59-05-032 Proceeding

44 Proceedings of the Princess Maha Chakri Sirindhorn Congress SINGLE CELL CHARACTERIZATION OF NEUTROPHIL ROLLING IN MICROFLUIDICS Chayakorn Petchakup 1 , Han Wei Hou 2 and King Ho Holden Li 3 Abstract : Neutrophils are themost abundant peripheral white blood cells that play a crucial role in innate immunity.The study of neutrophil functional phenotype usingmicrofluidics enables single cell analysis andwouldbe imperative in identifying surrogate cell-basedbiomarker for inflammation in chronic diseases such as cardiovascular diseases and diabetes. Here we developed amicrofluidics strategy to characterize neutrophil rolling on E-selectin, a cell adhesion molecule expressed on inflamed vessel walls during leukocyte recruitment. In our experiments, healthy neutrophils were purified fromwhole blood and treatedwith various inflammatory stimulus such as tumor necrosis factor alpha (TNF- α ) and phorbol myristate acetate (PMA). Healthy and treated neutrophils were then pumped into the E-selectin coated microchannel under physiological shear conditions (~2 dyne/cm 2 ) and neutrophils velocity distribution and rolling trajectory were analyzed using a cell tracking algorithmOur preliminary results showed distinct differences in cell rolling velocity and trajectory between healthy and inflamed neutrophils, clearly demonstrating the potential of using cell rolling as an intrinsic functional biomarker for clinical testing. Keywords : Image processing, Inflammatory stimulus, Microfluidics, Neutrophil rolling I. Introduction Neutrophils are the most abundant white blood cells in the bloodstream which play an essential part in immune systems. During leukocyte recruitment to inflamed blood vessels, neutrophils will first bind to endothelial cells to initiate rolling followed by firm adhesion and extravasation to the site of injures [1]. Previous studies have reported abnormal functional changes in neutrophils during inflammation in chronic diseases such as cardiovascular diseases and diabetes [2-4]. The motivation of this study is to characterize the rolling phenotype of neutrophils using microfluidics which can mimic physiological environment in microvasculature. Neutrophils purified from human blood were allowed to flow through a microchannel functionalized with E-selectin, one of the key rolling ligands expressed on inflamed endothelium under physiological conditions. Time lapse imaging was used to capture cell rolling behavior and image processing was implemented to track cell motion. This offers high throughout single cell analysis to detect subtle changes in cell rolling phenotypes. Here we demonstrated the feasibility to characterize distinct neutrophil rolling on microchannel by utilizing both microfluidic and image processing. 1 Mechanical and Aerospace Engineering, Nanyang Technological University, SINGAPORE 2 Lee Kong Chian School of Medicine, Nanyang Technological University, SINGAPORE 3 Mechanical and Aerospace Engineering, Nanyang Technological University, SINGAPORE