Biology Reference
In-Depth Information
100
75
50
25
0
Crude
feed
Clarified
feed
Concentrated
feed
HIC semipurifed
material
AEX purifed
material
Purity
Fig. 1
CAV-2 downstream processing. Crude CAV-2 vector stocks are subjected
to a series of DSP steps in order to attain the desired level of purity. With each
additional step vector purity considerably improves but the overall process yield
declines as shown in the scheme
2
Materials
2.1 Clarification
by Dead-End
Microfiltration
1. Peristaltic pump, silicone tubings, and pressure gauge.
2. ULTA™ cap PP membrane (0.6 μm) capsule filtration device
(GE Healthcare, Uppsala, Sweden).
3. Milli-Q H
2
O.
4. Buffer A: 20 mM Tris-HCl, 2 mM MgCl
2
, 10 mM NaCl,
2.5 % glycerol, pH 8 filter-sterilized buffer.
2.2 Concentration
by Cross-Flow
Ultrafiltration
1. Advanced Midjet
®
system (GE healthcare).
2. Hollow fiber MidGee
®
cartridge, MWCO 100,000 (GE
Healthcare).
3. Connecting silicone tubing (size 14).
4. Diafiltration buffer: 20 mM Tris-HCl, 2 mM MgCl
2
, 10 mM
NaCl, 2.5 % glycerol, pH 8 filter-sterilized buffer.
5. Cleaning solutions: 0.5 M NaOH, 20 % EtOH, Milli-Q H
2
O.
6. Storage solution: 20 % EtOH.
2.2.1 Midjet
®
System
(Up to 200 mL)
1. Quixstand
®
system (GE healthcare) and Watson Marlow peri-
staltic pump.
2. Hollow fiber Xampler
®
cartridge, MWCO 100,000 (GE
Healthcare).
3. Connecting tubing (size 18).
4. Diafiltration buffer: 20 mM Tris-HCl, 2 mM MgCl
2
, 10 mM
NaCl, 2.5 % glycerol, pH 8 filter-sterilized buffer.
2.2.2 Quixstand
®
System
(Up to 10 L)
