Experimental Design
Experimental – CELLine
Media Tested
•
BalanCD
™
CHO Growth A
•
IS CHO-CD G10.7
Vessels Used
•
350 CELLine (350mL)
•
1000 CELLine (1000mL)
Seeding Density
•
1.5 x 10
6
cells/mL
Culture Protocols
•
28 days
•
Cell count & IgG quantification every 3-4 days
•
Complete media refreshed every 7 days, split cells 1:4
Control – Shake Flask
Media Tested
•
BalanCD
™
CHO Growth A (-/+) BalanCD
™
CHO Feed 1
•
IS CHO-CD G10.7 (-/+) BalanCD
™
CHO Feed 3
Vessel Used
•
125mL flask (30mL working vol.)
Seeding Density
•
0.3 x 10
6
cells/mL
Culture Protocols
•
14 days
•
Cell count daily and IgG quantification @ day 7, 10 and 14
•
Feed initial 5% working volume of feed to fed-batch conditions
on days 3-6
Seed
•
9 x 10
6
Cells
Day 3
•
Count
•
Sample IgG
•
Feed 5%
Day 4
•
Count
•
Feed 5%
Day 5
•
Count
•
Feed 5%
Day 6
•
Count
•
Feed 5%
Day 7
•
Count
•
Sample IgG
Day 10
•
Count
•
Sample IgG
•
Feed Glucose
Day 14
•
Count
•
Harvest IgG
Fed-Batch 125mL Shake Flask Process
CELLine 1000 Bioreactor Process
Seed
•
22.5 x 10
6
Cells
Day 3
•
Count
•
Sample IgG
Day 7
•
Count
•
Harvest IgG
•
Reduce Cells 1:4
•
Change Media
Day 10
•
Count
•
Sample IgG
Day 14
•
Count
•
Harvest IgG
•
Reduce Cells 1:4
•
Change Media
Day 17
•
Count
•
Sample IgG
Day 21
•
Count
•
Harvest IgG
•
Reduce Cells 1:4
•
Change Media
Day 24
•
Count
•
Sample IgG
Day 28
•
Count
•
Harvest IgG
Media Information
•
BalanCD
™
CHO Growth A is a chemically-defined growth medium, containing no animal-derived components. The formulation was
developed using Irvine Scientific’s Rational Culture Media Design
®
strategy with three model cell lines, and specifically designed to support
growth and productivity in a variety of CHO cells in both batch and fed-batch cultures.
•
IS CHO-CD G10.7 is part of Irvine Scientific’s CHO Media Development Kit. It is a chemically-defined growth media with no animal
derived components that was designed to help control metabolic overflow and support growth and productivity in a variety of CHO cells
in both batch and fed-batch cultures.
•
BalanCD
™
CHO Feed 1 and BalanCD
™
CHO Feed 3 are both chemically defined, animal component free and protein-free feed media
designed to provide cell culture nutrients for fed-batch applications in conjunction with BalanCD
™
CHO Growth A. These feeds have been
developed to promote cell growth and MAb production in a variety of CHO cells.
CELLine
™
Bioreactor Flask
Materials and Methods
Shake Flask Culture
Shake flasks (125mL) were used in conjunction with BalanCD
™
CHO Growth A or IS CHO-CD G10.7, 30mL initial working volume. Five percent feed of
BalanCD
™
CHO Feed 1 or BalanCD™ CHO Feed 3, was provided on days 3, 4, 5 and 6 for a total of 20% initial volume. Conditions: shake speed 120 RPM,
5% CO
2
, and 37°C humidified incubator. Cells were counted with Beckman Coulter Vi-Cell and IgG quantified by Forte’ Bio Octet kEQ. Initial seeding density
was 0.3 x 10
6
cells/mL.
CELLine Bioreactor Flask Culture
CELLine bioreactor flasks were used in conjunction with BalanCD
™
CHO Growth A or IS CHO-CD G10.7. Cells were split 1:4 every 7 days. Media was refreshed
every seven days. Conditions: 5% CO
2
, and 37°C humidified incubator. Cells were counted with Beckman Coulter Vi-Cell and IgG quantified by Forte’ Bio Octet
kEQ. Initial seeding density was 1.5 x 10
6
cells/mL.
