This paper describes a simple, versatile method of generating gradients in composition in solution or on surfaces using microfluidic systems. This method is based on controlled diffusive mixing of species in solutions that are flowing laminarly, at low Reynolds number, inside a network of microchannels. We demonstrate the use of this procedure to generate (1) gradients in the compositions of solutions, measured directly by colorimetric assays and (2) gradients in topography of the surfaces produced by generating concentration gradients of etching reagents, and then using these gradients to etch profiles into the substrate. The lateral dimensions of the gradients examined here, which went from 350 to 900 μm, are determined by the width of the microchannels. Gradients of different size, resolution, and shape have been generated using this method. The shape of the gradients can be changed continuously (dynamic gradients) by varying the relative flow velocities of the input streams of fluids. The method is experimentally simple and highly adaptable, and requires no special equipment except for an elastomeric relief structure that can be readily prepared by rapid prototyping. This technique provides a new platform with which to study phenomena that depend on gradients in concentration, especially dynamic phenomena in cell biology (chemotaxis and haptotaxis) and surface chemistry (nucleation and growth of crystals, etching, and Marangoni effects).

Although a wealth of knowledge about chemotaxis has accumulated in the past 40 years, these studies have been hampered by the inability of researchers to generate simple linear gradients instantaneously and to maintain them at steady state. Here we describe a device microfabricated by soft lithography and consisting of a network of microfluidic channels that can generate spatially and temporally controlled gradients of chemotactic factors. When human neutrophils are positioned within a microchannel, their migration in simple and complex interleukin-8 (IL-8) gradients can be tested. The cells exhibit strong directional migration toward increasing concentrations of IL-8 in linear gradients. Neutrophil migration halts abruptly when cells encounter a sudden drop in the chemoattractant concentration to zero ("cliff" gradient). When neutrophils are challenged with a gradual increase and decrease in chemoattractant ("hill" gradient), however, the cells traverse the crest of maximum concentration and migrate further before reversing direction. The technique described in this paper provides a robust method to investigate migratory cells under a variety of conditions not accessible to study by earlier techniques.