Objective: To evaluate the recognition of NS4B mimotope, as multiple antigen peptide (MAP), by dengue antibodies presents in serum samples from patients with different serotype infections. Methods: A MAP containing mimotope sequence was synthesized and used to evaluate the recognition of NS4B mimotope as MAP by a panel of 66 human sera from dengue cases by an indirect ELISA assay. Results: The MAP differentiated between sera from dengue viruses infected patients and sera from healthy individuals and the best reactivity was shown by serum from dengue type 3 virus patients. The recognition was more intense with serum from patients with secondary infection. Conclusions: The findings suggest the potential use of NS4B mimotope on the development of a multi-epitope diagnostic tool. These results are important for further immunogenicity studies.