Indian Journal of Virology

Dengue virus infection has recently taken endemic proportion in India implicating all the four known dengue serotypes. Since there is neither specific therapy nor any effective vaccine available, the timely and rapid diagnosis plays an important role in patient management. DIII is the receptor binding domain eliciting serotype specific neutralizing antibody response. Thus, there is a need to develop an alternate cost effective recombinant antigen to replace the whole virus antigen in diagnostic tests, which has several drawbacks. Envelope domain III gene of dengue virus type 3 was cloned in pQE 30UA expression vector and expressed in E. coli strain M15 (pREP4). The recombinant dengue-3 domain III protein was purified from inclusion bodies by affinity chromatography under denaturing conditions. The reactivity of this protein was checked by Western blot using patient serum sample. This protein was further evaluated by an in-house ELISA employing a panel of dengue infected patient serum samples. The final yield of dengue virus 3 recombinant domain-III protein was 13 mg/l. The result of in-house ELISA using recombinant Dengue 3 DIII protein was compared with that of Pan-Bio IgM capture ELISA and Pan-Bio IC test. The overall agreement among all three tests was found to be more than 90% with serum samples. The sensitivity of the in-house ELISA was found to be more than 92% and specificity with reference to both commercial assays was found to be 95%. This study showed that rD3DIII protein could be used as diagnostic reagent in ELISA for detection of dengue infection.