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A new crystallization system is described, which makes it possible to use an evaporation-based microfluidic crystallization technique for protein crystallization. The gas and water permeability of the used polydimethylsiloxane (PDMS) material enables evaporation of the protein solution in the microfluidic device. The rates of evaporation are controlled by the relative humidity conditions, which are adjusted in a precise and stable way by using saturated solutions of different reagents. The protein crystals could nucleate and grow under different relative humidity conditions. Using this method, crystal growth could be improved so that approximately 1 mm-sized lysozyme crystals were obtained more successfully than using standard methods. The largest lysozyme crystal obtained reached 1.57 mm in size. The disadvantage of the good gas permeability in PDMS microfluidic devices becomes an advantage for protein crystallization. The radius distributions of aggregrates in the solutions inside the described microfluidic devices were derived from in situ dynamic light scattering measurements. The experiments showed that the environment inside of the microfluidic device is more stable than that of conventional crystallization techniques. However, the morphological results showed that the protein crystals grown in the microfluidic device could lose their morphological stability. Air bubbles in microfluidic devices play an important role in the evaporation progress. A model was constructed to analyze the relationship of the rates of evaporation and the growth of air bubbles to the relative humidity.
