Chondroitin sulfate (CS) is a sulfated glycosaminoglycan (GAG) that is usually found attached to proteins as part of a proteoglycan. The use of CS-based biomaterials in the field of tissue engineering applications has been intensively growing over the past decades because CS is a biopolymer with the major advantages of being biodegradable, biocompatible, easily available, and highly versatile. In vitro and in vivo studies have shown that CS-based biomaterials upregulate cartilage-specific gene expression in chondrocytes and mesenchymal stem cells and also support osteogenic differentiation by increasing the effectiveness of bone anabolic growth factors. Chondroitin sulfate proteoglycan plays a key role during the acute recovery stage after spinal cord injury by activating microglia/macrophages and modulating neurotrophic factor secretion. In addition, CS-based materials promote the wound-healing process and stimulate the regeneration of skin defects. Moreover, CS can be used to construct high toughness gels by having a double network structure. Taken together, CS-based biomaterials would be a useful material for successful replacement and regeneration of damaged cartilage, bone, skin, and neural tissues.