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This chapter discusses various techniques such as micro- and nanotechnologies, used in the fabrication of gradient biomaterials or scaffolds suitable for engineering tissue interfaces and how the gradient features of the biomaterials influence cellular behaviors such as adhesion, migration, differentiation, and heterotypic interactions during tissue organization. In addition, an overview of various gradient biomaterials and their physical, chemical, and biological classifications is provided. Finally, potential challenges and future directions of the emerging field of interface tissue engineering (ITE) are discussed.