High‐resolution computer simulation of electrophoretic mobilization in isoelectric focusing

Abstract

Cationic and anionic electrophoretic mobilization for focusing of hemoglobins (Hb's) in the presence of 100 carrier ampholytes covering a pI range of 6.00-7.98 was studied by computer simulation at a constant current density of 300 A/m(2). Electropherograms that would be produced by whole column imaging and by single detectors placed at different locations along the focusing column are presented. Upon mobilization, peak heights of the Hb zones decrease, but the zones retain a relatively sharp constant profile and are migrating at a constant velocity. A further peak decrease occurs during readjustment at the locations of the original buffer/column interfaces, indicating that detection sensitivity is the lowest at these locations. An anionic carrier ampholyte mobility smaller than that of its cationic species produces a cathodic drift which is smaller than the transport rate used for electrophoretic mobilization. Compared to the case with equal mobilities of carrier ampholyte species, a small increase (decrease) is predicted for the cationic (anionic) mobilization rate within the focusing column. Simulation data suggest that electrophoretic mobilization after focusing and focusing with concurrent electrophoretic mobilization are comparable isotachophoretic processes that occur when there is an uninterrupted flux of an ion through the focusing column. Cathodic drift caused by unequal mobilities of the species of carrier ampholytes, electrophoretic mobilization, and decomposition occurring at the pH gradient edges are related electrophoretic processes.

Dynamic computer simulations of electrophoresis: A versatile research and teaching tool

“…15), electrophoretic mobilization after focusing [132] and IEF with concurrent electrophoretic mobilization [160] are ITP based processes (see Section 4.3). In a different application, Dubrovčáková et al [72] performed simulations of ITP separations in which a neutral complexing agent (a cyclodextrin in this case) was added to the electrolytes.…”

“…With the significant improvement in computer power over the last twenty years, the capacity of GENTRANS could be increased to handle up to 150 components and to perform high-resolution IEF simulations at experimental current densities [53,54,132,159,160,177]. Mao et al [53] presented simulation data of a pH gradient spanning seven pH units from 3-10, with the pH gradient made from 140 Electrophoresis 2010, 31, 726-754 ampholytes, and low molecular mass amphoteric dyes as sample constituents.…”

“…This was remarkable agreement given that the input data (pK and mobility values) of carrier ampholytes and the dyes used were not accurately known and that the composition of the commercial ampholyte mixture is largely unknown. A subsequent study by Mosher and Thormann [132,159] revisited the focusing and stabilization phases of IEF under high-resolution conditions. The short (less than 10 min) separation phase is followed by a lengthy stabilization phase, which is characterized by changes that progress from the column ends towards neutrality and the formation of non-linear pH gradients.…”

“…This also caused a gradual pH loss at the anodic end, while the cathodic end remained unchanged causing a change in the profile of the pH gradient. Similarly, focusing with mobilization towards the anode is obtained having salt added to the anolyte instead of the catholyte [132]. Cationic and anionic electrophoretic mobilization after focusing of hemoglobins in presence of 100 carrier ampholytes covering a pI range from 6.00 to 7.98 was studied by computer simulation at a constant current density of 300 A/m 2 [132].…”

“…New models with sophisticated integration schemes emerged and the number of applications increased. Models that are currently in use in various laboratories are the 1-D simulators GENTRANS of Mosher et al [44,49,[51][52][53][54][129][130][131][132] and SIMUL5 of Hruška et al [48] (available at http://www.natur.cuni.cz/ Gas), which succeeded SIMUL 4.0 [47]. Interesting new models are those of Bercovici et al [69] (available as the Stanford's Public Release Electrophoretic Separation Solver (SPRESSO), http://microfluidics.stanford.edu/spresso), the 2-D model of Shim et al [70,71,135,136] and the models of Yang and his collaborators [68,138].…”