strain. The resulting 26 clones produced hemagglutination inhibition (HI) antibody against the homologous virus. In order to clarify the immunological characteristics of these 26 monoclonal antibodies, the HI reactivity of each was tested against the 6 flaviviruses; JE, Murray Valley encephalitis (MVE), Egypt 101 strain of West Nile (WN), St. Louis encephalitis (SLE), Russian spring-summer encephalitis (RSSE) and Dengue type 1 viruses.We obtained 14 JE type-specific monoclonal antibodies. In order to study the antigenic difference among JE virus strains, the 14 JE type-specific antibodies were tested against the 4 JE virus strains; Nakayama-RFVL, Nakayama-Yoken, JaGAr01 and Kamiyama strains.The results obtained are as follows:1. In the HI test against the 6 flaviviruses, the 26 monoclonal antibodies fell into 4 groups:1) 14 clones produced JE type-specific antibodies.2) 6 clones produced antibody reactive to JE and MVE viruses.3) 3 clones produced antibody reactive to 3 or 4 viruses in the WN JE group; WN, MVE, SLE and JE viruses.4) 3 clones produced antibody cross-reactive to all 6 flaviviruses.In this pattern of reactivity, it was suggested that JE virus was immunologically most closely related to MVE virus and more closely related to WN and SLE viruses than to RSSE or Dengue-1 viruses.2. According to the HI test against the 4 JE virus strains, the 14 JE type-specific monoclonal antibodies consisted of the following antibodies:1) 2 clones produced Nakayama strain-specific antibodies.2) 3 clones produced antibody which reacted to Nakayama and Kamiyama strains, and slightly or not at all to JaGAr01 strain.3) 1 clone produced antibody which reacted to Nakayama and jaGAr01 strains, but not to Kamiyama strain.4) 8 clones produced antibody which had almost the same titer against all 4 strains. There was antigenic variation among 3 strains; Nakayama, jaGAr01 and Kamiyama strains. However, there was little antigenic difference between Nakayama-RFVL and Nakayama-Yoken strains.3. According to the analysis of immunoglobulin secreted, the 26 monoclonal antibodies were 4 IgM, 8 IgG1, 10 IgG2a, 1 IgG2b, 2 IgG3 and 1 IgA. A majority of the monoclonal antibodies which belonged to the IgM class were strain-specific. However, the igG class antibodies reacted to all 4 JE virus strains, or to other flaviviruses.4. Chromosome analysis strongly indicated that the hybrid cell was made by the fusion of one spleen cell with one myeloma cell.