Abstract
Human tissues and organs exhibit complex hierarchical and gradient structures that are essential to their function and should be recapitulated within biomaterial scaffolds targeting their regeneration. Unidirectional freezing, an ice templating technique where ice acts as a porogen, is uniquely suited to recapitulating the architectural anisotropy, gradients, and hierarchical transitions of human tissues, but ice templating of polymeric systems, including silk fibroin, remains less well understood than their colloidal counterparts. To address this, a versatile and accessible freezing setup for silk that allows tuning of freezing parameters including the polymer cooling rate (30 degrees C min(-1) to 2 degrees C min(-1)) and ice solidification velocity (2.5 to 0.6 mm min(-1)) using liquid nitrogen, is developed. Real time visual and thermal monitoring of the freezing process for multiple silk concentrations (2-10% wt/v) and material states (liquid, hydrogels) is performed and the conditions are correlated with pore morphology. Unprecedented control over pore size (100-90 000 mu m(2)) and pore morphology (cellular-lamellar), consistent pore alignment, and generation of gradient porosity in silk scaffolds are demonstrated. For the first time, impact of shear thinning behavior of silk in ice crystal formation is demonstrated, showing non-linear and complex freezing phenomena in silk.