Patterning proteins and cells using soft lithography

Abstract

This review describes the pattering of proteins and cells using a non-photolithographic microfabrication technology, which we call `soft lithography’ because it consists of a set of related techniques, each of which uses stamps or channels fabricated in an elastomeric (`soft') material for pattern transfer. The review covers three soft lithographic techniques: microcontact printing, patterning using microfluidic channels, and laminar flow patterning. These soft lithographic techniques are inexpensive, are procedurally simple, and can be used to pattern a variety of planar and non-planar substrates. Their successful application does not require stringent regulation of the laboratory environment, and they can be used to pattern surfaces with delicate ligands. They provide control over both the surface chemistry and the cellular environment. We discuss both the procedures for patterning based on these soft lithographic techniques, and their applications in biosensor technology, in tissue engineering, and for fundamental studies in cell biology.

Introduction

This review describes techniques for patterning the properties and structures of surfaces at the molecular level, and for using these patterns to control both the adsorption of proteins to these surfaces and the attachment of cells to them. The ability to generate patterns of proteins and cells on surfaces is important for biosensor technology [1], [2], [3], [4] for tissue engineering [5], and for fundamental studies of cell biology [6], [7], [8]. The placement of biological ligands at well-defined locations on substrates is required for certain biological assays, for combinatorial screening, and for the fabrication of biosensors. Biosensors based on living cells [3], [9], [10], [11], [12] can also be used for environmental and chemical monitoring; accurate positioning of the cells used for sensing on these devices is critical for monitoring the status of the cells. Control over the positioning of cells is also important for cell-based screening, in which individual cells need to be accessed repeatedly to perturb them and to monitor their response. Tissue engineering may require that cells be placed in specific locations to create organized structures. Patterning techniques that control both the size and shape of the cell anchored to a surface, and the chemistry and topology of the substrate to which the cell is attached, are also extremely useful in understanding the influence of the cell–material interface on the behavior of cells [7], [8], [13], [14].

Photolithography is the technique that has been used most extensively for patterning proteins and cells. For example, photolithography can be used to generate patterns by photoablating proteins preadsorbed to a silicon or glass surface [15], by immobilizing proteins on thiol-terminated siloxane films that have been patterned by irradiation with UV light [16], and by covalently linking proteins to photosensitive groups [17]. Although photolithography is a technique that is highly developed for patterning, the high costs associated with photolithographic equipment, and the need for access to clean rooms, make this technique inconvenient for biologists. Photolithography is not well suited for introducing either specific chemical functionalities, or delicate ligands required for bio-specific adsorption, onto surfaces. Photolithography cannot be used to pattern non-planar substrates. While photolithography can be used to produce patterns with features smaller than 1 μm, this resolution may be unnecessary for many applications of patterning in cell biology.

We have developed a set of microfabrication techniques that is an alternative to photolithography for patterning surfaces used in biochemistry and biology. We call this set `soft lithography’ [18], [19], [20], [21], [22], [23], because each of the techniques uses stamps or channels fabricated in an elastomeric (`soft') material for pattern transfer or modification. Soft lithographic techniques are not expensive, are procedurally simple, can be used to pattern a variety of different planar and non-planar substrates, and do not require stringent control over the laboratory environment for their successful application.

This review will focus on the patterning of proteins and cells using soft lithographic techniques, in programs carried out in our group and by others. The patterning of proteins and cells by other techniques will not be discussed in this review, other than by reference (Table 1).