Limits...
Monitoring and scoring counter-diffusion protein crystallization experiments in capillaries by in situ dynamic light scattering.

Oberthuer D, Melero-García E, Dierks K, Meyer A, Betzel C, Garcia-Caballero A, Gavira JA - PLoS ONE (2012)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Institute for Biochemistry and Molecular Biology, Laboratory for Structural Biology of infection and inflammation, University of Hamburg, Hamburg, Germany.

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.

Show MeSH

Related in: MedlinePlus

Comparison of data obtained from DLS measurements, optical microscopy records and the numerical simulation.The data at 18.7 and 24.5 mm have not been drawn because it does not show disturbance of the size distribution. The green horizontal line corresponds to original optical microscopy data plotted as ‘time vs distance’ instead of ‘distance vs time’, using the same limits as those used to plot the DLS and the simulation data. The axis of the original optical microscopy data have not been drawn for clarity.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3366972&req=5

pone-0033545-g003: Comparison of data obtained from DLS measurements, optical microscopy records and the numerical simulation.The data at 18.7 and 24.5 mm have not been drawn because it does not show disturbance of the size distribution. The green horizontal line corresponds to original optical microscopy data plotted as ‘time vs distance’ instead of ‘distance vs time’, using the same limits as those used to plot the DLS and the simulation data. The axis of the original optical microscopy data have not been drawn for clarity.

Mentions: In order to compare DLS data with optical microscopy observations, we prepared a counter-diffusion experiment under identical conditions and recorded the progress of the crystallization process by registering the distance of the furthest observable crystal from the entrance of the capillary at different times. Figure 3 illustrates how the disturbance observed in the size distribution at each position occurs earlier in time than the interpolated appearance of crystals, as expected. Only at the last point of measurement, where we know that the nucleation rate is already low, the perturbation of the DLS signal took a bit longer to be detected.


Monitoring and scoring counter-diffusion protein crystallization experiments in capillaries by in situ dynamic light scattering.

Oberthuer D, Melero-García E, Dierks K, Meyer A, Betzel C, Garcia-Caballero A, Gavira JA - PLoS ONE (2012)

Comparison of data obtained from DLS measurements, optical microscopy records and the numerical simulation.The data at 18.7 and 24.5 mm have not been drawn because it does not show disturbance of the size distribution. The green horizontal line corresponds to original optical microscopy data plotted as ‘time vs distance’ instead of ‘distance vs time’, using the same limits as those used to plot the DLS and the simulation data. The axis of the original optical microscopy data have not been drawn for clarity.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3366972&req=5

pone-0033545-g003: Comparison of data obtained from DLS measurements, optical microscopy records and the numerical simulation.The data at 18.7 and 24.5 mm have not been drawn because it does not show disturbance of the size distribution. The green horizontal line corresponds to original optical microscopy data plotted as ‘time vs distance’ instead of ‘distance vs time’, using the same limits as those used to plot the DLS and the simulation data. The axis of the original optical microscopy data have not been drawn for clarity.
Mentions: In order to compare DLS data with optical microscopy observations, we prepared a counter-diffusion experiment under identical conditions and recorded the progress of the crystallization process by registering the distance of the furthest observable crystal from the entrance of the capillary at different times. Figure 3 illustrates how the disturbance observed in the size distribution at each position occurs earlier in time than the interpolated appearance of crystals, as expected. Only at the last point of measurement, where we know that the nucleation rate is already low, the perturbation of the DLS signal took a bit longer to be detected.

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Institute for Biochemistry and Molecular Biology, Laboratory for Structural Biology of infection and inflammation, University of Hamburg, Hamburg, Germany.

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.

Show MeSH
Related in: MedlinePlus