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Genigraphics® has been producing output from PowerPoint® longer than anyone in the industry; dating back to when we helped Microsoft® design the PowerPoint® software. US and Canada: 1-800-790-4001 Email: info@genigraphics.com [This sidebar area does not print.] Effect of Yeast Cell Shearing in Relation to Various Patterned Microchips Turner Adornetto 1 ; Usoshi Chatterjee 1 ; Luke Forshey 1 ; Ryan Schoell 1 1 The Ohio State University’s Fundamentals of Engineering Honors Program Team X6 would like to thank Mr. Paul Clingan, Andrew Theiss, Aaron Strickland, Martin Spang, Erica Brackman, Ani Tarimala, and the rest of the FEH instructional staff. Team X6 would also like to thank the FEH program at The Ohio State University, and its sponsors for providing this research opportunity. Acknowledgements 1.M. Mercier-Bonin et al. Study of bioadhesion on a flat plate with a yeast/glass model system. Journal of Colloid and Interface Science. 2004, 271, 342-350 2. Statistics Package. www.carmen.osu.edu. April 2014. 3. Lab 03-1 Chip Molding Procedure. February 2014. 4. Lab 03-2 Yeast Adhesion and Shearing Procedure. February 2014. References The objective of this project was to determine the effect of surface topography on yeast cell adhesion. Yeast cells were allowed to attach to hexagonal-well patterned and unpatterned micro channel and then were sheared off by allowing water to flow through these micro channels. Objectives As the height of the water column increased, the channel experienced a higher shear stress due to an increase in pressure Average shear stress for unpatterned surface: 2.6 x 10 2 dynes/cm 2 Average shear stress for patterned surface: 2.45 x 10 2 dynes/cm 2 Number of yeast cells counted for the unpatterned chip (from initial flush to 3 rd height) ranged from 14,580 to 102, and the same range for the patterned chip was 22,272 to 180 More yeast cells were removed from the channel at higher shear stress Change in cell adhesion between low pressure flush and water flow from height of 45 cm for both patterned and unpatterned surfaces are shown in Figure 3 Figure 4 shows the difference between the shear stresses on the unpatterned and patterned surfaces for the collected data points Introduction Microfluidics system was assembled as shown in Figure 2 below Yeast-water suspension was injected into the channel and allowed to adhere for 20 minutes Water was introduced into the channel from a height of 35 cm for 5 minutes for a low pressure flush to remove excess yeast cells Channel was examined under a microscope and a picture was taken to determine the number of yeast cells on the channel surface The channel was then subjected to increasing shear stresses by increasing the height of water flow in increments of 5 cm Each time the water was allowed to flow for 2 minutes and the images of the channel was recorded by focusing on the same location as that of the first picture Number of cells present in the channel were counted from the pictures Methods A statistical analysis was performed to decide whether the results were statistically significant. With an α value of 0.05, a type 2, 2-tailed t-test was performed on the data, and the p-value from the t-test was 0.977. Statistical Analysis Based on shear stress calculations and using a 2 tailed t-test, no statistically significant difference was observed between the shear stress needed to dislodge cells from the patterned hexagonal welled surface and the blank chip, as the p-value (0.977) was greater than α/2 (0.025). Null hypothesis was not rejected, and surface topography does not significantly affect cell adhesion. Since the average diameter of the yeast cells (Saccharomyces cerevisiae) used in the experiment was 5 to 10 µm, it is possible that the 8 µm wells on the patterned surface could not accommodate the larger yeast cells; thus, reducing the ability of the cells to adhere. Conclusions are applicable to the particular species of yeast cells tested and patterned surface used for the experiment. For a more generalized conclusion, the study needs to be expanded to include a larger, more diverse range of patterned surfaces, variety of yeast cells, and significantly more data points. Conclusions Results and Discussion Figure 1. Exploded View of Standard Chip Figure 3. Comparison of unpatterned (above) and patterned (below) micro channels with yeast cells after initial low pressure flush, and 1 st tested height. Magnification of 100x. Figure 4: Comparison of unpatterned and patterned micro channel shear stress based on pressure in dynes/cm 2. It appears that higher stress was required to shear cells from the unpatterned surface compared to the patterned surface. Medical device design, in particular implantable devices, is dependent on the degree of cellular interactions with surface topography. A high degree of cell adhesion is expected during the process of regeneration of human tissue, based on tissue engineering, as this depends on the ability of the cells to attach and grow on a material. However for cancerous cells, cell adhesion should be minimum to discourage their growth. The goal of this study is to understand how cells attach to different surfaces. Complex flow environments, as found in human body (blood vessels), can be mimicked using micro fluidic devices, and our experiment focuses on understanding cell adhesion on micro channels. Mercier-Bonin et al. tested shear induced detachment of various types of yeast and its effects. This project utilized micro fluidic flow concepts and determined flow rate and shear stress values based on the yeast cells adhered to the channel wall. Hypothesis Null Hypothesis: Surface topography will have no effect on yeast cell adhesion Alternative Hypothesis: Surface topography will have a statistically significant effect on yeast cell adhesion Alpha Level: 0.05 A chip with micro channels, a smooth chip top, and a patterned chip were fabricated using polydimethylsiloxane (PDMS) The chip holder top and bottom were made of acrylic, and finished with a laser cutter Yeast-water suspension was prepared by mixing dry yeast with water and incubating for 20 minutes Material Fabrication Significance Level α:0.05 α/2:0.025 Degrees of Freedom:7 P-value0.977 Figure 2. Experimental Set-Up Figure 4: Statistical Analysis.