Eppendorf In touch with life

Transcription

1 Eppendorf In touch with life

2 Scalable Cell Culture Using Rigid-Wall, Single-Use Bioreactors Eppendorf BioBLU Single-Use Vessels analytica, Dr. Christof Knocke, May 10, 2016

3 Sample Handling Cell Handling Liquid Handling The Eppendorf Experience The Eppendorf Competence Areas Manual pipettes Electronic pipettes Pipette tips Automated pipetting systems Dispensers Combitips Pipette controllers Fermentors and bioreactors Single-use bioreactors Cell culture and imaging consumables CO 2 incubators Biological shakers Photometers and Spectrometers Micromanipulators and injectors Electroporators Centrifuges and vacuum concentrators Rotors and accessoires Mixers and temperature control systems Thermal cyclers Ultra-low temperature freezers Multiwell plates Test tubes 3

4 Premium partner in life sciences History of Bioprocessing at Eppendorf 70 years of experience in the life science market Foundation of Eppendorf- Netheler-Hinz GmbH Acquisition of New Brunswick Scientific Co Acquisition of DASGIP GmbH Foundation of New Brunswick Scientific Co Foundation of DASGIP GmbH Eppendorf Bioprocess 4

5 Scalable Systems Our 3S Value Proposition Scalable systems Comprehensive Single-use options Ready-to-use Software solutions Innovative Eppendorf bioprocess systems cover a wide range of working volumes in autoclavable, single-use and SIP. Ready-to-use single-use bioreactors of different volumes complement the scale range. Comprehensive software packages offer innovative solutions for advanced process control and information management. 5

6 Scalable Systems Scalable Systems Bioprocessing needs scalable systems > Process optimization (feeding, gassing, ) in the milliliter-scale > Pilot/production scale in up to thousands of liters > Scale-down model development to implement changes The primary scope of this project was to investigate the scale-up capabilities of Eppendorf BioBLU Single-Use Vessels for cell culture 6

7 Scale-Up Criteria

8 Demands and objectives Bioprocess Scale-Up Criteria Geometry > Geometrical numbers Performance > Mixing > Tip speed, shear stress > Power input, > Mixing times > Gassing (k L a) 8

9 Eppendorf BioBLU Single-Use Vessels

10 Single-use options BioBLU Single-Use, but Rigid Wall: The Best of Both Worlds > Inspired by industrial rigid-wall designs > Typical stirred-tank (STR) dimensions > Fast and efficient mixing with magnetically coupled overhead drives > USP Class VI certified and animal component free material, e.g. virgin monolayer polystyrene Scalable and reproducible results 10

11 BioBLU c Family BioBLU c Concept Weldable or easy to connect tubing Overlay and sparge gas options to best suit your application Full featured highly integrated headplate Effective exhaust treatment, heat blanket or liquid-free (Peltier) Magnetic drive assembly Standard impeller sizes and vessel dimensions Non-invasive sensors 11

12 Single-use options BioBLU c The Widest Range of Rigid-Wall, Single-Use STRs BioBLU 0.3c BioBLU 1c BioBLU 3c BioBLU 50c Working volume ml 320 ml 1.25 L L L Stirring rpm rpm rpm rpm System/controller DASbox BioFlo 320, DASGIP BioFlo,DASGIP, adaptor kits BioFlo/CelliGen, adaptor kits R&D and Scale Down Product/Process Process Development Pilot Scale Production 12

13 Scalable Systems Geometry Vessel name 0.3c 1c 3c 50c Working vol : total vol Impeller diameter : Vessel ID V max height / vessel ID *: V max height = height from bottom of the vessel to liquid top surface at maximum vessel volume. 13

14 > Tip speed range > Numerical CFD analysis Mixing

15 Scalable Systems Tip Speed > Tip speed as criteria for the comparison of vessels > Scale-up based on maintaining a constant tip speed is common approach for shear sensitive cells > Range of available speeds by the controllers limits the considered tip speeds in the range of 0,3 0,7 m/s Tip speed (m/s) 0.3c (rpm) 1c (rpm) 3c (rpm) 50c (rpm)

