Effects of ampholyte concentration on protein behavior in on‐chip isoelectric focusing

The effects of mobility corrections on carrier ampholytes are studied at various ampholyte concentrations to understand protein behavior during IEF. IEF simulations are conducted in the presence of 25 biprotic carrier ampholytes within a pH range of 6–9 after applying the Onsager–Debye–Hückel correction to the carrier ampholytes. Two model proteins with ten charge states but without ionic strength corrections are allowed to focus under an electric field of 300 V/cm in a 1 cm long channel. The IEF simulation results show that higher ionic strengths (50 – 100 mM) cause significant changes in the transient movement as well as the final focused profiles of both ampholytes and proteins. The time required for a single, well‐defined peak to form increases with ionic strength when Onsager corrections are applied to the carrier ampholytes. For a particular ampholyte concentration, the space‐averaged conductivity does not change during the final focusing stage, but the magnitude of space averaged conductivity is different for different ampholyte concentration. The simulation results also reveal that at steady‐state ionic strength profiles remain flat throughout the channel except at the locations of proteins where a significant change in ampholyte concentration is obtained.

[1]  Athonu Chatterjee,et al.  Generalized numerical formulations for multi-physics microfluidics-type applications , 2003 .

[2]  J. Dufrêche,et al.  Transport equations for concentrated electrolyte solutions: Reference frame, mutual diffusion , 2002 .

[3]  W. Thormann,et al.  Computer simulation and experimental validation of the electrophoretic behavior of proteins. 2: Model improvement and application to isotachophoresis , 1992 .

[4]  Prashanta Dutta,et al.  Isoelectric focusing in a poly(dimethylsiloxane) microfluidic chip. , 2005, Analytical chemistry.

[5]  T. van de Goor,et al.  Electrophoretic mobilities of large organic ions in nonaqueous solvents: Determination by capillary electrophoresis in propylene carbonate, N,N‐dimethylformamide, N,N,‐dimethylacetamide, acetonitrile and methanol , 2002, Electrophoresis.

[6]  Bohuslav Gas,et al.  Optimization of background electrolytes for capillary electrophoresis: II. Computer simulation and comparison with experiments , 2002, Electrophoresis.

[7]  Champak Das,et al.  Conductivity properties of carrier ampholyte pH gradients in isoelectric focusing , 2005, Electrophoresis.

[8]  D. Saville,et al.  Mathematical modeling and computer simulation of isoelectric focusing with electrochemically defined ampholytes. , 1981, Biophysical chemistry.

[9]  D. C. Henry.  The cataphoresis of suspended particles. Part I.—The equation of cataphoresis , 1931 .

[10]  M Bier,et al.  Electrophoresis: mathematical modeling and computer simulation. , 1983, Science.

[11]  Prashanta Dutta,et al.  Multistage isoelectric focusing in a polymeric microfluidic chip. , 2005, Analytical chemistry.

[12]  B. Gaš,et al.  Determination of cationic mobilities and pKa values of 22 amino acids by capillary zone electrophoresis , 2004, Electrophoresis.

[13]  V. Kašička,et al.  Investigation of the effect of ionic strength of Tris‐acetate background electrolyte on electrophoretic mobilities of mono‐, di‐, and trivalent organic anions by capillary electrophoresis , 2005, Electrophoresis.

[14]  B. Gaš,et al.  Determination of limiting mobilities and dissociation constants of 21 amino acids by capillary zone electrophoresis at very low pH. , 2006, Journal of chromatography. B, Analytical technologies in the biomedical and life sciences.

[15]  D. Saville,et al.  Computer simulation and experimental validation of the electrophoretic behavior of proteins. , 1989, Analytical chemistry.

[16]  C. Lucy,et al.  Prediction of electrophoretic mobilities. 3. Effect of ionic strength in capillary zone electrophoresis. , 1999, Analytical chemistry.

[17]  B. Gale,et al.  Effect of carrier ionic strength in microscale cyclical electrical field-flow fractionation. , 2006, Analytical Chemistry.

[18]  Bohuslav Gas,et al.  Simul 5 – Free dynamic simulator of electrophoresis , 2006, Electrophoresis.

[19]  Prashanta Dutta,et al.  Modeling and simulation of IEF in 2‐D microgeometries , 2007, Electrophoresis.

[20]  E. Bender.  Numerical heat transfer and fluid flow. Von S. V. Patankar. Hemisphere Publishing Corporation, Washington – New York – London. McGraw Hill Book Company, New York 1980. 1. Aufl., 197 S., 76 Abb., geb., DM 71,90 , 1981 .

[21]  J. Deconinck,et al.  Relaxation effect on the Onsager coefficients of mixed strong electrolytes in the mean spherical approximation. , 2007, The journal of physical chemistry. B.