vaccines (regardless if they are used as a substrate for virus propagation and re-

combinant DNA products or as vaccine components). A detailed description of the

cell bank system including procedures used to avoid microbial/viral contamina-

tion, safety precautions in case of catastrophic events, and storage conditions must

be provided [89]. Moreover, the cells of the MCB must be completely character-

ized and must be able to show a complete history. Characterization includes, but is

not limited to, biochemistry (cell surface markers, mRNA, etc.), specific identifiers

(morphology, serotype, etc.), karyology and tumorgenicity, virulence markers,

genetic markers, and media (e.g., serum) [89]. The WCB is directly derived from

the MCB and characterized for cell viability before use in the manufacturing

process. In contrast to the MCB, documentation outlining procedures for storage

and assays used for qualification, purity, and characterization has to be provided

for each new WCB [89].

5.9.2

VIRUS SOURCES

Similar to cellular sources, seed lot systems including a master seed lot (MSL) and a

working seed lot (WSL) are typical components of a vaccine production process. A

full characterization of the WSL is required, but depends on several factors such as

the nature of the virus strain/vector, genetic modifications, and the history of the cell

source used in seed lot preparation [90]. Nevertheless, genetic and phenotypic

properties of the virus including a side-to-side comparison with the parental virus

must be included. Genetic characterization typically involves next-generation se-

quencing of the virus, PCR analysis, southern blotting, and restriction mapping.

Moreover, verification of possible endogenous retroviruses present in the viral

genome is critical to ensure that no additional virus material is introduced. Special

attention should be paid regarding genetic stability ensuring no changes in regions

involved in e.g., attenuation. For this, genetic stability studies of the vaccine seed of

a passage level comparable to a production batch should be conducted. Phenotypic

characterization focuses on in vitro and in vivo studies measuring markers for at-

tenuation/modification and expression of heterologous antigens. Particularly for

viral vaccines, infectious virus titers, total particle numbers, virus yields, and in vivo

growth characteristics in a suitable animal model are a critical part of this char-

acterization. WSL are usually prepared by passage of the MSL in the cellular source

used in the production process. Here, special attention must be paid regarding

passage numbers, ideally limiting the number of passages to minimize the possi-

bility of genetic and phenotypic changes of the virus [90].

5.9.3

CELL GROWTH AND HARVESTING

Production processes for various vaccine types are likely to be similar. In many

cases, there will be minimal downstream processing and the basic requirements for

control and manufacturing are the same [90]. For large-scale production, the

working cell bank as well as the seed virus may have to be expanded before in-

oculation of the production cell culture. As discussed above, the number of passages

between WSL and production lot should be kept low. Whatever the final production

Upstream processing for viral vaccines

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