2016 Archived Content

Cambridge Healthtech Institute’s 2nd Annual
Protein Purification Technologies

Streamlining Processes to Achieve Quality

3 - 4 November, 2016 | EPIC SANA Lisboa Hotel | Lisboa PORTUGAL

As demand for biological drugs increases, so does the need for purified protein. Whether for research or to create biotherapeutics, researchers must purify protein following expression and scale-up. The plethora of novel and complex protein-based molecules pose greater challenges for purification professionals, including bispecifics and antibody-drug conjugates. Protein’s tendency to aggregate is also a nagging problem, as is working with membrane proteins. The “Protein Purification Technologies” meeting explores the strategies and technologies employed to streamline steps and keep up with industry’s growing demands.

This conference also examines issues of scale, and innovating ‘traditional’ technologies such as Protein A and chromatography. Purifying proteins in different expression systems will also be addressed including mammalian, bacterial, and insect cells.

Final Agenda

Day 1 | Day 2 | Speaker Biographies | Download Brochure


12:30 Registration

13:00 Dessert Break in the Exhibit Hall with Poster Viewing


13:30 Chairperson’s Opening Remarks

Ana Cecília Afonso Roque, Ph.D., Assistant Professor, Chemistry, UCIBIO, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa


Biopharmaceuticals: Past, Present & Future

Frank_RiskeFrank Riske, Ph.D., Senior Consultant, BioProcess Technology Consultants (BPTC)

This presentation will cover the evolution of bioprocessing starting with the production of molecules such as insulin, HGH and alpha-interferon and bringing us up to today; a market dominated by monoclonal antibodies but where gene and cell therapy have become increasingly commonplace, biogenerics have made their mark and a number of new therapies are in the works. How has bioprocessing changed to accommodate this diversity? The field has changed from producing and purifying small proteins expressed in bacteria to large, post translationally, modified molecules produced in multiple types of cells (but dominated by CHO). Although Mab’s dominate the todays’ market there is a wide diversity of molecules in the pipeline ranging in size from several thousand to several million Dalton. Instead of just Mab’s we now have their offspring; nanobodies, minibodies, diabodies, bispecifics and other forms. Furthermore, synthetic chemistry continues to push the size limit of what can be produced by solid phase synthesis (several thousand Daltons) and then refolded or modified. 
Bioprocess scientists need the flexibility to produce this wide variety of molecules. They need flexible facilities and processes and have turned to disposables, new technologies (e.g. single use filters, charged and hydrophobic membranes, pre-packed columns, single pass TFF, affinity capture) and new approaches (continuous systems) to bridge this gap. This presentation will explore these approaches, the current state of the industry, and a potential future.

14:20 Molecular Properties of Antibodies and Their Derivatives Impacting Manufacturing

Stefan_SchmidtStefan Schmidt, Ph.D., MBA, Vice President, Process Science & Production, Rentschler Biotechnology

Certain molecular properties and process conditions can lead to aggregation, truncation and chemical modifications complicating downstream processing. Particularly highly engineered molecules differing from the original antibody format require a careful evaluation of parameters like contact time, ligand density, elution pH, capacity, etc. to identify optimal settings for the initial capture step. Several examples and case studies illustrate the general approaches undertaken to deliver high-quality antibodies and derivatives taking into account their molecular properties.

14:50 Quality Control of Recombinant Proteins: Best Practice Recommendations

Mario_LebendikerMario Lebendiker, Ph.D., Head, Protein Purification Facilities, Wolfson Centre for Applied Structural Biology, The Hebrew University of Jerusalem
Around 200 specialists in protein production and in biophysical characterization of biomolecules, of the most important Core facilities in Europe and Israel, have formed a joint initiative to establish guidelines on recombinant protein quality.
The Production and Purification Partnership in Europe (P4EU) and the Association of Resources for Biophysical Research in Europe (ARBRE-MOBIEU) aim to develop a best practice/minimal standard for the quality control of recombinant proteins to ensure that the input material used in biophysical and biochemical research is of high quality, which, in turn, will result in optimized data quality. The prescribed tests must be both feasible by all protein production labs, and, at the same time, acceptable for biophysical or structural biology labs as admission criteria. A series of ‘best-practice’ methods is suggested for further characterization.  To include an extended discussion with conference delegates.

15:20 Refreshment Break in the Exhibit Hall with Poster Viewing


16:05 Development of an Automated Parallel System for Production of High-Quality Lead Candidate Antibodies – Keeping Pace with Researchers’ Appetites

Lars_LindenLars Linden, Ph.D., Head, Protein Biochemistry, Bayer HealthCare

In early research, project teams profile multiple antibody candidates in functional and often cell-based assays. Within a growing portfolio, this leads to an increasing resource need for the production of mg amounts of high-quality antibodies. To meet this demand, transient transfection and expression were automated, a novel high-throughput 2-step chromatography system was set up, and an integrated automation concept for sample handling was developed.

