Stereomask lithography for multi-protein patterning

The advances of biologically-friendly micropatterning technologies have benefited many areas of biological and medical research, including quantitative biochemical assay, point-of-care devices, biosensing and regenerative medicine. Conventional micropatterning techniques, for example, photolithography and soft lithography, have seen encouraging adaptation to creating biological micropatterns in the last decades. However, they still have not completely addressed the major needs of constructing multi-object biological microarrays with single-cell resolution without requiring cleanroom access. In this chapter, we present a novel versatile biological lithography technique to achieve integrated multi-object patterning with high feature resolution and high adaptability to various biomaterials, referred to as stereomask lithography (SML). A novel three-dimensional stereomask has been developed for successive patterning of multiple objects. The stereomask consists of both patterned through holes, which layout new micropatterns and non-through recesses, which protect pre-existing features on the substrate. Furthermore, high-precision reversible alignment among multiple bio-objects is achieved by adopting a peg-in-hole design between the substrate and stereomasks. As demonstration, we have successfully used the SML technique to construct complex biological microenvironment with various bio-functional components at single-cell resolution.