Monoclonal antibody raised against envelope glycoprotein peptide neutralizes Japanese encephalitis virus.

Epitopes on envelope glycoprotein of Indian strain of Japanese encephalitis virus were delineated by prediction methods. Monoclonal antibodies (MAb) raised against a putative B cell epitope peptide, reacted with the virion in ELISA and immunofluorescence assays. One MAb was also able to neutralize the virus. The reactivity of this MAb against a Sri Lankan strain was checked, since this strain had a substitution within the B cell epitope at position Egp 153 (G-->W). The MAb was able to bind to, but was not able to neutralize the Sri Lankan isolate. The data indicated that the predicted B cell epitope is a neutralizing epitope and may be included in a peptide-based vaccine against the virus.

An IgM monoclonal antibody to Japanese encephalitis virus recognizing a cross-reactive epitope on nuclear histones.

An IgM class of monoclonal antibody (MAb) raised against 'envelope' (E) glycoprotein of Japanese encephalitis (JE) virus, cross reacted with nuclear histones, in addition to recognizing the viral antigen present in the cytoplasm of infected cells by indirect fluorescent antibody (FA) technique. The experiments on histone depletion by the acid treatment of uninfected PS (porcine kidney) cells, revealed the loss of nuclear immunofluorescence (IF) which was regained after the reconstitution of acid treated cells with histones, prior-to reacting with MAb NHA-2. The IgM MAb recognized specifically the viral antigens expressed on the surface of JE virus infected PS cells by a modified indirect FA. The adsorption of MAb NHA-2 with calf thymus histones (type II-AS) showed a comparative higher drop in the reactivity to JE virus (54.2% reduction) as compared to that against uncomplexed histones (33.3%) by ELISA, thus indicating a higher MAb affinity to the former. In contrast, the adsorption of MAb with chicken RBC nuclei resulted in comparatively more reduction in the reactivity to the uncomplexed histones (52.4% reduction) as against JE virus (37.5%), suggesting that DNA plays some role in modifying and presenting these epitopes. The cross-linkage of epitopes by glutaraldehyde treatment of JE virus antigen and histones showed a 2-fold and higher rise in the MAb reactivity as against those with unfixed or methanol fixed antigens (no cross-linkage), suggesting that the epitope is conformation dependent. Thus, histones seem to share a partial conformational homology with 'E' glycoprotein of JE virus and immune reaction with histones might lead to an autoimmune disorder.

Monoclonal antibody to Japanese encephalitis virus cross-reacting with histones present in the cell nuclei.

An immunoglobulin G (IgG2b) class of monoclonal antibody (MoAb, NHA-1) raised against Japanese encephalitis virus (JEV) E glycoprotein, reacted with the viral antigen expressed in cytoplasm of the infected cells and also with the cell nuclei, by an indirect fluorescent antibody technique (FA). The NHA-1 reactivity to nuclei was found to be due to its recognizing a JEV cross-reactive epitope present on the nuclear histones. Adsorption with calf thymus histones (type II-AS) showed a drop in NHA-1 reactivity to both JEV and histones by an enzyme-linked immunosorbent assay (ELISA) and indirect FA; the drop was higher against the histones. The MoAb recognized specifically the viral antigens expressed on the infected porcine kidney cell surface by a modified indirect FA. ELISA carried out with glutaraldehyde-fixed antigens showed an almost 2-fold increase in the reactivity over unfixed JEV antigen but none for the histones. Thus, the results indicate that histones share a sequential homology with E glycoprotein of JEV, which might lead to an autoimmune disorder induced due to the molecular mimicry between these two antigens.

Epitope analysis of strains of Japanese encephalitis virus by monoclonal antibodies.

Twenty one strains of Japanese encephalitis (JE) virus, including 16 from India, were compared antigenically on the basis of their reactivity in immunofluorescence (IF), haemagglutination inhibition (HI), ELISA with captured antigen (ECA), and neutralization (N) tests with JE monoclonal antibodies (MAbs). These MAbs represented three domains of distinct epitopes on the envelope protein, designated as Hs-1 to 4 (JE specific in HI), Hx-1 to 5 (flavivirus cross reactive in HI) and NHs-1 to 2 (non-HI JE virus specific). Fifteen of the 21 strains studied were placed in group I. These reacted with MAbs representing the three domains in all the tests indicating presence of the three types of epitopes with full functional activity. The remaining six strains were placed in group II and showed loss in HI reactivity with Hs MAbs but not with Hx MAbs. All the group II strains also reacted in IF and ECA with NHs-1. Hs epitopes in three strains, G9473 (Tamil Nadu), 641686 (Tamil Nadu) and 822199 (Karnataka), appeared to have mutated partially, indicating loss in HI reactivity with Hs MAbs only, while there was retention of other reactivities, viz., IF, ECA and to some extent N test with G9473 and 641686. The remaining three strains, 691004 (Sri Lankan), 755468 (West Bengal) and Yoken (Japan) of group II showed almost complete loss of Hs-1 and Hs-2 epitopes as there was absence of reactivity in IF, ECA and N test in addition to HI. However, Hs-3 MAb showed reactivity in IF with these strains.(ABSTRACT TRUNCATED AT 250 WORDS)