16:35 Enhancing Operational Efficiency during Protein Purification

Martin_BaderMartin Bader, Ph.D., Head, Biochemistry, Roche Pharmaceuticals

Purification of bispecific antibodies is often more labor-intensive due to complex impurity profiles. We have systematically analyzed and optimized individual process steps during pre-clinical protein supply. We have developed work flows that reduce operator time and combine critical pooling decisions with high-throughput analytics. In addition, generic steps are either automated or outsourced. In summary, we present a highly efficient purification platform for next-generation proteins.

17:05 End of Day

17:00 Dinner Short Course Registration

Recommended Dinner Short Course*

SC7: Protein Purification Strategies: Dealing with Proteins that are Prone to Aggregate

*Separate registration required

Day 1 | Day 2 | Speaker Biographies | Download Brochure


08:00 Registration and Morning Coffee


08:30 Chairperson’s Remarks

Stefan Schmidt, Ph.D., MBA, Vice President, Process Science & Production, Rentschler Biotechnology

08:35 Continuous Aqueous Two-Phase Extraction of Proteins – Novel Process Windows by Increasing Protein Solubility

Christoph_BrandenbuschChristoph Brandenbusch, Ph.D., Group Leader, Department of Biochemical and Chemical Engineering, Laboratory of Thermodynamics, TU Dortmund University

Increasing product titers in pharmaceutical protein production led to a demand for novel workup strategies (e.g. protein extraction by Aqueous Two-Phase Systems). However, the effort and costs required in selecting appropriate ATPS and process windows hinder the industrial applicability. Hybrid modeling approaches enable the estimation of efficient phase formers and novel displacement agents. This allows for both a selective purification and stabilization of the protein in solution by increasing solubility.

09:05 Resolving Perplexing Purification Paradigms with CD-FAST: Calcium Dependent Fragment Assembly Separation Technology

David_OConnellDavid O’Connell, Ph.D., Lecturer & Director, Biomolecular & Biomedical Science, University College Dublin, and Founder, YourProteome

Membrane proteins and chaperone-assisted cytosolic proteins prove to be particularly perplexing when it comes to isolating them in sufficient quantity and purity for downstream assays. Most rationales involve multi-step strategies that consume valuable amounts of protein. The application of CD-FAST technology is a powerful innovation in the single step isolation of pure, functional proteins with recent advances in GPCR and aggregation-prone kinase purification and downstream assay reported here.

09:35 A Saposin-Based Nanoparticle System for Stabilization of Membrane Proteins

Jens_FrauenfeldJens Frauenfeld, Ph.D., CEO, Salipro Biotech AB

We present a saposin-lipoprotein nanoparticle system, which allows for the reconstitution of membrane proteins in a lipid environment. We demonstrate the applicability of the method on purified membrane proteins as well as by the direct solubilization and nanoparticle incorporation of membrane protein complexes from the virus membrane. The Salipro system allows for high-resolution cryo-EM of membrane proteins and is applicable for the development of novel drugs, vaccines and therapeutic antibodies.

10:05 Coffee Break in the Foyer with Poster Viewing


10:35 Versatile Affinity Ligands for Bioseparation Processes

Ana_RoqueAna Cecília Afonso Roque, Ph.D., Assistant Professor, Chemistry, UCIBIO, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa

We employed biological and chemical combinatorial libraries supported by computational design tools to develop robust peptidomimetics based on different scaffold molecules. The scaffold molecules ranged from small synthetic ligands, to artificial β-hairpin peptides and small protein domains produced chemically. We studied the potential of these scaffold affinity reagents to find binding partners against several targets (e.g. recombinant proteins, phosphorylated peptides, and virus-like particles), and to develop affinity-based purification processes.

11:05 Affitins as a Novel Tailored Class of Robust Reagents for Affinity Chromatography Purification of Antibodies and Non-Immunoglobulin Proteins

Frederic_PecorariFrédéric Pecorari, Ph.D., Researcher, IRS-UN, Cancer Research Center Nantes-Angers, University of Nantes

This presentation reveals how Affitins can be used as ligands to design affinity columns for one-step purification of human immunoglobulin G (hIgG), with 95% purity and recovery of 100%. Affitin production in E. coli makes it possible to produce these affinity columns at low cost. Our results validate Affitins as a new class of tailored ligands for the affinity purification of potentially any proteins of interest including biopharmaceuticals.

11:35 A Novel IgG-Binding Purification Matrix for Mild Elution

Sophia_HoberSophia Hober, Ph.D., Professor, Biotechnology, KTH-Royal Institute of Technology

Antibodies are widely used affinity molecules in many fields of biological science and the therapeutic field for monoclonal antibodies is constantly growing. The most common method for purification of antibodies is protein A affinity chromatography. It offers high-selectivity, and yields pure and concentrated antibodies. One of the major issues with protein A purification is the low pH that is essential to elute the bound antibody from the column. Here, we have addressed this by protein engineering of the IgG-binding domain from protein A.

