Biomaterial science has made enormous progress in the last few decades. Nonetheless, innovative biomaterials are still urgently needed to provide in vitro cell-culture models that more closely resemble three-dimensional (3-D) cell interactions and cyto-architectures in bodies and tissues. In this review, the recent advances toward this goal through molecular engineering of various designer self-assembling peptide scaffolds are discussed. These peptide scaffolds can be commercially and custom-tailor synthesized materials with high purity and may be not only useful for specific 3-D tissue cell cultures but also for tissue repair and regenerative therapies. Furthermore, these designer self-assembling peptide scaffolds have recently become powerful tools for regenerative medicine to repair nervous tissue, to stop bleeding in seconds, to repair infarctuated myocardia, as well as being useful medical devices for slow drug release.