Error analysis in equilibrium dialysis: evaluation of adsorption phenomena.
Various sources of error in equilibrium dialysis may lead to inaccurate results of binding experiments: (i) the finite time of dialysis; (ii) the Donnan effects; (iii) the adsorption of ligand to the membrane; and (iv) release of contaminating material from dialysis casings. These errors were analyzed for a polynucleotide-oligopeptide model system with particular regard to adsorption phenomena and the underlying mechanisms. Adsorption data were treated according to Freundlich and Langmuir isotherms. The latter turned out to be more appropriate for the consideration of adsorption phenomena with respect to a minimum error propagation. Furthermore, it was shown that the degree of adsorption varies with ionic strength and temperature and could be interpreted in terms of polyelectrolyte theory. The kinetics of both adsorption and of the ligand distribution between the polymer and buffer compartments follow first order at the beginning of dialysis which is in line with a simple diffusion process. After 13-15 h data deviate from first order kinetics indicating an alteration in the transport mechanism. The effects of errors on binding parameters were determined and a detailed protocol for correction is presented allowing one to obtain binding data from equilibrium dialysis experiments with the required degree of accuracy. The fundamental principles and results for the system under investigation generally apply to all protein-ligand systems.
Desoye
G
G
Max-Planck-Institute for Biophysical Chemistry, Göttingen, F.R.G.
D000327:Adsorption; D003956:Dialysis; D007700:Kinetics; D015203:Reproducibility of Results