Purification of an Aggregation Prone Protein - Challenges and Solutions for Polymeric IgM Production
Julia Hennicke, Graduate Student, Bio Tech, University of Natural Resources and Life Sciences (BOKU)

12:35 Problem-Solving Breakout Discussions with a Light Snack in the Foyer

One Affinity Tag to Rule Them All: Is Any Single Tag Good Enough for Complex Protein Purification Rationales?

David_OConnell David O’Connell, Ph.D., Lecturer & Director, Biomolecular & Biomedical Science, University College Dublin

  • When is IMAC/FPLC just not good enough?
  • Single step affinity purification - fact or fiction?
  • Membrane protein purification with affinity tags
  • Affinity purification of disordered proteins & chaperones
  • Can Protein A chromatography be replaced with Affinity Tag Purification?

Protein Quality Standards that Should be Implemented by Journal Reviewers to Improve the Quality of Publications

Sabine_Suppmann Sabine Suppmann, Ph.D,, Head, Recombinant Protein Production, Biochemistry Core Facility, Max Planck Institute of Biochemistry

There is an urgent need for a protein quality standard in the academic environment to improve reliability of publications. This roundtable discussion is meant to be a brain-storming focused on the following questions:

  • How would we define such a standard?
  • How would we spread the standard?
  • How would we convince the scientists to perform protein quality assessment?
  • How would we convince funding agencies and editors of journals to insist on the documentation of protein quality assessment?

13:35 Session Break


14:00 Chairperson’s Remarks

Christoph Brandenbusch, Ph.D., Group Leader, Department of Biochemical and Chemical Engineering, Laboratory of Thermodynamics, TU Dortmund University

14:05 Peptides in Headlock: A Novel Nanobody-Derived Capture/Detection System

Ulrich_RothbauerUlrich Rothbauer, Ph.D., Professor, Pharmaceutical Biotechnology, University of Tübingen

Single-domain antibodies (nanobodies) have emerged as an attractive alternative to traditional antibodies and have become highly valuable tools for numerous bioanalytical and biotechnical applications. Here we present a novel nanobody-derived capture/detection system that enables a fast and efficient isolation of epitope-tagged proteins from prokaryotic and eukaryotic expression systems. The high-affinity-binding and modifiable peptide tag of this system renders it a versatile and robust tool to combine biochemical analysis with microscopic studies.

14:35 Case Study: Human Kinase Crystallization, Phosphatase Co-Expression

Mario_LebendikerMario Lebendiker, Ph.D., Head, Protein Purification Facilities, Wolfson Centre for Applied Structural Biology, The Hebrew University of Jerusalem

We present a case study of production of a human Kinase co-expressed in E.coli cells together with phosphatase in order to get a homogeneous, non-phosphorilated, and crystallizable protein. The reproducible production of different batches in E.coli allow us to successfully crystallize our target with a great variety of ligands with possible therapeutic implications. We describe our expression and purification approach, bottlenecks, etc. Since this protein was crystallized in the past, this case study emphasizes the need to establish “minimal protein quality information” in publications in order to assure reproducibility of results.

15:05 Use of Succinylated Trypsin as a Manufacturing Aid

Nina_MadsenNina Madsen, M.S., Scientist, Downstream Process Development, Bristol-Myers Squibb

We present a robust method for the charge modification of lysine residues on trypsin, by way of an acylation reaction between succinic anhydride and the lysine functional group. We then demonstrate the use of this modified enzyme as a tool in the downstream manufacture of a recombinant microbial product. Our protein is expressed in E. coli as a prohormone precursor and is enzymatically converted to the desired product using trypsin.

15:35 Development of a Robust Ultrafiltration/Diafiltration Step for Highly Concentrated Drug Substance Using High-Throughput Methods and Aggregation Modelling

Dejan_ArzensekDejan Arzensek, Ph.D., Research Scientist, Downstream Process Development, Sandoz Biopharmaceuticals

The present study focused on evaluating critical parameters within the Ultrafiltration/Diafiltration (UF/DF) operation that lead to aggregation due mainly to the physical instability of therapeutic mAbs. Insight into the physical instability of the protein in solution was achieved by using a directed approach based on a more detailed biophysical characterization required to ensure successful process development. Aggregation and particle formation were monitored using multiple analytical techniques for UF/DF conditions tested.

16:05 A New Shaped Rocking Bioreactor for Insect and Mammalian Cells

Sabine_SuppmannSabine Suppmann, Ph.D,, Head, Recombinant Protein Production, Biochemistry Core Facility, Max Planck Institute of Biochemistry

Wave-rocking bioreactors have increasingly replaced traditional stainless steel stirred tank reactors for large-scale cultivation of insect or mammalian cells during the last decade. Disposable wave bags are available in many shapes and sizes from several commercial providers. In research laboratories, however, production in wave bags is much less established for cost reasons and still mainly performed in shaking flasks. We will discuss how we have developed a re-usable, fixed-shaped bioreactor, which can be used on the diverse rocking unit platforms that control temperature, aeration rate, rocking speed, and rocking angle.

16:35 End of Conference

Day 1 | Day 2 | Speaker Biographies | Download Brochure