Dispersion of protein bands in a horseshoe microchannel during IEF

Shim, Jaesool; Dutta, Prashanta; Ivory, Cornelius F.

doi:10.1002/elps.200800728

Cited by 13 publications

(14 citation statements)

References 32 publications

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“…It is assumed that the ionic strengths do not affect the electrophoretic mobilities of ampholytes and proteins, as reported in Ref. 10. The initial concentrations of ampholytes and proteins are set as 0.2 mM and 2 l M, respectively, throughout the separation column.…”

Section: Resultsmentioning

confidence: 99%

“…Recently, due to the availability of high performance computers and the development of efficient parallel algorithms, it is possible to simulate several hundred ampholytes in order to form smooth profiles for both narrow-and broad-range IEF. 6,7 Although the aforementioned IEF simulations have been used to explain experimental behavior in 1-D 8,9 and twodimensional (2-D) [10][11][12] problems qualitatively, none of these models have accounted for the effect of temperature. All previous IEF models were based on the same reference temperature, with no consideration of Joule heating in the capillary tube or microchannels.…”

Section: Introductionmentioning

confidence: 99%

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Effect of Joule heating on isoelectric focusing of proteins in a microchannel

2014

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Electric field-driven separation and purification techniques, such as isoelectric focusing (IEF) and isotachophoresis, generate heat in the system that can affect the performance of the separation process. In this study, a new mathematical model is presented for IEF that considers the temperature rise due to Joule heating. We used the model to study focusing phenomena and separation performance in a microchannel. A finite volume-based numerical technique is developed to study temperature-dependent IEF. Numerical simulation for narrow range IEF (6 < pH < 10) is performed in a straight microchannel for 100 ampholytes and two model proteins: staphylococcal nuclease and pancreatic ribonuclease. Separation results of the two proteins are obtained with and without considering the temperature rise due to Joule heating in the system for a nominal electric field of 100 V/cm. For the no Joule heating case, constant properties are used, while for the Joule heating case, temperature-dependent titration curves and thermo-physical properties are used. Our numerical results show that the temperature change due to Joule heating has a significant impact on the final focusing points of proteins, which can lower the separation performance considerably. In the absence of advection and any active cooling mechanism, the temperature increase is the highest at the midsection of a microchannel. We also found that the maximum temperature in the system is a strong function of the DpK value of the carrier ampholytes. Simulation results are also obtained for different values of applied electric fields in order to find the optimum working range considering the simulation time and buffer temperature. Moreover, the model is extended to study IEF in a straight microchip where pH is formed by supplying H þ and OH À , and the thermal analysis shows that the heat generation is negligible in ion supplied IEF. V C 2014 AIP Publishing LLC.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Effect of Joule heating on isoelectric focusing of proteins in a microchannel

2014

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“…Dynamic computer simulation of IEF in presence of carrier ampholytes has demonstrated considerable value as a research tool . Simulations performed with a one‐dimensional (1D) model have been used to predict separation dynamics of low molecular mass amphoteric components and proteins, pH gradient formation and stability, as well as electrophoretic mobilization .…”

Section: Introductionmentioning

confidence: 99%

“…In carrier ampholyte‐based focusing, sample components are typically applied as a zone mixed with the carrier ampholytes. This configuration provided the basis for all IEF simulation studies mentioned above . The main purpose of the work described in the present paper was to explore different sampling strategies by computer simulation in order to establish optimized sampling for CIEF with electroosmotic mobilization and detection toward or at the cathodic capillary end .…”

Section: Introductionmentioning

confidence: 99%

Sampling strategies for capillary isoelectric focusing with electroosmotic zone mobilization assessed by high‐resolution dynamic computer simulation

et al. 2012

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The impact of initial sample distribution on separation and focusing of analytes in a pH 3–11 gradient formed by 101 biprotic carrier ampholytes under concomitant electroosmotic displacement was studied by dynamic high-resolution computer simulation. Data obtained with application of the analytes mixed with the carrier ampholytes (as is customarily done), as a short zone within the initial carrier ampholyte zone, sandwiched between zones of carrier ampholytes, or introduced before or after the initial carrier ampholyte zone were compared. With sampling as a short zone within or adjacent to the carrier ampholytes, separation and focusing of analytes is shown to proceed as a cationic, anionic, or mixed process and separation of the analytes is predicted to be much faster than the separation of the carrier components. Thus, after the initial separation, analytes continue to separate and eventually reach their focusing locations. This is different to the double-peak approach to equilibrium that takes place when analytes and carrier ampholytes are applied as a homogenous mixture. Simulation data reveal that sample application between two zones of carrier ampholytes results in the formation of a pH gradient disturbance as the concentration of the carrier ampholytes within the fluid element initially occupied by the sample will be lower compared to the other parts of the gradient. As a consequence thereof, the properties of this region are sample matrix dependent, the pH gradient is flatter, and the region is likely to represent a conductance gap (hot spot). Simulation data suggest that sample placed at the anodic side or at the anodic end of the initial carrier ampholyte zone are the favorable configurations for capillary isoelectric focusing with electroosmotic zone mobilization.

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“…Also, electric fieldinduced electroosmotic flow is not considered here because in most experimental work the channel surfaces are coated with methylcellulose or other chemicals to suppress electroosmosis. Ionic strength-dependent mobility corrections are not applied in this study, as their contribution is negligible [25].…”

Section: Assumptionsmentioning

confidence: 99%

A method to determine quasi‐steady state in constant voltage mode isotachophoresis

2011

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Identification of the steady state is very challenging in isotachophoresis (ITP); especially in complex microgeometries, such as dog-leg channels or cross-channel junctions. In this work, an elastic matching method is applied to determine the quasi-steady state in microscale ITP. In the elastic matching method, the similarity between two profiles is calculated by comparing intensity distribution of two images or profiles. To demonstrate this similarity-based analysis technique for ITP, a constant voltage mode ITP model is developed and applied to a five-component ITP system. Hydrochloric acid and caproic acid are used as the leader and terminator, respectively, while histidine is used as the counter-ion. Two sample components, acetic acid and benzoic acid, are separated under the action of an applied electric field in both straight and dog-leg microchannels. This analysis shows that conductivity profiles provide a better measure to determine the quasi-steady state in an ITP process. For a straight microchannel, the quasi-steady state is achieved in less than a minute with a total potential drop of 100 V in a 2 cm long channel. In a straight channel, a true steady state can be achieved for ITP with appropriate countercurrent flow where stationary zones are formed, but the time it takes to reach the steady state is much longer than the without counter flow case. The numerical results indicate that a steady state cannot be reached in a dog-leg microchannel because of sample dispersion and refocusing at and near the intersections and at the branch channels. However, the elastic matching method can be used to determine the quasi-steady state in a dog-leg microchannel.

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Dispersion of protein bands in a horseshoe microchannel during IEF

Cited by 13 publications

References 32 publications

Effect of Joule heating on isoelectric focusing of proteins in a microchannel

Effect of Joule heating on isoelectric focusing of proteins in a microchannel

Sampling strategies for capillary isoelectric focusing with electroosmotic zone mobilization assessed by high‐resolution dynamic computer simulation

A method to determine quasi‐steady state in constant voltage mode isotachophoresis

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