Fabrication of patterned multicomponent protein gradients and gradient arrays using microfluidic depletion

We demonstrate that depletion effects in the fluids used to fill a poly(dimethylsiloxane) microfluidic device can be used in conjunction with its design rules to generate patterned protein gradients. The linear portions of these structures can be designed to present gradients of bound protein coverage-varying from near-saturation to effectively zero-over distances ranging from a few hundred micrometers to more than I cm by design. Such patterns can be developed in a simple, single-channel form as well as in a multichannel gradient array of more complex design. The patterning protocols also support the use of multiple protein sources, and we demonstrate an assembly process mediated by a protein that inhibits adsorption to generate a gradient array in pixel form. We describe examples of multiple protein gradient patterns along with simple immunoassays to illustrate the scope of the methodology, the activity of the patterned proteins, and their recognition in gradient form on a surface. These gradients should prove useful to studies in biosensor and bioassay development and as substrates for cell culture to study growth and motility.