gradients, and surface molecules etc. Our work has focused on development of microfluidic assays for cellular mechanics measurements in the context of human blood diseases. For instance, a fundamental mechanism of vascular occlusion in sickle cell disease has been recognized as a result of deoxygenated intracellular sickle hemoglobin. Sickle hemoglobin forms into fibers at low partial pressure of oxygen, causing rigid sickle cells that can severely impact blood flow in narrow vessels when they interact with other cell types in a complex manner [32]. Since many factors can be associated with deteriorated cellular mechanical properties, such as molecular level-variations in hemoglobin types and concentrations as well as local micro environmental factors, a testing platform with high flexibility will be useful to elucidate the role of each individual factors. 13.2 Microfluidic Techniques 13.2.1 Cellular Rheology Under Hypoxia We developed an in vitro microvascular model for sickle cell disease. This model allowed rheological measurements of single cells under controlled transient hypoxia through a double-layer polydimethylsiloxane (PDMS) structure. Development and initial applications of this model have been reported in our previous publications [33, 34]. The key idea of this model is to control the oxygen level in cell suspension by diffusing gas mixtures of specific oxygen concentrations through a gas-permeable PDMS film, while forcing individual sickle cells moving through a series of periodic micro gates under a differential pressure at body temperature (Fig. 13.1). Initially, sickle cells were deformable and can move freely through the micro gates in exposure to a gas mixture with atmospheric oxygen levels. When a hypoxic gas mixture was introduced, oxygen concentration in the cell channel reduced to the same level as the hypoxic gas due to a diffusion equilibrium. As a result, the deoxygenated sickle hemoglobin inside cells form into rigid fibers, turning initially deformable cells into rigid sickled cells. Fig. 13.1 In vitro microvascular model for sickle cell occlusion under transient hypoxia. (a) Schematic double-layer PDMS structure for gas diffusion. (b) Schematic of variations in cellular morphological sickling and obstructions (blue—rigid cells; red—deformable cells). (c) Experimental observation of single-cell rheology in microfluidic mimic of microvasculature. Highlighted cells are deformable while other cells are unable to pass through micro grates 90 E. Du
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