An introduction to a novel
ATF-manufacturing Platform
filtration system
Microcarrier Process Unit Operations
Vaccines
Gene Therapy
Stem Cells
Microcarrier Screen Filter Module
• All USP Class VI materials (Polysulfone, Polyester, epoxy, 316L SS)
• 70um screen
• Special flow path design to reduce fouling and attachment
• Multiple low shear, rapid, separations, without settling:
– Media exchanges, wash steps
– Seed transfer
– Harvest: cell debris & virus filtration from microcarriers
Microcarriers: Process Overview
Reactor seed train 5x volumes
X
X
Maybe ?
SIP: PBS & ucarriers
50+ flasks…
Wash ucarriers
SIP: PBS & ucarriers
Innoculate
Wash ucarriers
SIP: PBS & ucarriers
Batch/perfusion growth
Innoculate
Wash ucarriers
Trypsinization
Batch/perfusion growth
Innoculate
Wash
Trypsinization
Batch/perfusion growth
Cell transfer
Wash
Media reduction
Cell transfer
Infection
Media addition
Harvest
3
Microcarrier Processes: ATF System Advantages
• Major advantage is liquid and microcarrier
handling: ease and speed to remove or
transfer media or cells
– Rapid liquids removal without shear
– Wash steps for carriers
– Low shear cell/seed transfers
– Rapid Harvest
– Microcarriers do not need to settle
• Cell ”health” improved, allowing higher
productivity per cell
• Media removal can be to a low volume
• Simple to operate and scales to manufacturing
• External system which allows easy exchange
of filter if required
4
Microcarriers: ATF2 System Setup
5
Microcarriers: ATF10 System Setup
6
Microcarriers: Preparation & Washing
1. Reactor filled with
carriers and PBS
2. Reactor is SIP’d
3. ATF is autoclaved
4. Steamed connection
made
1. PBS removed at 1vv/hr for 40-50 mins
2. Addition of media
3. Diafiltrate for 5 minutes
ATF rate constant
at 4-5x filtrate rate
4. Close harvest, media added to
working volume
5. Innoculate with cells
7
Microcarriers: Perfusion
1. Perfusion carried out at small or large scale
2. Rates at 0.5vv/day to 4vv/day
3. ATF rate at 30+ higher than filtrate rate
8
Microcarriers: Seed Transfer
1.
2.
3.
4.
5.
Cell growth to maximum in batch or perfusion mode
Media reduction at 1vv/hr for 30-40 mins
Trypsinise cells
Harvest cells through ATF leaving behind carriers
Diafiltrate with new media, to capture maximum
number of cells
6. Cells transferred directly to next reactor
7. ATF rate at 3-5x filtrate rate
9
Microcarriers: Media Exchange, Infection, Harvest
1. Remove media at 1vv/hr
for 40-50 mins
2. Addition of new media
3. Infect cells with virus
4. Optional: Perfusion
1. Harvest cells and debris at 1
vv/hr for 40-50 mins
ATF rate constant
at 4-5x filtrate rate
2. Diafiltrate for 10-20 minutes
3. Optional: Clarification with
ATF and 0.2u filter on
holding tank
10
Adherent Cells: Combined DSP
Buffer, Wash
Vaccine Process – Adherent Cells
Ionic Solution
(hc-dna precipitation)
70u filtered
stream
0.2u or 0.5u
filtered stream
Production
reactor
Microcarrier-free
Harvest
Clarified
Harvest