Sartorius - November 2021 - Simplifying Adeno Associated Virus - 17

Titer 96 h
01.5
1
The cells were inoculated in 24 microbioreactors
2 hours before infection
at 27 ˚C, DO 30%, and stirring speed
750 rpm. We infected cells at a range
of VCCs using a baculovirus ratio: 1:1
and a MOI Baculovirus Helper | Baculovirus
Vector of 0.1:0.1 and lysed the
cells using a range of Lysis Reagent 2
concentrations.
0.5
-0.5
We identified that the optimum conditions
in this run produced high rAAV
titers > 5 × 10¹¹ vg/mL, as well as high
VCCs in Sf9 Medium 2 with all process
conditions (Figure 7).
We used the MODDE®
-1
software to
-1.5
Sf9 Medium 1
Sf9 Medium 2
Ratio
Cell Densitiy
Lysis Reagent 1Lysis Reagent 2Ratio×Ratio
Titer 96 h (N=22, DF=16, R2=0.99)
Titer 120 h
01.5
Using the software, we analyzed these
factors to generate a dynamic profile
plot and found the multi-factor setpoint
for optimum rAAV productivity
is a VCC at infection of 8 × 10⁶ cells/mL
and 1% Lysis Reagent 2.
1
generate coefficients plots for titer
and found that the multiple factors
which had the most positive impact
on titer are Sf9 Medium 2, VCC and
Lysis Reagent 2 concentration.
0.5
-0.5
-1
Figure 6: Coefficients plots for rAAV vector
genome titer versus a range of cell culture
factors obtained from Sf9 cells in Ambr®
15
-1.5
Sf9 Medium 1
Sf9 Medium 2
Titer 120 h (N=22, DF=16, R2=0.99), Confidence = 0.95
Ratio
Cell Densitiy
Lysis Reagent 1Lysis Reagent 2Ratio×Ratio
microbioreactors. Titers were determined at
96 hours (up) and 120 hours (down) after
infection with two baculoviruses.
17 | GENengnews.com
Titer [vg/mL]
Titer [vg/mL]

Sartorius - November 2021 - Simplifying Adeno Associated Virus

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