Abstract
In this paper, we demonstrate the feasibility of using in situ Dynamic Light Scattering (DLS) to monitor counter-diffusion crystallization experiments in capillaries. Firstly, we have validated the quality of the DLS signal in thin capillaries, which is comparable to that obtained in standard quartz cuvettes. Then, we have carried out DLS measurements of a counter-diffusion crystallization experiment of glucose isomerase in capillaries of different diameters (0.1, 0.2 and 0.3 mm) in order to follow the temporal evolution of protein supersaturation. Finally, we have compared DLS data with optical recordings of the progression of the crystallization front and with a simulation model of counter-diffusion in 1D.
Highlights
The crystallization of any given protein [1,2,3] is a process that can be outlined in three successive and interrelated stages
In situ Dynamic Light Scattering (DLS) Measurements in Capillaries The volume of solution analysed by the DLS system has been previously estimated at around 1 pL [14]
Initially it was necessary to establish whether it is possible to obtain a clean DLS signal similar to that obtained in quartz cuvettes without any special distortion due to the geometry of the capillary or the plastic walls of the Granada Crystallization Boxes-Domino (GCB-D)
Summary
The crystallization of any given protein [1,2,3] is a process that can be outlined in three successive and interrelated stages. The rate at which supersaturation is achieved is one of the most important parameters in protein crystallization since it defines a very specific temporal pathway through the phase diagram [5]. The nucleation step [5] can be understood as a process by which, stochastically in space and time, a new phase with lower chemical potential appears in solution. Stable protein nuclei are formed from a dynamical size distribution of density fluctuations that are driven by the supersaturated state of the solution. Crystal growth, the stable aggregates or nuclei that have formed during nucleation start to grow at the expense of the remaining protein in solution. The growth of crystals consumes the protein, thereby reducing the supersaturation value of the solution and driving the system back towards equilibrium
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