The aim of this study was to demonstrate electronic speckle pattern interferometry (ESPI) as a powerful tool in determining diffusion coefficients and partition coefficients for proteins in gels. ESPI employs a CCD camera instead of a holographic plate as in conventional holographic interferometry. This gives the advantage of being able to choose the reference state freely. If a hologram at the reference state is taken and compared to a hologram during the diffusion process, an interferometric picture can be generated that describes the refraction index gradients and thus the concentration gradients in the gel as well as in the liquid. MATLAB is then used to fit Fick's law to the experimental data to obtain the diffusion coefficients in gel and liquid. The partition coefficient is obtained from the same experiment from the flux condition at the interface between gel and liquid. This makes the comparison between the different diffusants more reliable than when the measurements are performed in separate experiments. The diffusion and partitioning coefficients of lysozyme, BSA, and IgG in 4% agarose gel at pH 5.6 and in 0.1 M NaCl have been determined. In the gel the diffusion coefficients were 11.2 +/- 1.6, 4.8 +/- 0.6, and 3.0 +/- 0.3 m(2)/s for lysozyme, BSA, and IgG, respectively. The partition coefficients were determined to be 0.65 +/- 0.04, 0.44 +/- 0.06, and 0.51 +/- 0.04 for lysozyme, BSA, and IgG, respectively. The current study shows that ESPI is easy to use and gives diffusion coefficients and partition coefficients for proteins with sufficient accuracy from the same experiment.
A study of the effect of light deflection during diffusion studies
of ethanol into agarose gel using holographic laser interferometry is
presented. Furthermore it also demonstrates how a diffusive flux could give
rise to a convective flux in holographic laser interferometry experiments. The
convective and diffusive mass transfer is also theoretically compared in both
a liquid phase and a gel phase for the ethanol-agarose system used.
The current study shows that errors due to light deflection in holographic
laser interferometry are extremely small and can be neglected. It also shows
the importance of designing the diffusion experiments to avoid natural
convection. In gels the convective flow is cancelled by the friction forces
between the liquid and the polymer network. However, in the liquid phase the
natural convection could occur even though the density differences in the
phase are small.
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