Growth Kinetics of Rabies Virus in BHK-21 Cells Using Fluorescent Activated Cell Sorter (FACS) Analysis and a Monoclonal Antibody Based CellELISA
Journal Title: Journal of Immunology and Vaccine Technology - Year 2014, Vol 1, Issue 1
Abstract
Introduction: Rabies is a highly fatal, neurotropic viral zoonosis of warm-blooded animals including man caused by rabies virus belonging to genus Lyssavirus of the family Rhabdoviridae under order Mononegavirales. Now-a-days rabies cell culture vaccines are being used as a curative and prophylactic immunization in animals. The present investigation aimed at refinement/improvement of the existing cell culture rabies vaccine protocol, envisaging critical investigation on kinetics of growth of Pasteur virus strain in BHK-21 clone 13 cells. Materials & Methods: Mouse monoclonal antibodies were generated against rabies virus for the development of a cell-ELISA as an in-process control assay. Pasteur virus strain of rabies virus was ropagated in BHK-21 cells. Virus samples were collected at different time intervals for investigation of growth kinetics using ell-ELISA, FAT and fluorescence activated cell sorter analysis. Results: The monoclonal antibody based cell-ELISA developed during the present study may serve as a useful assay with proven repeatability for in-process monitoring of growth of rabies virus in BHK-21 cells. This assay although less sensitive, correlated well with FAT for the determination of optimal period of harvesting the virus. Fluorescence Activated Cell Sorter (FACS) analysis indicated its suitability and higher level of sensitivity for monitoring of rabies virus propagation in BHK-21 cells. Growth pattern of the virus using all the three tests indicated that virus as eclipse phase at around 9-12 h post infection. The study on rabies virus growth kinetics suggested that the optimum quantity of infectious virus particles could be harvested at around 48 h of infection using 0.1 multiplicity of infection of rabies virus. Conclusion: The findings of the present investigation if applied successfully are likely to impact the production protocol f cell culture rabies vaccine with the elimination of hurdles like concentration of virus and use of mice for vaccine virus titration, etc.
Authors and Affiliations
Anandan Paldurai
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