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Clinical and Vaccine Immunology, January 2006, p. 59-67, Vol. 13, No. 1
1071-412X/06/$08.00+0 doi:10.1128/CVI.13.1.59-67.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.
Recombinant Multiepitope Protein for Early Detection of Dengue Infections
Ravulapalli AnandaRao,1 Sathyamangalam Swaminathan,1 Sirimali Fernando,2 Asha M. Jana,3 and Navin Khanna1*International Centre for Genetic Engineering and Biotechnology, New Delhi-110067, India,1 Faculty of Medical Sciences, University of Sri Jayewardenepura, Gangodawilla, Nugegoda, Sri Lanka,2 Division of Virology, Defense Research and Development Establishment, Ministry of Defense, Government of India, Jhansi Road, Gwalior-474002, India3
Received 23 May 2005/ Returned for modification 15 August 2005/ Accepted 12 October 2005
Dengue fever is a mosquito-borne viral disease prevalent mainly in tropical countries. As the clinical manifestations of dengue are not very unique, laboratory diagnosis is crucial in identifying cases of dengue infection. Detection of dengue infection based on the identification of antidengue antibodies has emerged as a practical and reliable means of diagnosing dengue fever. We recently developed a customized recombinant dengue multiepitope protein (r-DME-G) that can specifically detect the immunoglobulin G (IgG) class of antidengue antibodies in patient sera. Using this strategy, we have now created another dengue multiepitope protein, r-DME-M, with specificity for the IgM class of antidengue antibodies. A synthetic gene encoding the r-DME-M protein was expressed as a maltose-binding protein fusion in Escherichia coli. The recombinant protein was purified in a single affinity chromatographic step to obtain yields of 
15 mg purified protein/liter of culture. The purified protein was used to develop an in-house IgM enzyme-linked immunosorbent assay (ELISA) and tested using a panel of 172 patient sera characterized using the commercially available Dengue Duo rapid strip test from PanBio, Australia. The IgM ELISA results showed that the r-DME-M protein not only recognized all IgM+ samples identified by the PanBio test but also identified samples missed by the latter test. We also successfully adapted the r-DME-M protein to a rapid strip test format. This approach of creating customized antigens coupled to overexpression in E. coli and simple purification offers a promising alternative option to dengue diagnosis with the potential to circumvent the drawbacks of the whole virus antigen-based commercial kits.
* Corresponding author. Mailing address: RGP Group, International Centre for Genetic Engineering and Biotechnology, P.O. Box 10504, Aruna Asaf Ali Marg, New Delhi-110067, India. Phone: 91-11-26177357, ext. 272. Fax: 91-11-26162316. E-mail: navin{at}icgeb.res.in.
Clinical and Vaccine Immunology, January 2006, p. 59-67, Vol. 13, No. 1
1071-412X/06/$08.00+0 doi:10.1128/CVI.13.1.59-67.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.
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