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A gradient-free method for the purification of infective dengue virus for protein-level investigations
Stephanie M Jensen1, Celina T Nguyen1, John C Jewett1
1The Department of Chemistry and Biochemistry, The University of Arizona, 1306 E. University Blvd., Tucson, AZ 85721, USA.
Abstract:
Dengue virus (DENV) is a mosquito-transmitted flavivirus that infects approximately 100 million people annually. Multi-day protocols for purification of DENV reduce the infective titer due to viral sensitivity to both temperature and pH. Herein we describe a 5-h protocol for the purification of all DENV serotypes, utilizing traditional gradient-free ultracentrifugation followed by selective virion precipitation. This protocol allows for the separation of DENV from contaminating proteins - including intact C6/36 densovirus, for the production of infective virus at high concentration for protein-level analysis.
Insights
A new 5-hour Dengue virus (DENV) purification protocol enhances infective virus yield. This rapid method improves viral concentration for crucial protein-level analysis.
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- Dengue virus (DENV) is a significant global health threat, infecting millions annually.
- Conventional DENV purification methods are lengthy and reduce viral infectivity due to sensitivity to temperature and pH.
Purpose of the Study:
- To develop a rapid and efficient protocol for purifying all Dengue virus serotypes.
- To improve the yield of infective DENV for downstream protein-level analyses.
Main Methods:
- A novel 5-hour purification protocol combining gradient-free ultracentrifugation and selective virion precipitation.
- Separation of DENV from contaminating proteins, including C6/36 densovirus.
Main Results:
- Successful purification of all DENV serotypes within 5 hours.
- High concentration of infective DENV achieved, suitable for protein analysis.
- Effective removal of contaminating proteins and intact C6/36 densovirus.
Conclusions:
- The developed protocol offers a significantly faster alternative for DENV purification.
- This method preserves viral infectivity and yields high concentrations of purified DENV.
- Enables advanced protein-level investigations of Dengue virus.
