We describe a protocol for the microfabrication of the gradient-generating microfluidic device that can generate spatial and temporal gradients in well-defined microenvironment. In this approach, the gradient-generating microfluidic device can be used to study directed cell migration, embryogenesis, wound healing, and cancer metastasis.
A. Microfabrication of the gradient-generating microfluidic device
B. Experimental setup
Cells exposed to stable concentration gradients of EGF in a microfluidic device migrated toward higher concentrations. The directional orientation of cell migration, chemotactic index, motility of migrating cells were investigated by cell tracking analysis. Therefore, this gradient-generating microfluidic platform could be useful for studying cancer metastasis, embryogenesis, and axon guidance.
Material Name | Tipo | Company | Catalogue Number | Comment |
---|---|---|---|---|
Dextran-FITC | Reagent | Sigma Aldrich | FD10S | Fluorescein isothiocyanate (FITC) conjugated-dextran (10kD) |
hr-EGF | Invitrogen | 13247-051 | human recombinant Epidermal growth factor | |
PDMS | K.R. Anderson Co. Inc. | 2065622 | Poly(dimethylsiloxane) (PDMS), Dow Corning Sylgard 184 (8.6 lb) | |
Negative photoresist | Microchem | SU-8 50 | ||
Si wafer | silicone wafer, 4 inch | |||
Petri dishes | ||||
Polyethylene tubing | Becton Dickinon | PE 20 | ||
PBS | Invitrogen | |||
Fibronectin | ||||
NIH 3T3 | cell-line | fibroblast cells | ||
Inverted microscope | Nikon | TE 2000 |