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Method Article
Edward Hutchinson
Fodor Group, University of Oxford
Corresponding Author
Ervin Fodor
Fodor Group, University of Oxford
Corresponding Author
License:
This work is licensed under a CC BY-NC 3.0 License. Read Full License
It is often useful to concentrate and purify the pleomorphic, enveloped virions of influenza viruses prior to analysis. This protocol describes a standard method for purifying ‘spherical’ influenza virions, the most common morphology of laboratory-adapted strains, by rate-zonal ultracentrifugation through a discontinuous density gradient. The protocol, which takes around eight hours to complete, is designed for virions shed into the growth media of infected cells, and would also be suitable for concentrating virions from the allantoic fluid of embryonated chicken eggs. Two different density gradients are described: a conventional gradient of sucrose and a gradient of iodixanol (OptiPrep™ density gradient medium). A high degree of virion purity can be achieved, but small quantities of contaminants will remain. These can be removed by an optional haemadsorption-elution (HAd) step, which takes an additional one and half hours. This allows extremely efficient removal of the original contaminants, but introduces other, distinct contaminants at low levels.
Keywords
Influenza, ultracentrifugation, haemadsorption
The authors declare no competing financial interests.
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Posted 26 Aug, 2014
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Subject Areas
Microbiology
Method Article
Edward Hutchinson
Fodor Group, University of Oxford
Corresponding Author
Ervin Fodor
Fodor Group, University of Oxford
Corresponding Author
Version History
CURRENT STATUS: Posted
Version 1
Posted 26 Aug, 2014
No community comments so far
Metrics
Comments: 0
HTML Views: ...
Subject Areas
Microbiology
License:
This work is licensed under a CC BY-NC 3.0 License. Read Full License
It is often useful to concentrate and purify the pleomorphic, enveloped virions of influenza viruses prior to analysis. This protocol describes a standard method for purifying ‘spherical’ influenza virions, the most common morphology of laboratory-adapted strains, by rate-zonal ultracentrifugation through a discontinuous density gradient. The protocol, which takes around eight hours to complete, is designed for virions shed into the growth media of infected cells, and would also be suitable for concentrating virions from the allantoic fluid of embryonated chicken eggs. Two different density gradients are described: a conventional gradient of sucrose and a gradient of iodixanol (OptiPrep™ density gradient medium). A high degree of virion purity can be achieved, but small quantities of contaminants will remain. These can be removed by an optional haemadsorption-elution (HAd) step, which takes an additional one and half hours. This allows extremely efficient removal of the original contaminants, but introduces other, distinct contaminants at low levels.
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