Journal
JOURNAL OF IMMUNOLOGICAL METHODS
Volume 282, Issue 1-2, Pages 1-11Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jim.2003.07.008
Keywords
chemotactic chamber; silicon wafer; chemokine; neutrophils; eosinophils; basophils
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We have developed an optically accessible, horizontal chemotaxis apparatus consisting of an etched silicon substrate and a flat glass plate, both of which form two compartments with a 5-mum-deep microchannet in between. The device is held together with a stainless steel bolder with holes for injecting cells and a chemoattractant to the different compartments. Migration of cells in the channel is traced with time-lapse intervals using a CCD camera. By developing a method for aligning cells at the edge of the channel, we could successfully reduce the number of cells required for a chemotactic assay, depending on the experiment, to 100 or less. To prevent ceaseless flow of contents between the adjacent compartments via the communicating microchannel, a space at the top end of the holder was filled with medium after aligning the cells. By using a fluorescent probe, we demonstrated experimentally that a stable concentration gradient could be maintained. Furthermore, we determined theoretical details of the gradient established using a model chemokine and a computational fluid dynamics code. Reproducible kinetic results of cell migration were obtained using human neutrophils and IL-8 as a model. Migration of other cells such as eosinophils, basophils and Jurkat lymphocytes toward the appropriate chemokines were also demonstrated. (C) 2003 Elsevier B.V. All rights reserved.
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