The recycling, development, and application of keratin-containing waste (e.g., hair, wool, feather, and so on) provide an important means to address related environmental pollution and energy shortage issues. The extraction of keratin and the development of keratin-based functional materials are key to solving keratin-containing waste pollution. Keratin-based biofilms are gaining substantial interest due to their excellent characteristics, such as good biocompatibility, high biodegradability, appropriate adsorption, and rich renewable sources, among others. At present, keratin-based biofilms are a good option for various applications, and the development of keratin-based biofilms from keratin-containing waste is considered crucial for sustainable development. In this paper, in order to achieve clean production while maintaining the functional characteristics of natural keratin as much as possible, four important keratin extraction methods—thermal hydrolysis, ultrasonic technology, eco-friendly solvent system, and microbial decomposition—are described, and the characteristics of these four extraction methods are analysed. Next, methods for the preparation of keratin-based biofilms are introduced, including solvent casting, electrospinning, template self-assembly, freeze-drying, and soft lithography methods. Then, the functional properties and application prospects of keratin-based biofilms are discussed. Finally, future research directions related to keratin-based biofilms are proposed. Overall, it can be concluded that the high-value conversion of keratin-containing waste into regenerated keratin-based biofilms has great importance for sustainable development and is highly suggested due to their great potential for use in biomedical materials, optoelectronic devices, and metal ion detection applications. It is hoped that this paper can provide some basic information for the development and application of keratin-based biofilms.