Related Experiment Video
Updated: Jul 19, 2026
Production of E. coli-expressed Self-Assembling Protein Nanoparticles for Vaccines Requiring Trimeric Epitope Presentation
Published on: August 21, 2019
Production, purification and characterization of recombinant dengue multiepitope protein
Nagesh K Tripathi1, Ambuj Shrivastva, Priyabrata Pattnaik
1Division of Virology, Defence Research and Development Establishment, Jhansi Road, Gwalior 474002, India. tripathink@gmail.com
Abstract:
Dengue is an acute mosquito-borne viral disease of humankind. Dengue fever, dengue haemorrhagic fever and dengue shock syndrome have become global public health problems in recent years. rDME-G (recombinant dengue multiepitope protein that can specifically detect IgG) was produced in a 5-litre fermenter in Escherichia coli for use in diagnosis. The culture was induced with 1 mM isopropyl beta-D-thiogalactoside and cells were further grown for 4 h before harvesting. After fermentation, dry cell weight resulted in approx. 16.2 g/l. The rDME-G protein was purified from inclusion bodies using affinity chromatography. The final yield of purified rDME-G protein from fermentation resulted in approx. 168 mg/l of pure biologically active rDME-G protein. The purity of rDME-G protein was checked by SDS/PAGE analysis and the reactivity of this protein was further determined by Western blotting. The purified protein was used to develop an in-house dipstick ELISA and tested using a panel of 60 patient sera characterized using the commercially available tests for detection of dengue antibody. We compared our results with IgG-capture ELISA (Pan-Bio, Windsor, QLD, Australia) and rapid IC (immuno-chromatography) test (Pan-Bio). By using rDME-G protein as an antigen, in the dipstick ELISA, the results were in excellent agreement with commercial rapid IC test and IgG capture ELISA. These results show that the product has a promising potential to be used for diagnosis of dengue in both laboratory- and field-based detection systems with minimum cost and a high degree of sensitivity and specificity.
Insights
A new recombinant dengue multiepitope protein (rDME-G) was developed for dengue diagnosis. This protein shows high sensitivity and specificity, offering a cost-effective solution for laboratory and field detection systems.
Area of Science:
- Virology
- Immunology
- Biotechnology
Background:
- Dengue fever, dengue hemorrhagic fever, and dengue shock syndrome are significant global public health concerns.
- Accurate and accessible diagnostic tools are crucial for managing dengue outbreaks.
Purpose of the Study:
- To produce and purify a recombinant dengue multiepitope protein (rDME-G) for diagnostic applications.
- To evaluate the efficacy of rDME-G in an in-house dipstick ELISA for detecting dengue-specific IgG antibodies.
Main Methods:
- Recombinant dengue multiepitope protein (rDME-G) was produced in Escherichia coli and purified using affinity chromatography.
- The purified rDME-G protein was utilized to develop a dipstick ELISA.
- The dipstick ELISA performance was validated against commercial IgG-capture ELISA and rapid immunochromatography tests using patient sera.
Main Results:
- High yield of purified, biologically active rDME-G protein was achieved (approx. 168 mg/l).
- SDS/PAGE and Western blotting confirmed the purity and reactivity of rDME-G.
- The in-house dipstick ELISA using rDME-G demonstrated excellent agreement with commercial diagnostic tests.
Conclusions:
- The developed rDME-G protein is a promising antigen for dengue diagnosis.
- The dipstick ELISA offers a cost-effective, sensitive, and specific method for dengue detection.
- This diagnostic approach has potential for both laboratory and field-based applications.
