Device to monitorcontrol differentiation of stem cells to β.pptVIP

Device to Monitor/Control Differentiation of Stem Cells to β-islet Cells Dhaval Desai – Team Leader Jon Baran – BWIG Tim Pearce – BSAC Tess Rollmann – Communications Client : Victoria Browning, Ph.D. Advisor: Naomi Chesler, Ph.D. Motivation Type I diabetes patients cannot produce insulin Current treatment methods take insulin from a donor Stem cells show promise to differentiate into insulin secreting cells ?eliminate the need for a donor Problem Statement Differentiate foregut committed progenitor cells into insulin-producing pancreatic β-islet cells Replace or supplement transplanted donor beta cells Test different concentrations of growth factors (GFs) for their ability to affect conversion of progenitor cells into mature insulin-secreting cells Continuous linear growth factor gradient Design Contraints Capable of holding ≥ 100 cells (1000-5000 ideal) Compatible with imaging Capable of withstanding immunofluorescense Able to withstand 7-28 day incubation period at 37 degrees Celsius Minimal amount of GF required Total cost of under $500 Previous Work Created linear gradient in a microfluidic channel Channel filled with matrigel to create a high resistance barrier Characterized the gradient formation using modeling software Design Change Problems in 3D Results cannot be compared to Mashima et al paper Expensive imaging for 3D set up Unconventional cell culture protocol Redesign in 2D Goals for Redesign Create a GF gradient within a microfluidic channel which allows for 2D cell culture Integrate and test viability of cells in the channel Validate our microenvironment culture conditions by comparing with standard culture conditions as seen in the Mashima et al paper Method 1 High resistance membrane 0.8 micron pore polyester membranes Placed between 2 PDMS layers Cells suspended in media introduced from the sink Membrane Results Successes Gradient formation Plasma bonding sealed PDMS layers to prevent leakage Using non-fluorescent dye to ensure fluid conne