Protein crystallization remains a bottleneck in the goal to determine the protein's structure. A microfluidic device denoted the Crystal Optimizer (XOpt) has been developed to optimize the kinetic pathway for the nucleation and growth of protein crystals once the lead conditions have been identified by other screening methods. The device drives each of hundreds of drops of the candidate protein solution to different degrees of supersaturation for variable durations and then reduces the supersaturation to a low value. The concept is to find the two-step kinetic pathway that firstly quenches the drop for the right amount of time to nucleate just a few crystals and secondly reduces the supersaturation to a level that suppresses nucleation of additional crystals, but is still supersaturated enough for the crystals to grow slowly. Besides controlling concentration, the XOpt measures the solute and protein concentrations as a function of time so quantitative information on the actual kinetic pathway traversed by the samples can be obtained and exploited to design improved kinetic pathways in subsequent rounds of optimization. Addition of a simple temperature stage that generates a thermal gradient across the chip is used to find the optimal temperature of the optimal kinetic path. Two key points make the Crystal Optimizer innovative and useful. First, protein crystallization is an activated, non-equilibrium process and the XOpt is the sole microfluidic device designed to systematically optimize this physical feature in a controlled and reversible manner. Second, the XOpt is simple to use and inexpensive to manufacture.
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