Several H-2 defined cell lines were examined for their ability to support infection and replication of Japanese encephalitis virus (JEV) before their use in and stimulation protocols for generating cytotoxic T lymphocytes (CTLs) against JEV. Among 11 different cell lines tested, two H-2 macrophage tumour lines (P388D1, RAW 264.7), an H-2 hybridoma (Sp2/0), an H-2KD neuroblastoma (Neuro 2a), and H-2 fibroblast cell line (L929) were found to support JEV infection and replication. These cell lines were used to generate anti-JEV CTLs by using immunization followed by stimulation of BALB/c mice. We observed that not only syngeneic and allogeneic infected cells but also JEV-infected xenogeneic cells could prime BALB/c mice for the generation of JEV-specific CTLs upon subsequent stimulation of splenocytes with JEV-infected syngeneic cells. Although infected xenogeneic cells were used for immunization, the anti-JEV effectors that were generated lysed infected syngeneic targets but not JEV-infected xenogeneic or allogeneic target cells in a 5 h Cr release assay. These anti-JEV effectors recognized syngeneic target cells infected with West Nile virus to a lesser extent and were shown to be Lyt-2.2 T cells. The results of unlabelled cold target competition studies suggested alterations in the cell surface expression of viral antigenic determinants recognized by these CTLs. We further demonstrate that the JEV-specific CTLs generated could virtually block the release of infectious virus particles from infected P388D1 and Neuro 2a cells .


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