Studies of cell-extracellular matrix (ECM) interactions within fibrous systems such as collagen or fibrin are challenging, particularly if peri-cellular stiffness cannot be monitored. Here we present our light-based method for non-invasive patterning of molecular crosslinking combined with multi-axes optical tweezers active microrheology to map ECM stiffness landscapes. This method allows us to generate prescribed stiffness gradients and associated anisotropies, which model stiffness of the natural peri-cellular ECM. Patterned crosslinking induces strain hardening and measured stiffness gradients are in agreement with predicted strain fields. Migratory cells respond to these gradients as assessed by change in F-actin distribution and morphological properties.
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