Researcher

Matthias Meininger
Laura Fischer

Additional Information

Collaborations:
Bioprocess Engineering at the Otto-von-Guericke-University, Magdeburg

Start:
2005/01/01

End:
2010/01/01

Affinity Chromatography of Influenza Virus

Affinity Chromatography of Influenza Virus

Affinity chromatography is a method of selectively and reversibly binding target compounds e.g. proteins to immobilized molecules on solid support matrices based on the exploitation of known physico-chemical and biological affinities between the interacting molecules. The interaction is based on structural properties, such as the interaction of antibodies with antigens, lectins with carbohydrates, enzymes with substrate analoges, hormones with receptors, and nucleic acids with binding proteins.

In our laboratory we are designing and optimizing different affinity matrices for the downstream processing of cell culture based (Madin Darby canine kidney cells) influenza vaccines. Currently studied affinity interactions are centered on lectins, antibodies and chemical analoges of enzymatic substrates.

Lectin-affinity chromatography (LAC) is based on a selective affinity of lectins, which are carbohydrate specific proteins of non-immune origin, for a carbohydrate or a group of carbohydrates. The influenza A virus contains two spike glycoproteins on its surface: hemagglutinin (HA) and neuraminidase (NA). HA is the most abundant surface protein and also the main antigenic protein of the influenza A virus. It is a trimeric glycoprotein containing per subunit 3 to 9 N-linked glycosylation sites depending on the viral strain. Detailed analysis of these sites and their individual glycan structures are presently performed in house.
Lectins immobilized on different column and membrane based supports have been successfully applied for the capturing of various influenza virus strains from MDCK cell culture broths. Thereby contaminating host cell nucleic acid as well as proteins could be highly reduced while virions were concentrated.

Fig 1. Target process schema for downstream processing of cell culture derived influenza virus vaccine production Zoom Image
Fig 1. Target process schema for downstream processing of cell culture derived influenza virus vaccine production

Chemical substrate analoges of the surface glycoprotein neuraminidase represent also an interesting capturing tool for viral particles of influenza. A considerable advantage of chemically synthesized ligands is their relatively low production costs compared to recombinant proteins. Therefore, a specific capture step based on chemical analoges of the neuraminidase substrates would allow an exciting economization of the overall cell-culture based vaccine production processes.

Immunoaffinity chromatography harnesses the specificity and avidity of the interaction between an antigen and its antibody to purify the antigen. It is a powerful tool for the specific capturing of viral particles and virally encoded membrane proteins direct from the cell culture broth. However, due to antigen drifts immunoaffinity based virus capturing has a disadvantage compared to other affinity interactions.

Fig 2. Light scattering signal (90°, Dawn EOS, Wyatt Technology Inc.) during LAC purification of MDCK cell derived human influenza A virus Zoom Image
Fig 2. Light scattering signal (90°, Dawn EOS, Wyatt Technology Inc.) during LAC purification of MDCK cell derived human influenza A virus [less]

Literature:
Opitz, L., Salaklang, J., Büttner, H., Reichl, U., Wolff, M.W. 2007. Lectin-affinity chromatography for downstream processing of MDCK cell culture derived human influenza A viruses. Vaccine 25: 939-947.
Opitz, L., Lehmann, S., Zimmermann, A., Reichl, U., Wolff, M.W. 2007. Impact of adsorbents selection on capture efficiency of cell culture derived human influenza viruses. Journal of Biotechnology 131: 309-317.
Wolff, M.W., Opitz, L., Reichl, U. Method for purification of viral proteins. European Patent: Application number: EP07007759, Application date: 17.04.2007 USPTO: Application number: US 60/912,305; Application date: 17.04.2007

Interconnections with other projects of Bioprocess Engineering

Coupled Processes:

Influenza Vaccine Production in Microcarrier Systems
Downstream Processing of Influenza Virus
Particulate formulation of influenza virus antigen

 
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