The ambr™ advanced microscale bioreactor, from The Automation Partnership
The Penridge Suite, London, N11 1NL, UK : Thursday, 20th Oct 2011 09:00 - 17:00
Following on from the success of the 2010 event, Professor Gary Lye is Chairing the second meeting in the series “Miniaturisation - Micro Scale Bioprocess Development”. Miniaturisation and automation of bioprocess development continues to be a rapidly expanding area of interest since the technologies promise to reduce biopharmaceutical development time and cost. This meeting aims to highlight recent technologies used in high throughput bioprocess development, from clone selection through to analysis of final product and formulation. A series of expert speakers will describe the development and use of current miniaturisation technologies together with the technical and regulatory hurdles that must be overcome to facilitate wider industrial uptake.
Meeting Chair: Professor Gary J Lye, PhD, CEng, CSci, FIChemE, Director Industrial Doctoral Training Centre, Deputy Head of Department, The Advanced Centre for Biochemical Engineering, Department of Biochemical Engineering, University College London, UK
This event has CPD accreditation and will have a discussion panel session.
9:00 – 9:45 Registration
9:45 – 10:00 Introduction by the Chairs: Professor Gary J Lye, University College London
10:00 – 10:30 Advanced microscale bioreactor (ambrTM): Concepts, applications and field data.
Dr Barney Zoro, TAP Biosystems, UK
Implementation of high throughput automated systems is recognised as a valuable approach in the field of bioprocess development. The ambrTM system combines the benefits of single use labware and automated liquid handling to provide a high throughtput microscale bioreactor system with capability for both batch and fed-batch cell cultures. This talk introduces the ambr, its potential in bioprocess development and, through discussion of field data gathered to date, demonstrates the effectiveness of the ambr in CHO cell culture applications: Clone Screening, Process Optimisation, and Design of Experiments.
10:30 – 11:00 Integration and usage of chemical optical sensors into small scale cultivation vessels
Dr Gernot Thomas John, PreSens Precision Sensing GmbH
Monitoring of important culture parameters is indispensable / essential for bioprocess development and production. Chemical optical sensors allow to measure oxygen, pH and CO2 non-invasively and in real time. This technology contributes to the optimization of culture media and reproducible culture conditions in the main applications of biotechnology. PreSens Precision Sensing GmbH offers a variety of sensors for integration in smallscale vessels. The talk will describe the basics of chemical optical sensors and show examples of their integration into vessels of different sizes & shapes. In addition, data of these systems will be presented showing the efficiency of these
measurement tools.
11:00– 11:30 Mid-morning break and Poster Viewing
11:30 – 12:00 Microbioreactor Development: driven by demand and the availability of enabling technologies.
David Laidlaw, Small Scale Technologies Manager, Applikon Biotechnology.
Micro-scale bioreactors exist only because of a demand-driven intersection of several enabling, but unrelated, technologies. Having come into mainstream existence only six (or so) years ago, this market segment of instruments now has many solution-providers competing to meet the needs of a global scientific community. Though some commonalities exist, instruments presented to the market have been as diverse in their design as they have been in their success. This talk will first review general miniature reactor formats and their enabling technologies to highlight considerations necessary before their implementation into process development efforts. A new 24 deep square well plate will be introduced and links to resources available to help users implement such tools will be provided. Finally, a micro-dose liquid feeding technology, applicable over a wide range of bioreactor vessel sizes, will be introduced and its potential for application to controlled process development at the small scale will be presented.
12:00 – 12:30 Making the Link Between Small Scale Cell Culture Technologies and Larger Scale Bioreactors
Professor Gary J Lye, University College London
Miniature and small scale bioreactor technologies generally operate at scales ranging from 0.1-10 mL (fluidic or shaken systems) to 50–500 mL (stirred systems). In contrast most conventional bioreactors operate at 2-5 L scale or above. PreviousUCLresearch has addressed the characterisation of miniature shaken and stirred bioreactor technologies and their application to the fed-batch cultivation of mammalian cells. Most recently we have invested in a new Responsive Bioprocessing Facility to address research and training on single-use bioreactors. This presentation will summarise our experience in working with a range of automated, miniature and single-use bioreactor technologies. In particular it will be shown how it is possible to scale-up predictively between shaken, rocked and stirred bioreactors of different size and geometry.
