Controlling Protein Crystal Nucleation by Droplet‐Based Microfluidics

Herein, we demonstrate the potential of droplet‐based microfluidics for controlling protein crystallization and generating single‐protein crystals. We estimated the critical droplet size for obtaining a single crystal within a microdroplet and investigated the crystallization of four model proteins to confirm the effect of protein molecular diffusion on crystallization. A single crystal was obtained in microdroplets smaller than the critical size by using droplet‐based microfluidics. In the case of thaumatin crystallization, a single thaumatin crystal was obtained in a 200 μm droplet even with high supersaturation. In the case of ferritin crystallization, the nucleation profile of ferritin crystals had a wider distribution than the nucleation profiles of lysozyme, thaumatin, and glucose isomerase crystallization. We found that the droplet‐based microfluidic approach was able to control the nucleation of a protein by providing control over the crystallization conditions and the droplet size, and that the diffusion of protein molecules is a significant factor in controlling the nucleation of protein crystals in droplet‐based microfluidics.