Graphene-based nanomaterials, such as graphene oxide and reduced graphene oxide, have been attracting increasing attention in the field of biology and biomedicine over the past few years. Incorporation of these novel materials with drug, gene, photosensitizer and other cargos to construct novel delivery systems has witnessed rapid advance on the basis of their large surface area, distinct surface properties, excellent biocompatibility and pH sensitivity. Moreover, the inherent photothermal effect of these appealing materials enables them with the ability of killing targeting cells via a physical mechanism. Recently, more attentions have been attached to tissue engineering, including bone, neural, cardiac, cartilage, musculoskeletal, and skin/adipose tissue engineering, due to the outstanding mechanical strength, stiffness, electrical conductivity, various two-dimensional (2D) and three-dimensional (3D) morphologies of graphene-based nanomaterials. Herein, emerging applications of these nanomaterials in bio-imaging, drug/gene delivery, phototherapy, multimodality therapy and tissue engineering were comprehensively reviewed. Inevitably, the burgeon of this kind of novel materials leads to the endeavor to consider their safety so that this issue has been deeply discussed and summarized in our review. We hope that this review offers an overall understanding of these nanomaterials for later in-depth investigations.