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TUESDAY, JANUARY 10
8:15 Chairperson’s Remarks
Rolf Douwenga, Ph.D., Vice President, Research & Development, DSM Biologics
8:20 Protein Purification in Vaccine Development - Challenges and Opportunities
Yan-ping Yang, Ph.D., Director, Downstream Purification, Bioprocess Research and Development, sanofi pasteur - Biography
Purification scientists face many technical challenges and time constraints in vaccine process development, from bench-top experimentation, scale up, process understanding and control, to clinical lot manufacturing and process industrialization. An efficient process development time with an optimized downstream process is the key in achieving overall success in vaccine product development. The combination of modern technology and innovation with conventional wisdom to achieve process efficiency and quality is the formula for overall downstream process excellence. Case studies will be presented to illustrate (1) the benefit of state-of-the-art technologies in accelerating vaccine protein purification process development; and (2) the use of purification and knowledge management tool box to achieve protein process development efficiency.
8:50 One-Second Isolation/Immunoprecipitation of Weak Protein Complexes Using a Microfluidic IFAST Device
Richard R. Burgess, Ph.D., Professor, Oncology, McArdle Lab for Cancer Research, School of Medicine, University of Wisconsin, Madison - Biography
We describe the use of a technique, Immiscible phase Filtration Assisted by Surface Tension (IFAST), recently pioneered in the lab of Dave Beebe to simplify immunoprecipitation. Its microfluidic nature makes IFAST easy to operate (surface tension-dominant fluid mechanics), it is amenable to arrayed processing (planar microfabrication), and exploits the many inherent advantages of microfluidics, including reduced reagent consumption, increased automation, lower device cost, and enhanced throughput, but does not require multiple electronic, pneumatic, or hydraulic connections to manipulate fluids in complex pathways.
9:20 Analysis of Protein Samples by Microfluidics: The Pros and Cons
Sabine Suppmann, Ph.D., Head, Recombinant Protein Production, Microchemistry Core Facility, Max-Planck Institute of Biochemistry MPIB - Biography
The trend from conventional SDS PAGE to LabChip systems has mainly been driven by the need for high-throughput analysis of protein samples. Beyond that, the speed, data quality and data management render microfluidics technology just as attractive for non-HTP protein research. As a Core Facility we provide an academic service for (non-automated) recombinant protein production. We replaced SDS PAGE by Agilent’s 2100 BioAnalyzer to monitor expression levels and protein fate during protein purification. In this talk I will present both pros and cons of this technology.
9:50 POSTER HIGHLIGHT
Purification of Antibody Subclasses Produced in Response to HSV-2 gC Immunization in Mice, Poster A206
Rachel Xoconostle, Biochemist, Vaccine Basic Research, Merck & Co Inc.
10:05 Networking Coffee Break in the Exhibit Hall with Poster Viewing
10:45 Differential Filtration Analysis: An Enabling Technology for Membrane Proteins (and Soluble Proteins too!)
Michael C. Wiener, Ph.D., Associate Professor, Molecular Physiology and Biological Physics, Center for Membrane Biology, University of Virginia - Biography
Structural studies on integral membrane proteins are routinely performed on protein–detergent complexes (PDCs) consisting of purified protein solubilized in a particular detergent. Identification of detergents that maintain the solubility and stability of a membrane protein is a critical step and can be a lengthy and ‘‘protein-expensive” process. We have developed an assay that characterizes the stability and size of membrane proteins exchanged into a panel of 94 commercially available and chemically diverse detergents. This differential filtration assay (DFA), using a set of filtered microplates, requires sub-milligram quantities of purified protein and small quantities of detergents and other reagents and is performed in its entirety in several hours. Additionally, we have characterized the effects of detergents upon the activity of proteases utilized to remove affinity tags from recombinant fusion proteins, and find detergent sensitivity to vary significantly among different proteases.
11:15 Novel Purification of Soybean Peroxidase (Glycine max) and SBP Applications in Detecting Bisphenol A
William Ward, Ph.D., Associate Professor, Biochemistry, Center for Research & Education in Bioluminescence & Biotechnology (CREBB), Department of Biochemistry & Microbiology, Cook College, Rutgers University
Soybean Peroxidase was extracted from 1.5 kg of soybean hulls. Following initial settling of large particles, the still-cloudy suspension was passed through Miracloth to remove remaining large particles. However, even careful filtration leaves a very cloudy suspension that invariably clogs flow columns. However, following batch-mode stirring with large, dense ResinTech SBACR beads, an easily aspirated fine gray sediment forms over the rapidly settling beads that have trapped virtually all of the SBP. The ResinTech step simplifies all subsequent purification steps. We have adapted this later reaction as a spot test for bisphenol A (BPA) in thermally sensitive store receipt paper.
11:45 Protease-Based Nanomachines
Philip N. Bryan, Professor, Institute for Bioscience and Biotechnology Research, and Department of Bioengineering, University of Maryland - Biography
We will describe the engineering of the serine protease subtilisin and its prodomain inhibitor to create sophisticated enzymatic machines. Three basic aspects of protease function were modified: activation, specificity, and inhibition. In these enzymatic machines, the protease occupies a role analogous to a transistor in an electronic circuit and is able to mediate such functions as detection, purification, activation, or inactivation (destruction) of other proteins. We have developed a protein purification system that uses a simple protease machine for protein purification and provides the conceptual foundation for other devices based on tightly-regulated proteases.
12:15 pm Close of Morning Session
12:30 Luncheon Presentation I
Rhobust® Technology – Process Intensification Through Direct Product Capture
Rolf Douwenga, Ph.D., VP, Research and Development, DSM Biologics
DSM Biologics’ Rhobust® technology is based on the second generation Expanded Bed Adsorption and represents an excellent solution for direct protein capture from crude harvests and viscous feed streams. The direct protein capture circumvents the need for separate unit operations for cell removal such as dead-end filtration or centrifugation, thereby reducing the processing times and cost of goods. The Rhobust® technology utilizes a new generation of Agarose beads that are prepared from a homogeneous mixture of Agarose and Tungsten carbide (10 vol%) and consequently have a high density (3 g/mL) that allows processing in the expanded bed mode and at higher flow rates than with standard packed bed chromatography systems. The Rhobust® technology combines particularly well with high cell density and high titer harvests, such as those achieved using DSM Biologics’ proprietary XD® technology. Here, we present the Rhobust® technology as the ideal solution for Downstream Processing of XD® harvests, with more than 100 million cell/mL, where product was captured in one single step while still retaining the same product quality when compared to traditional methodologies.
1:00 Luncheon Presentation II
Direct Detect IR-Based Platform Revolutionizes Biomolecule Quantitation
Ivona Strug, Senior Biochemical Scientist, EMD Millipore
By directly measuring amide bonds in protein chains, Direct Detect accurately determines an intrinsic component of every protein without relying on amino acid composition. Within minutes and without any bio- or immuno-chemical staining, protein concentration in an extremely wide range from about 0.2 mg/ml to >20 mg/ml can be determined directly from the undiluted solution. In addition, quantitation by Direct Detect is not subject to interference from many common buffer components such as detergents, reducing agents and chelators.
2:00 BuzZ Session A
2:45 Networking Refreshment Break in the Exhibit Hall with Poster Awards
3:30 BuzZ Session B
4:15 End of Conference
4:30 – 7:30 Concurrent Dinner Short Courses (SC5-SC9)
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Links to Companion Meetings
Higher Throughput Protein Purification
Protein Aggregation and Emerging Analytical Tools