assays are used for a different genotype of the virus, both show the
same features necessary for assay application.
2.3.2 Quantification
of Contaminants
The quantification of contaminants is performed by using assays in
the 96-well microplate format. Reading is handled in a plate reader
(Cytation3 [BioTek, Winooski, VT, USA]).
Residual Free DNA (dsDNA) (See Note 2)
l
Quant-iT™ PicoGreen™ dsDNA Kit (Thermo Fisher Scientific,
Waltham, MA, USA).
l
Plate
shaker
(Thermomixer
compact
[Eppendorf,
Hamburg, GER]).
l
96-Well plates, flat bottom, black (Greiner CELLSTAR
®
[Sigma-Aldrich, St. Louis, MO, USA]).
l
Sterile, nuclease-free water (Thermo Fisher Scientific, Waltham,
MA, USA).
Total Protein via Lowry (BCA) Assay (See Note 3)
l
Pierce™ BCA Protein Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA).
l
Plate
shaker
(Thermomixer
compact
[Eppendorf,
Hamburg, GER]).
l
Incubator (HERAcell™ 240i [Thermo Fisher Scientific]),
37
C.
l
96-Well plates, flat bottom, transparent (Greiner CELLSTAR
®
[Sigma-Aldrich, St. Louis, MO, USA]).
l
Water.
l
PBS for dilutions (Biochrom, Berlin, GER).
2.3.3 Clarification via
Depth Filtration
The DSP process is performed using the benchtop filtration system
SARTOFLOW
® Slice 200 Benchtop System (Sartorius, Go ¨ttingen,
GER). The system should be equipped with a peristaltic pump with
a Tandem 1082-pump head (Sartorius, Go ¨ttingen, GER) (see Note
4). Pressures are measured with SciLog
® SciPres
® pressure sensors,
connected to the pump.
l
Sartopure
® PP3 particle reduction filter element: size 8 (0.12 m
2
filtration area; 3 μm retention rate; filter material: polypropylene
fleeces); single-use (Sartorius, Go ¨ttingen, GER).
l
Sartopure
®
PP3 particle reduction filter element: size
4 (0.013 m
2 filtration area; 0.64 μm retention rate; filter material:
polypropylene
fleeces);
single-use
(Sartorius,
Go ¨ttingen, GER).
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373