Dengue virus (DEN), the pathogen behind dengue hemorrhagic fever, remains a public health problem in Asia and South America. Method: Monoclonal antibodies (MAbs) specifi cally against dengue serotype 2 were generated by fusion of immunized mouse spleen cells with NS1/1-Ab4-1 myeloma cells. DEN-2 strain PL046, a local strain isolated from Taiwanese dengue fever patients, was used in the immunization. Results: The specifi cities of MAbs were determined by Western blot and ELISA assays. Six anti-E and twenty-one anti-NS1 MAbs were generated from mice immunized with PL046 strain, two of the six anti-E MAbs strongly inhibited infection by PL046 and 16681 strains. Ten of the twenty-one anti-NS1 MAbs were specifi cally against dengue serotype 2. Overall, three anti-E and twelve anti-NS1 MAbs were produced and characterized. The isotypes were also determined and described. Conclusions: The MAbs generated in this study may provide a powerful tool for antigenic analysis and clinical diagnosis.