Dual-photoresponsive substrates enabling light-induced single-cell patterning and sorting of nonadherent mammalian...

Single-cell arrays are essential tools for visualizing cellular heterogeneity and for linking cellular phenotypes observed by imaging to intracellular molecular systems through molecular analyses of cells retrieved after imaging. However, conventional single-cell array techniques often suffer from limited applicability due to the narrow microstructures required for single-cell trapping. In this study, we developed a dual-photoresponsive surface that enables single-cell photopatterning without any microstructures, as well as the selective light-induced release of individual cells of interest. The surface is based on a photoclickable material coating, onto which a photocleavable cell-anchoring material is photopatterned for single-cell trapping. We demonstrated light-induced patterning of mammalian immune cells through binding to patterned cell-anchoring materials, enabling microstructure-free single-cell array formation on a flat glass substrate. Furthermore, photopatterned individual cells were selectively released via localized light irradiation and subsequently retrieved at the single-cell level using cell pickers. These results represent the first demonstration of selective cell isolation from photoclickable surfaces following cell patterning and phenotypic observation. This surface is expected to be broadly applicable to a wide range of single-cell analyses by freely tailoring cell-anchoring molecules, designed cell patterns, and downstream molecular analyses.