requested that manufacturers specifically consider widely distributed viruses, which

are difficult to inactivate (e.g., PCV and PPV) and with zoonotic potential (e.g.

HEV); and that manufacturers conduct viral-risk assessment according to the

general principles outlined in Ph. Eur. 5.1.7 (Viral Safety). The same re-

commendation is also made for the analysis of serum in the EMA guideline on

viral-risk analysis [26]. This guidance strongly recommended that manufacturers of

biological medicinal products conduct risk analyses, taking into account the quality

and properties of the serum batches and the impact of these serum batches on the

quality of the finished product, prior to use.

4.3.7

RAW MATERIALS: FROM BIOLOGICAL MATERIALS TO CHEMICALLY DEFINED

MATERIALS FOR CELL LINES

Serum and trypsin are biological materials with potentially significant safety risks.

Following the strategic imperative of removing non-essential animal-derived ma-

terials, manufacturers of biologics including vaccines progressively have been de-

veloping serum-free cell-culture media. Some plant components have been used

such as rice extract. However, this was not the ideal solution because many new

plant viruses have also been identified over the last 20 years [27]. Chemically

defined cell-culture media is also being developed. Hence, over the years, upstream

manufacturing processes for new vaccines are becoming free of animal-derived

components. Cell and virus banks have benefited from this approach as well. It is

clear that the preferred option of risk management for cell culture media is to

prevent and remove the risks rather than to mitigate through detection.

4.4

ANALYTICAL CONSIDERATIONS AND CELL-BASED ASSAYS

4.4.1

SAFETY TESTS AND FUTURE

A panel of tests are requested to be performed by manufacturers at relevant stages

of the production process, to confirm that the biological products, and the biological

starting materials from which they are manufactured, are free of adventitious

agents. This extensive testing for adventitious agents prevents major contamination

events and potential adverse clinical consequences.

4.4.1.1

In Vitro and in Vivo Safety Tests

The current in vitro and in vivo technologies for the detection of specific ad-

ventitious contaminants of vaccines were developed more than 50 years ago.

The current in vitro virus-detection assays use different cell lines in accordance

with the relevant regulatory guidance (e.g., WHO, FDA guidance, Eur P, and USP,

etc.). The assays work by incubating the test sample in the cell culture and then

observing subsequent cytopathic effects (CPEs), or hemadsorption (HAD) or he-

magglutination (HA), by introducing red blood cells. In vivo virus-detection assays

require the inoculation of the sample in an animal susceptible to infection by the

virus under consideration. The presence of virus is demonstrated by animal mor-

tality or the presence of hemagglutinins in tissue at sites of potential infection.

70

Bioprocessing of Viral Vaccines