A novel optical assay system for the quantitative measurement of chemotaxis.
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-microm-deep microchannel in between. The device is held together with a stainless steel holder 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.
Kanegasaki
Shiro
S
Effector Cell Institute in Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguroku, Tokyo 153-8904, Japan. shirokan@ims.u-tokyo.ac.jp
Nomura
Yuka
Y
Nitta
Nao
N
Akiyama
Shuichi
S
Tamatani
Takuya
T
Goshoh
Yasuhiro
Y
Yoshida
Takashi
T
Sato
Tsuyoshi
T
Kikuchi
Yuji
Y
D001491:Basophils / Q000502:physiology; D002465:Cell Movement; D002634:Chemotaxis, Leukocyte; D004804:Eosinophils / Q000502:physiology; D004867:Equipment Design; D006801:Humans; D009504:Neutrophils / Q000502:physiology