Two flaviviruses, Japanese encephalitis (JE) virus and Dengue (DEN) virus which have high pathogenicity for humans, continue to pose a serious public health problem in tropical and subtropical countries of the world. In order to identify the immunodominant B-cell epitopes for diagnostic application, we have prepared a series of 15-mer synthetic peptides from JE virus core protein based on computer analysis. Four linear, immunodominant epitopes corresponding to amino acids 91-105 (P78), 1-15 (P73), 8-22 (P74), and 34-48 (P75) of JE virus core proteins were identified by employing an enzyme-linked immunosorbent assay (ELISA), using high-titered immune sera from JE-vaccinated children. P78 was found to be the most immunodominant. The sero-specificity of these peptides was tested by binding to seroconverted samples from JE and DEN-1 patients. P78 and P74 belonged to group-specific epitopes which reacted with both JE and DEN-1 patient sera. P73 and P75 belonged to subcomplex-specific epitopes which reacted only with JE but not with DEN-1 patient sera. The study suggests that these peptides corresponding to the immunodominant epitopes of JE virus core protein might have the potential to be used as peptide-based diagnostic reagents for the detection and differentiation of JE and DEN antibody responses.