16 Scalable Systems Scientific Cooperation Numerical investigation on the Eppendorf BioBLU 0.3c, 1c, 3c, and 50c Single-Use Vessels for cell culture ZHAW Zurich University of Applied Sciences Department Life Sciences und Facility Management Institute of Biotechnology, Biochemical Engineering and Cell Cultivation Technique, Wädenswil Scientists: > Prof. Dr. Dieter Eibl > Cedric Schirmer > Valentin Jossen 16

17 Scalable Systems CFD Modeling > Computational fluid dynamic investigations to predict power numbers and shear gradients BioBLU 50c mesh. Unstructured hexahedron volume mesh (A), and stirrer surface mesh (B) of the BioBLU 50c. The darker domain around the stirrer shows the multiple reference frame zone (MRF) for the implementation of the stirrer rotation. For the investigation of the acting forces on the stirrer blades, a regular finely woven mesh was used at the stirrer regions. 17

18 Scalable Systems CFD Fluid Pattern Simulation at a tip speed of 0,5m/s, maximum working volume 0.3c 1c 3c 50c Steady state fluid flow pattern of the BioBLU 0.3c, 1c, 3c and 50c cell culture systems. Steady state fluid flow pattern of the BioBLU 0.3c, 1c, 3c and 50c at a stirrer speed of 0.5 m/s at y-z-planes. The velocities are scaled to 90 % of the theoretical tip speeds. 18

19 Scalable Systems Power Input > Computational fluid dynamic investigations to predict power numbers and shear gradients > Identified scalable range of power inputs in function of tip speeds Power inputs of the BioBLU cell culture systems. Power inputs against tip speeds. 19

20 Scalable Systems Shear Gradients > The mean, local shear gradients as a function of impeller number, agitation speed, and agitation direction are below a critical range > Shear gradients of 500 s -1 to 5000 s -1 are considered to be harmful for cell cultures (Yim and Shamlou, 2000) 20

21 Scalable Systems k L a Values > k L a-based scale-up can be performed in the h -1 range BioBLU Single-Use Vessel k L a-values determined with the static gassing out method 21

22 > Case Study I: CHO cell culture 5L, 50L > Case Study II: CHO cell culture and protein production 250 ml, 3.75 L Case Studies

23 Case Study I Case Study I > Scale-up of CHO cell culture using New Brunswick CelliGen BLU stirred-tank bioreactors equipped with 5 L and 50 L single-use vessels. Application note AA257; Nick Kohlstrom, Joseph Capone, and Ma Sha, Eppendorf Inc., Enfield, CT, U.S.A. 23

24 Case Study II Case Study II Scale up of SEAP production with CHO XM suspension cells using the BioBLU 0.3c and BioBLU 5c single use bioreactors ZHAW Zurich University of Applied Sciences Department Life Sciences und Facility Management Institute of Biotechnology, Biochemical Engineering and Cell Cultivation Technique, Wädenswil Scientists: > Prof. Dr. Dieter Eibl > Katharina Blaschczok 24

25 Case Study II Case Study II Setup SEAP (secreted alkaline phosphatase) expressing CHO suspension cell line CHO XM Scale-up criteria > Equal mixing time > Equal sufficient oxygen mass transfer Schematic presentation of the BioBLU 0.3c bioreactor and feeding line setup. 25

26 Case Study II Case Study II Cell Growth and Protein Production Production volume: 250 ml Production volume: 3.75 L 26

27 Case Study II Case Study II Glucose and Lactate Profiles Production volume: 250 ml Production volume: 3.75 L 27

28 Summary > Eppendorf BioBLU c Single-Use Vessels of different sizes are geometrically and proportionally similar > Broad overlapping tip speed zone and k L a values for excellent scalability > Offer a low-shear environment > Case studies demonstrated the suitability of Eppendorf BioBLU c Single-Use Vessels for cell culture scale up 28

29 We Know Bioprocessing Contact:

30 Disclaimer Eppendorf, the Eppendorf logo, In touch with life and BioBLU are registered trademarks of Eppendorf AG, Germany. New Brunswick is a trademark of Eppendorf AG, Germany. BioFlo and CelliGen are registered trademarks of Eppendorf, Inc., USA. DASGIP and DASbox are registered trademarks of DASGIP Information and Process Technology GmbH, Germany. All rights reserved, including graphics and photos. Copyright 2016 by Eppendorf AG. 30