12:30 – 13:30 Lunch, and Poster Viewing
13:30 - 14:30 Question and Answer Session and Speakers photo
Delegates will be asked to submit questions to a panel of experts. Questions can be submitted before the event or on the day
14:30 - 15:00 How the usage of modern small scale cultivation technology improves clone and process development
Carsten Mueller, m2p-labs, UK
In a first part this talk will focus on the engineering parameters relevant for cultivation and their impact on clone selection and the first steps of process optimisation. At present, numerous technologies are available to enable the screening of clones under process relevant conditions. This empowers our users now to select the most promising strains for a later production already at the earliest stage. In the second part we will focus on how an online monitored micro bioreactor system (BioLector) can help you to speed up your process optimisation and make this step more efficient. In addition, currently available popular liquid handling systems can be used to boost this step even further. Even though this talk will focus on microbial cell cultivation the use of BioLector technology is not limited to growth of microorganisms. BioLector is also designed for the cultivation of insect and mammalian cell cultures.
15:00 - 15:30 Afternoon Tea/Coffee, and Last Poster Viewing
15:30 – 16:00 Optimisation of downstream bioprocess development – Applications of High throughput technology
Dr Martyn Hulley, Eden Biodesign Ltd, Liverpool, UK
16:00 – 16:30 High throughput techniques in downstream processing
Sharon Williams, Product Development and Downstream Processing Manager, ProMetic BioSciences Ltd. UK
16:30 – 17:00 Microfluidic Bioreactor Platforms for Early Bioprocess Development
Dr Nicolas Szita, Biochemical Engineering, University College London, UK
Microfluidic bioreactors offer exciting new opportunities to accelerate early bioprocess development. These include the reduced use of resources such a culture media, the capacity for real-time monitoring and control of process variables, ease of sterilisation via disposable polymer technology, reduced amount of labour due to automation, and increased throughput via parallelisation. This talk will present recent examples of microfluidic bioreactors for a range of applications in biocatalytic synthesis, suspension culture and regenerative medicine
17:00 Chairman’s summing up
Media Partners
Keywords: bioreactor, micro, automated, culture, single use, Microbioreactors, Well Plate, Liquid Feed,
Meeting Web Site: www.regonline.co.uk/mini2011
About the chair
Gary Lye is Professor of Biochemical Engineering within the Advanced Centre for Biochemical Engineering at University College London (UCL). He is Deputy Head of Department and Director of the Industrial Doctoral Training Centre (IDTC) in Bioprocess Engineering Leadership. He received his PhD in Biotechnology from the University of Reading in 1992. Between 1993 and 1996 he was successively a Research Fellow and then Lecturer in Chemical Engineering at Imperial College London and the University of Edinburgh. He joinedUCL in 1996. He has broad research interests on the application of microscale and automation techniques to rapid bioprocess design, optimisation and scale-up.
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Registration Web Site: www.regonline.co.uk/mini2011
About the Speakers
David Laidlaw’s graduate studies were in the Biotechnology Laboratory at the University of British Columbia, Vancouver, B.C. Mr. Laidlaw has worked as an Applications Specialist at MicroReactor Technologies and later in Fermentation Process Development at Genentech Inc. Dave uses his research and process development background to apply an end-user perspective to upstream screening and technology development at Applikon Biotechnology Inc.
Gernot Thomas John studied chemistry in Saarbrücken / Germany and did a doctorate with Professor Elmar Heinzle on the input of chemical-optical sensors in screening biotechnological processes. From 2000 till 2005 he worked as product manager with PreSens Precision Sensing GmbH in Regensburg / Germany. At the same time he graduated a correspondence course as industrial engineer (university of applied sciences). The topic of his diploma thesis was the distribution of innovative industrial goods. In 2005 he started as acting manager for the business area “Histology Solutions” with Carl Zeiss MicroImaging GmbH. In 2007 he returned to PreSens Precision Sensing GmbH as Director Marketing and Innovations. Currently he focuses on business development
Barney Zoro’s primary focus is delivering high quality bioreactor technology which meets or exceeds needs in the cell culture aspects of bioprocess development, including new technology developments. Barney’s personal and professional goal is to ensure industry wide acceptance and use of progressive bioprocess technologies, demonstrating technical capability and value to businesses, through an evidence-based approach. Barney read Chemical Engineering at Cambridge University and continued his study at University College London with a Masters degree and a Doctorate in Biochemical Engineering, with a focus on Tissue Engineering. He has published papers in Biotechnology and Bioengineering and presented a number of conference posters.
