English
 
User Manual Privacy Policy Disclaimer Contact us
  Advanced SearchBrowse

Item

ITEM ACTIONSEXPORT
  Flow cytometric monitoring of influenza A virus infection in MDCK cells during vaccine production

Schulze-Horsel, J., Genzel, Y., & Reichl, U. (2008). Flow cytometric monitoring of influenza A virus infection in MDCK cells during vaccine production. BMC Biotechnology, 8, 45. doi:10.1186/1472-6750-8-45.

Item is

Basic

show hide
Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0013-95FC-C Version Permalink: http://hdl.handle.net/11858/00-001M-0000-002C-8B36-5
Genre: Journal Article

Files

show Files
hide Files
:
eDoc_327772_2008.pdf (Publisher version), 460KB
Name:
eDoc_327772_2008.pdf
Description:
-
Visibility:
Public
MIME-Type / Checksum:
application/pdf / [MD5]
Technical Metadata:
Copyright Date:
-
Copyright Info:
© 2008 Schulze-Horsel et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Locators

show

Creators

show
hide
 Creators:
Schulze-Horsel, J.1, Author              
Genzel, Y.1, Author              
Reichl, U.1, 2, Author              
Affiliations:
1Bioprocess Engineering, Max Planck Institute for Dynamics of Complex Technical Systems, Max Planck Society, ou_1738140              
2Otto-von-Guericke-Universität Magdeburg, ou_1738156              

Content

show
hide
Free keywords: -
 Abstract: Background In cell culture-based influenza vaccine production the monitoring of virus titres and cell physiology during infection is of great importance for process characterisation and optimisation. While conventional virus quantification methods give only virus titres in the culture broth, data obtained by fluorescence labelling of intracellular virus proteins provide additional information on infection dynamics. Flow cytometry represents a valuable tool to investigate the influences of cultivation conditions and process variations on virus replication and virus yields. Results In this study, fluorescein-labelled monoclonal antibodies against influenza A virus matrix protein 1 and nucleoprotein were used for monitoring the infection status of adherent Madin-Darby canine kidney cells from bioreactor samples. Specific binding of monoclonal antibodies was shown for influenza A virus strains of different subtypes (H1N1, H1N5, H3N8) and host specificity (human, equine, swine). At high multiplicity of infection in a bioreactor, the onset of influenza virus replication was detected at about 2 to 4 h post infection by immunocytometry. In contrast, a significant increase in titre by hemagglutination assay was detected at the earliest 4 to 6 h post infection. In addition to the quantification of the infected cell fraction, influenza A virus nucleoprotein and matrix protein copy numbers in cells could be estimated from the measured fluorescence intensities. The fluorescence intensity threshold for infected cell quantification was equivalent to 76 000 nucleoprotein and matrix protein 1 copies per cell, which corresponds to 19 virions at the most. Conclusions It is shown that immunocytometry is a sensitive and robust method for the monitoring of viral infection and the quantification of virus protein content in fixed cells from bioreactor samples. Therefore, it is a valuable addition to other detection methods of influenza virus infection such as immunotitration and RNA hybridisation. Thousands of individual cells are measured per sample. Thus, the presented method is believed to be quite independent of the concentration of infected cells (multiplicity of infection and total cell concentration) in bioreactors. This allows to perform detailed studies on factors relevant for optimization of virus yields in cell cultures. The method could also be used for process characterisation and investigations concerning reproducibility in vaccine manufacturing. © 2008 Schulze-Horsel et al; licensee BioMed Central Ltd. [accessed June 6, 2008]

Details

show
hide
Language(s): eng - English
 Dates: 2008
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Method: -
 Identifiers: eDoc: 327772
Other: 13/08
DOI: 10.1186/1472-6750-8-45
 Degree: -

Event

show

Legal Case

show

Project information

show

Source 1

show
hide
Title: BMC Biotechnology
Source Genre: Journal
 Creator(s):
Affiliations:
Publ. Info: BioMed Central
Pages: - Volume / Issue: 8 Sequence Number: - Start / End Page: 45 Identifier: ISSN: 1472-6750
CoNE: /journals/resource/111000136906066