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Study of a Novel Microcarrier Scale-up Culture of Vero Cells

Author: FengJian
Tutor: QiNianMin
School: Shanghai Jiaotong University
Course: Pharmaceutical Engineering
Keywords: Viral vaccines Vero cells Microcarrier Bioreactor Bead to bead
CLC: TQ464
Type: Master's thesis
Year: 2012
Downloads: 178
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Abstract

The rapid development of the vaccine industry provides a strong guarantee for the prevention of human diseases. The mostly widely used vaccines include viral vaccines, bacterial vaccine and so on currently, among which, the viral vaccines are the earliest, most widespread, and most various ones. The conventional processes for viral vaccines production with primary cells and roller bottles are labor-intensive, with low virus titers and different in quality from batch to batch. Since 1967 the microcarrier culture technology made it possible to produce biological products with adherent animal cells automatically in large scale culture, Which has become the trend of the future development of adherent mammalian cells in large-scale culture. Many years ago the foreign scientists had begun to develop the large scale bioreactor culture process of passaged cell lines for production of viral vaccines, In our country , however, it has just started in small and lab scale stage in recent years.Vero cells is a passaged cell line of the African green monkey kidney cells, which is non-tumorigenic ,easy for large scale culture and stable in the nucleus types with no exogenous factors of pollution , the WHO recommended it strongly many years ago as the vaccine production substrate and it is already widely used in viral vaccines production. Cytodex microcarriers are microspheres produced by treating the surface of cross-linked dextran with or denatured collagen, which have become the most widely used cell culture microcarriers all over the world.The microcarrier culture process of Vero cells involves a lot of key technolies, among which the bioreactor culture modes and the scaling-up are the hottest.In the study, we first investigated the Vero cells culture in 2D and 3D batch culture, anylising the key points of Vero cell microcarrier culture. Later, we studied the feasibility of Vero cell recirculation culture process with spinner flasks by manually simulation. Then with regard to the key problems in scaling-up bioreactor culture of Vero cells on microcarries,we put forward a fully new idea of microcarrier bead-to-bead scale up culture and compared it with the two common methods to prove the feasibility. First, the key points of Vero cell microcarrier batch culture were analysed, in the recirculation culture,the highest cell density was achieved after 7 day culture ,with the recirculation rate from 2 VVD to 3VVD. In 24 wells and Petri dishes, we defined the“half digested status with trypsin”of Vero cells on microcarriers with trypsin detaching the cells on microcarriers 5 minutes, then we scaled up the culture and compared the new-way-treated Vero cell culture with the two formerly used bead-to-bead methods. After 8 days culture, Our results showed that the half-digested Vero cells with trypsin treating 5 min can effectively improve the efficiency of the bead to bead effiency of Vero cells, achieving a higher cell density of 7.9×105 /mL and a lower bare microcarrier percentage with 15%。The study offers a new idea for the development of the process of Vero cell bioreactor culture and bead to bead scale up culture, while providing clues for the development of a continuous in-situ bioreactor culture of Vero cells.

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