Therapeutic protein depots have limited clinical success because of the presence of critical preparation barriers such as low encapsulation, uncontrolled release, and activity loss during processing and storage. In the present study, we used our novel protein-nanoencapsulation (into sugar-glass nanoparticle; SGnP) platform to prepare a protein depot to overcome the abovementioned formidable challenges. The SGnP-mediated microparticle protein depot has been validated using four model proteins (bovine serum albumin, horseradish peroxidase, fibroblastic growth factor, and epidermal growth factor) and model biodegradable poly(lactic-co-glycolic acid) polymer system. The results show that our protein-nanoencapsulation-mediated platform provides a new generic platform to prepare a protein depot through the conventional emulsion method of any polymer and single/multiple protein systems. This protein depot has the required pharmaceutical properties such as high encapsulation efficiency, burst-free sustained release, and protein preservation during processing and storage, making it suitable for off-the-shelf use in therapeutic protein delivery and tissue engineering applications. © 2019 American Chemical Society.