Join the free seminar conducted on Real-Time Cell Viability Apoptosis and on Human iPSC-Derived Cell Types. The seminar is free of charge and will take place at O&N 4 at Campus Gasthuisberg.
- Campus Gasthuisberg I Room 04.330, O&N4 I Herestraat 49, 3000 Leuven Room 04.330, Onderwijs en Navorsing 4, Campus Gasthuisberg.
- Date: Wednesday, September 27, 2017
- Time: 9:30 AM–11:30 AM (CEST: Amsterdam, Berlin, Stockholm)
Contact: Lindsay Mesure email@example.com
No pre-registration needed!
Join the complimentary onsite seminar at Campus Gasthuisberg on 27 September 2017
The program will be presented by both a scientist from Promega and from Cellular Dynamics International - a FUJIFILM company
Title: Utilizing a Novel Engineered Shrimp-Derived Luciferase to Enable Real-Time In Vitro Measurement of Cell Viability and Apoptosis using a Plate-Reading Luminometer
Speaker: Dr. Terry Riss (Global Strategic Manager Cell Health, Promega Corporation)
We have engineered two versions of a shrimp-derived luciferase to develop new in vitro assay technologies for toxicological research. The first version of this new technology uses an engineered luciferase and a pro-substrate as a reagent to monitor cell viability in real-time by measuring the conversion of the pro-substrate into a luciferase substrate which is limited to live cells. Viable cell number is monitored by recording luminescence repeatedly from the same sample of cells for days, providing kinetic information on the status of the culture. The real-time viability assay also provides a new tool to enable repeated dose toxicity experiments extending for weeks. Assay reagent can be delivered to cultures and viable cell number recorded just prior to the normal schedule of changing culture medium and replenishing the toxin of interest. The reagents are well tolerated by live cells which enables subsequent multiplexing of other assays such as extraction of RNA to analyze gene expression to identify specific stress response pathways. The second version uses small and large fragments of luciferase that have been engineered as fusion proteins linked to annexin V for detecting of apoptosis in real-time using a plate-reading luminometer. The individual annexin V-luciferase fragment fusion pairs have low intrinsic affinity for each other and thus produce no or low luminescence in culture medium or in the presence of non-apoptotic cells; but, when the annexin V-luciferase fragment fusion proteins bind in close proximity to phosphatidylserine exposed on the surface of apoptotic cells, the luciferase fragments can reconstitute an active enzyme and generate a luminescent signal. The annexin v-luciferase fragment fusion proteins and a luciferase substrate are combined to form a reagent that is added directly to cells in culture to create a homogeneous protocol that does not require cell washing steps typically used with fluorescent annexin V binding assays. Monitoring luminescence over time shows the onset of apoptosis precedes secondary necrosis measured from the same sample by multiplexing with a non-permeable fluorogenic DNA binding dye to indicate membrane integrity. These new luciferase assay technologies to monitor cell viability and apoptosis in real-time from the same samples provide kinetic information not available from endpoint assays and greatly simplify assay protocols compared to using flow cytometry or high content imaging.
Title: Human iPSC-Derived Cell Types Enable Investigation of Molecular Mechanisms and Prediction of Drug Liabilities in Physiologically Relevant In Vitro Models.
Speaker: Giorgia Salvagiotto, Ph.D Senior Field Application Scientist, Cellular Dynamics International - a FUJIFILM company
Human cell types differentiated from induced pluripotent stem cells (iPSCs) offer an attractive source of cellular material for both basic research and drug discovery because of the biologically relevant systems they can represent in vitro. The iPSC technology is a key element for modeling human biology in a dish, which is otherwise difficult to explore using conventional cell lines, primary cells, or animal models. Our approach is to generate iPSC-derived cell types with high quality, purity, and unlimited quantities, design relevant assays with cells derived from apparently healthy donors, and develop disease models using environmental stimuli or disease-specific, patient-derived cells.
The presentation will focus on two iPSC-derived cell types, neurons and cardiomyocytes, their characterization, and their functional utility to study disease mechanisms or detect drug liabilities. In particular, we will discuss examples of applications of iPSC-derived neurons to model neurodegenerative diseases and neuropathy, including chemotherapy-induced neuropathy (CIPN). These diseases are difficult to study due to the lack of human models. We will show that the iPSC-derived neurons display the phenotypic characteristics of the disease in vitro, and offer the relevant sensitivity to detect drug neurotoxicity at clinical relevant concentrations. Similarly, we will present case studies that show how iPSC-derived cardiomyocytes could potentially be included in early detection of cardiac liability across different therapeutic classes, including anti-cancer drugs such as kinase inhibitors and monoclonal antibodies.
These advances in iPSC technology provide access to previously unattainable human cell types opening new opportunities to address the limitations of rodent primary cells and immortalized cell lines.