Carsten Müller studied mechanical engineering with the focus on biochemical engineering at the RWTH Aachen (Rheinisch-Westfaelische Technische Hochschule). He did his diploma work as DAAD stipendiary at the Indian Institute of Technology (IIT) in Delhi on the separation of proteins using ultrafiltration techniques. After his studies he worked as doctoral student at the chair of Biochemical Engineering (Prof. Jochen Büchs) at RWTH Aachen on the development and validation of a novel small-scale parallel bioreactor for continuous fermentation. Based on a technology developed at the university, he founded together with one of his colleagues (Frank Kensy) the company in 2005. Since then he is managing partner at m2p-labs GmbH.
Sharon Williams is the Product Development and Downstream Processing Manager for ProMetic BioSciences Ltd. This is a
biotechnology company that produces and supplies materials for the production and purification of biopharmaceuticals and for the
capture and removal of bio-contaminants. Her role is to manage the development of new products from conception to product launch
and subsequent use in the field. She also oversees the development downstream processes using these materials.
She has been working in downstream processing since she started her PhD in the Biochemical Recovery Group at University of
Birmingham, where she investigated adsorbent design for nanoparticle recovery. She continued this work as a Post Doctoral Research
Associate at the University of Cambridge where she investigated the affinity purification of viruses
Posters
STEADY STATE AND DYNAMIC CONTROL PERFORMANCE OF THE AMBR™ AUTOMATED MICRO BIOREACTOR SYSTEM IN A CHO CELL BATCH CULTURE.
Sinn Yee Yau, PhD, Kenneth Lee, PhD, Barney Zoro, EngD
TAP Biosystems, Cambridge, UK
Implementation of high throughput automated systems is recognised as a valuable approach in the field of bioprocess development, with broad acceptance that an efficient multi-parallel microscale bioreactor will become an important enabling technology. A standardised micro bioreactor system can unlock development bottlenecks in a variety of common bioprocess applications, such as cell line screening, media development, feed and process optimisation, QbD and DoE studies.
In this study we investigate the process control performance of the ambr™ micro bioreactor system, using 24 disposable micro bioreactors in parallel for a combination of steady state and dynamic control tests. We examine process control performance for three key process parameters (seeding cell density, pH, DO) and review cell count and glucose concentration profiles. Dynamic control tests challenge the capability of the ambr system to deliver common industrial process requirements such as pH and temperature shifts.
The results demonstrate the capability of the ambr system to support a range of typical steady state and dynamic control requirements for mammalian cell culture processes. The accurate and precise automated liquid handling and process control systems result in very low culture variation between replicate bioreactor conditions. Clear resolution in culture response (cell count, metabolism) is observed between different test conditions. The combination of tight process control performance, low replicate variation and high resolution between test results, illustrates the suitability of the ambr micro bioreactor system for large numbers of parallel bioreactor tests in a wide range of cell culture applications.
