Asymmetric multifunctional 3D cell microenvironments by capillary force assembly

The fabrication and characterization of advanced 3D cell culture microenvironments that enable systematic structure-property relationship studies are reported. The asymmetric multifunctional 3D cell microenvironments were obtained by capillary force-assisted assembly of microscale cubes at the water/air interface. The identical wettability of distinctly different microcubes, which exhibit different surface chemical functionalities or topographic nanostructures, drives the self-assembly into close-packed hexagonal aggregates with randomly arranged cubes. Hence with a library of a limited number of building blocks a very large number of distinct microenvironments with a unique trigonal pyramidal structure can be obtained in a combinatorial manner. The introduction of selective adsorption functionalities and initiator moieties for surface-initiated atom transfer radical polymerization (SI-ATRP), respectively, facilitated the immobilization of proteins such as fibronectin, and the grafting of passivating poly(acrylamide) (PAAm) brushes via SI-ATRP on selected side walls of the microwells. Thereby protein adsorption and protein mediated cell attachment was demonstrated to be varied according to the (a)symmetry of the 3D microenvironment. The random assembly of different cubes from a library comprising microcubes that are pre-functionalized or surface-structured exclusively on their top surface opens a pathway to generate a multitude of different microenvironments in a massively parallel combinatorial manner, enabling future systematic structure-property relationship studies with cells.