62
Viruses Used in Virus Validation Studies
Viruses that potentially may contaminate plasma, i.e., HIV,
WNV, HAV, and B19V, or their model viruses, BVDV and
CPV as specific model viruses for HCV and B19V, respec-
tively, and the herpes virus
Pseudorabies virus
(PRV), an
enveloped DNA virus as HBV, are chosen for virus validati-
on studies. Thus, assurance is provided that a wide range
of viruses with different physicochemical properties, that
could possibly be present in plasma, would be inactivated
and removed during the manufacturing process.
Methodology of Virus Reduction Validation
Major steps in the manufacturing process for Berinert
®
were validated in a virology laboratory. The starting materi-
al was obtained directly from the manufacturing plant and
was “spiked” with a known quantity of test virus, and the
virus inactivation/removal capacity of that step was measu-
red. The virus reduction factors for each step were then ad-
ded to result in the overall reduction factor for the process.
Results of virus validation studies are reported as x log
10
reductions. The value x is calculated by comparing the
virus load in the starting material to the virus load in
the material after the virus inactivation/removal step.
The log
10
of the ratio between these loads is the reduc-
tion factor. Each increment of 1 corresponds to a 10-fold
reduction in load. Thus, a log
10
reduction factor of 5 means
that the number of viruses remaining after the process has
been reduced 100,000-fold (10
5
-fold).
The virus reduction capacity of the two dedicated virus re-
duction steps pasteurization (in aqueous solution at 60ºC
for 10 hours) and virus filtration (nanofiltration) as well as
the HIC step was evaluated in a series of
in vitro
spiking
experiments. The mean overall virus reduction ranged from
≥
14.5 to
≥
19.9 log
10
as shown in Table 18. These validati-
on data for Berinert
®
show that pasteurization as well as
HIC and virus filtration (nanofiltration) effectively remove
a broad spectrum of viruses. This underscores the high
margin of safety of Berinert
®
in regard to the transmission
of viruses.
Virus
Manufacturing Stage
Virus Load of Spiked
Starting Material (log
10
)
Virus Load of
Final Sample (log
10
)
Virus Reduction
Factor (log
10
)
Overall Virus Reduction
Factor (log
10
)
HIV
Pasteurization
HIC
20N/15N virus filtration
6.1
8.5
7.6
≤
-0.5
≤
4.0
≤
2.5
≥
6.6
≥
4.5
≥
5.1
≥
16.2
BVDV
Pasteurization
HIC
20N/15N virus filtration
7.7
6.5
6.8
≤
-1.5
≤
1.8
≤
1.5
≥
9.2
≥
4.7
≥
5.3
≥
19.2
PRV
Pasteurization
HIC
20N/15N virus filtration
6.9
8.3
8.6
0.6
≤
1.8
≤
1.5
6.3
≥
6.5
≥
7.1
≥
19.9
WNV
Pasteurization
20N/15N virus filtration
7.5
9.5
≤
0.5
≤
1.5
≥
7.0
≥
8.0
≥
15.0
HAV
Pasteurization
HIC
20N/15N virus filtration
5.9
6.6
6.8
≤
-0.5
3.8
≤
1.5
≥
6.4
2.8
≥
5.3
≥
14.5
B19V
Pasteurization
5.4
1.5
3.9
NA
CPV
Pasteurization
HIC
20N/15N virus filtration
7.1
8.5
8.7
5.7
2.1
≤
1.5
1.4
6.4
≥
7.2
≥
15.0
HIV=
Human immunodeficiency virus
; BVDV=
Bovine viral diarrhea virus;
PRV=
Pseudorabies virus;
WNV=
West nile virus;
HAV=
Hepatitis A virus;
B19V=
Parvovirus B19;
CPV=
Canine parvovirus;
HIC=Hydrophobic interaction chromatography; NA=not applicable
Table 18 – Virus Reduction Capacity of the Manufacturing Process of Berinert
® 112