For questions about this poster, please contact Barney Zoro, TAP Biosystems Barney.zoro@tapbiosystems.com
AUTOMATED EVALUATION OF MICROSCALE LINKED PROCESS SEQUENCES FOR OPTIMISATION OF OXIDATIVE BIOCATALYTIC PROCESSES
J.Z. Baboo1, J.M. Ward2, G.J. Lye1, M. Micheletti1
Department of Biochemical Engineering1 & Institute of Structural and Molecular Biology2, University College London
Torrington Place, London WC1E 7JE UK
Corresponding author: j.baboo@ucl.ac.uk
Oxidative bioconversions offer valuable opportunities in industrial pharmaceutical synthesis such as using Baeyer-Villiger monooxygenases for antibiotic synthesis1. However, a limiting factor is the identification of scaleable hydroxylation biocatalysts. Coupling high-throughput microscale techniques with automation enables the operation of linked process sequences for faster identification and characterisation of optimal conditions2. A fully automated microscale sequence involving fermentation, bioconversion, liquid-liquid extraction for sample analysis and analytical techniques has been developed for the evaluation of whole cell Baeyer-Villiger monooxygenases. The automated approach has been shown to be robust and reproducible over multiple runs producing consistent results on different days. Rapid automated collection of quantitative kinetic data on a number of parameters has been applied to optimise biocatalyst expression and to identify new bioconversion substrates. Such studies have enabled the processing time to be halved and the biocatalysts specific activity to be more than doubled. Using the same methodology this automated platform is being applied to investigate novel cytochrome P450 biocatalysts for hydroxylation reactions3. Their vast reaction ability and essential role in both drug metabolism and bioremediation makes novel P450 biocatalysts an area of much interest. By using a matched oxygen transfer coefficient (kLa) approach both fermentation and bioconversion operations have been successfully scaled up to 75 L scale confirming the use of microscale approaches for process optimisation.
References:
- Strukul G (1998) Transition Metal Catalysis in the Baeyer-Villiger Oxidation of Ketones. Angewandte Chemie International Edition 37:1198-1209
- Lye GJ, Ayazi-Shamlou P, Baganz F, Dalby PA, Woodley JM (2003) Accelerated design of bioconversion processes using automated microscale processing techniques. Trends in Biotechnology 21:29-37
- Hussain HA & Ward JM (2002) Enhanced heterologous expression of two Streptomyces griseolus cytochrome P450s and Streptomyces coelicolor ferredoxin reductase as potentially efficient hydroxylation catalysts. Applied and Environmental Microbiology 69: 373-382
Microscale Characterisation of a Manufacturing Route for Lentiviral Vectors
H.M. Guy1,2, K.A. Mitrophanous1,*, G.J. Lye2, and T.K. Mukhopadhyay2
1The Advanced Centre of Biochemical Engineering, Department of Biochemical Engineering, University College London, Torrington Place, London, WC1E 7JE.
2Oxford BioMedica plc, Medawar Centre, Robert Robinson Avenue, The Oxford Science Park, Oxford, OX4 4GA.
The production and purification of high quality lentiviral vectors (LVs) at commercial scale presents many novel bioprocessing challenges. Microwell technologies may help to address some of these challenges by generating critical bioprocess information early in the development timeline
To date, material for LV clinical trails has typically been generated by transient transfection of adherent HEK293(T) cells grown in multilayer cell factories. Defining an upstream process based on suspension-adapted producer cell lines, however, is highly desirable for the commercial-scale production of LVs. ProSavin®, a LV engineered for the treatment of Parkinson’s disease, is presently in Phase I/II clinical trails. A ProSavin® producer cell line (PS46.2), which yields comparable titres to the current GMP HEK293T transfection method, was previously developed. However, recent adaptation of this cell line to suspension growth led to a significant drop in titre (approximately 10-fold).
In the present study, microscale fermentations were utilised in conjunction with statistical experimental design (DoE) to identify and optimise the key factors influencing ProSavin® production from suspension-adapted PS46.2 cells. A standard 24-well plate shaken system with Duetz sandwich lid was utilised for batch culture of the PS46.2 cell line. Samples were taken from sacrificial wells for offline monitoring of cell density, pH, osmolality, metabolites, pO2 and CO2. In addition, vector-containing supernatants were harvested from sacrificial wells for the analysis of total particle production (RNA copies/mL) and vector titre (transducing units, TU/mL).
Four rounds of DoE were employed to first screen (nine factor fractional factorial design) and then optimise (central composite designs) the critical factors influencing ProSavin® titre. Post-induction period, liquid fill volume, and concentration of inducer (doxycycline) were identified as the most critical factors. Optimisation of these factors led to an overall three-fold improvement over reference conditions (final titre approximately 6×104 TU/mL), and insight into the relationship between functional titre, cell growth and total particle production was obtained.
This work details the first application of a microscale and DoE approach to LV upstream development. The study demonstrates the value microwell technologies can bring to new and expanding fields, and highlights some of the benefits and drawbacks of the basic 24-well approach for LV production.
*Corresponding author: email: k.mitrophanous@oxfordbiomedica.co.uk
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