Massively Multiplexed Tip-Based Photochemical Lithography under Continuous Capillary Flow

Multiplexed microarrays-where different biological probes are spatially encoded onto a surface into spots with micrometer-scale diameters-have facilitated the rapid advancement of "omics'' research. Further miniaturization of feature diameters could increase the number of probes in a microarray, reduce the sample required for analysis, and decrease costs. Tip-based lithography (TBL) has gained popularity for patterning delicate, biologically active materials, but no versatile TBL-based multiplexing strategy has been devised. Here, we combine microfluidics, beam pen lithography, and photochemical surface reactions to create multiplexed arrays. For proof of concept, the thiol-ene reaction was optimized, and the reaction kinetics were analyzed. Subsequently, we created several patterns containing multiple fluorescent alkenes, where each pattern was designed to demonstrate a different capability of this instrument. This patterning strategy is a powerful approach to studying and optimizing organic reactions on surfaces and creating massively multiplexed arrays and, as such, could provide an entirely new approach for miniaturizing biochips or understanding interfacial reactivity.