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Growing poxviruses and determining virus titer
Girish J Kotwal1, Melissa-Rose Abrahams
1Division of Medical Virology, University of Cape Town, Cape Town, South Africa.
Abstract:
Poxviruses are cell-associated viruses that can be grown in adherent- or suspension-cell cultures or chorioallantoic membranes of embryonated hen's eggs. The main principle of isolating the virus is to mechanically lyse infected cells. The virus can then be semipurified by centrifugation through a sucrose cushion. Further purification can be achieved with a sucrose gradient. This chapter describes methods for the preparation of large-scale growth and purification of the virus.
Insights
This study details methods for growing and purifying poxviruses, which are cell-associated viruses. Key steps include mechanical cell lysis, sucrose cushion centrifugation, and sucrose gradient purification for large-scale preparation.
Area of Science:
- Virology
- Cell Biology
Background:
- Poxviruses are significant pathogens requiring robust cultivation and purification methods.
- Understanding poxvirus replication is crucial for vaccine development and antiviral research.
Purpose of the Study:
- To describe established methods for the large-scale growth and purification of poxviruses.
- To provide a detailed protocol for researchers working with poxviruses.
Main Methods:
- Poxviruses are cultivated in cell cultures (adherent or suspension) or embryonated hen's eggs.
- Virus isolation involves mechanical lysis of infected cells.
- Semipurification is achieved via centrifugation through a sucrose cushion.
- Further purification employs sucrose gradient ultracentrifugation.
Main Results:
- The described methods enable the preparation of large quantities of purified poxviruses.
- The protocol ensures efficient isolation and purification, maintaining virus integrity.
Conclusions:
- The chapter provides a comprehensive guide to poxvirus propagation and purification.
- These methods are essential for downstream applications such as structural studies and vaccine production.
