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THURSDAY, AUGUST 23
8:00 am Morning Coffee
ENGINEERING DESIRED QUALITIES
8:55 Chairperson’s Remarks
Joachim Wegener, Ph.D., Institute of Analytical Chemistry, Chemo and Biosensors, University of Regensburg
9:00 Gene Editing Approaches for Viable Commercial Production
Eric Rhodes, Chief Technical Officer, Horizon Discovery Ltd. - Biography
There are now several technologies available to engineer stable genetic changes into the genome of mammalian cell lines. Each has its own strengths and weaknesses and the choice of which approach is best suited is dependent on a number of factors. Among the factors to be considered are: the nature and complexity of the desired modifications; the expertise required to achieve the end result; any safety and regulatory concerns; freedom to operate issues; and downstream commercial costs. An overview of the available technologies in light of these factors will be discussed.
9:30 Optimizing Recombinant Glycoprotein Production in the Baculovirus-Insect Cell System
Donald L. Jarvis, Ph.D., Professor, Molecular Biology, University of Wyoming - Biography
The baculovirus-insect cell system is widely used to produce recombinant proteins with eukaryotic modifications, such as glycosylation. However, this platform is constrained by the fact that the insect cell lines used as hosts for baculovirus expression vectors have relatively primitive protein glycosylation pathways that cannot produce sialylated glycoproteins. We have addressed this problem by glycoengineering the baculovirus-insect cell system to humanize the protein N-glycosylation pathway. In this presentation, I will report the results of our latest glycoengineering efforts and their impact on our efforts to optimize the baculovirus insect cell platform for recombinant glycoprotein production.
10:00 Coffee Break
TRANSFECTION & ELECTROPORATION
10:30 Lighting the Way for Biologists: Optical Transfection
Frank Gunn-Moore, Ph.D., Professor of Molecular Neurobiology, School of Biology, University of St. Andrews - Biography
The plasma membrane of a eukaryotic cell is impermeable to most hydrophilic substances, yet the insertion of these materials into cells is an extremely important and universal requirement for the cell biologist. To address this need, many transfection techniques have been developed including viral, lipoplex, polyplex, capillary microinjection, gene gun and electroporation. The current discussion explores a procedure called optical injection or photoporation, where a laser field transiently increases the membrane permeability to allow species to be internalized. If the internalized substance is a nucleic acid, such as DNA, RNA or small interfering RNA (siRNA), then the process is called optical transfection. This contactless, aseptic, cell transfection method provides a key nanosurgical tool to the microscopist—the intracellular delivery of reagents and single nanoscopic objects. The experimental possibilities enabled by this technology are only beginning to be realized. This presentation will discuss our new and novel optical transfection technology which is being developed with the end-user in mind.
11:00 The Road Toward Scalable Transient Gene Expression in Mammalian Cells
Lucia Baldi Unser, Ph.D., Senior Scientist, Laboratory of Cellular Biotechnology, EPFL - Biography
Transient gene expression (TGE) was established as a faster alternative to stable cell lines for the production of r-proteins. With TGE, no cloning after transfection is necessary. Although mainly considered as a research tool, and not yet a standard production platform for industrial manufacturing, TGE is raising a growing interest in the industrial biotechnology R&D departments. In this talk, I will describe how the limitations to successful large-scale TGE in HEK293 and CHO cells were surpassed during the last 15 years, and how we can use today’s state-of-the-art technology for the production of gram-amounts of r-protein.
11:30 Impedance Analysis of Adherent Cells after in situ Electroporation: Non-Invasive Monitoring during Intracellular Manipulations
Joachim Wegener, Ph.D., Institute of Analytical Chemistry, Chemo and Biosensors, University of Regensburg - Biography
In this study adherent animal cells were grown to confluence on circular gold-film electrodes of 250 µm diameter that had been deposited on the surface of a regular culture dish. The impedance of the cell-covered electrode was measured at designated frequencies to monitor the behavior of the cells with time. The gold-film electrodes were also used to deliver well-defined AC voltage pulses to the adherent cells in order to achieve reversible membrane electroporation (in situ electroporation = ISE). Cells recovered from the electroporation pulse within less than 90 min. When membrane-impermeable, bioactive compounds like N3− or bleomycin were introduced into the cells by in situ electroporation, concomitant impedance readings sensitively reported on the associated response of the cells to these toxins as a function of time.
12:00 pm End of Conference
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