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Effects of Ampholyte Dissociation Constants on Protein Separation in On-Chip Isoelectric Focusing

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Abstract:

Numerical simulations are presented for ampholyte-based isoelectric focusing in 2D microgeometries. In this study, model proteins are focused in the presence of 25 biprotic ampholytes under an applied electric field. Each protein is considered as a simple polypeptide having ten charge states, while the biprotic ampholytes are selected to generate a shallow pH range of 6 to 9. Straight and contraction-expansion microchannels are considered here, and a nominal electric field of 300 V/cm is maintained for separation of proteins. Six distinct values of ΔpKs between 1 and 3.5 are investigated for ampholytes to form pH profiles in a 1 cm long microchannel. Simulation results show that relatively larger values of ΔpKpK > 3 are required to form stepless pH profiles in the system. The peak heights and differential resolutions of focused proteins are much higher for lower values of ΔpK for which a stepped pH profile is evident. For each protein, the time it takes for the two edges of a peak to merge increases linearly with ΔpK, while the focusing time goes up exponentially with increasing ΔpK. Both merging and focusing times of protein are higher for contraction-expansion microchannel than those of straight microchannel. For a particular value of ΔpK, the contracted "Zoom" region of contraction-expansion channel is able to form more tightly focused bands than the expanded region.

Keywords: BIPROTIC AMPHOLYTES; ISOELECTRIC FOCUSING; MICROCHANNEL; PH GRADIENT; PROTEIN

Document Type: Research Article

DOI: https://doi.org/10.1166/jnn.2008.011

Publication date: 2008-07-01

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  • Journal for Nanoscience and Nanotechnology (JNN) is an international and multidisciplinary peer-reviewed journal with a wide-ranging coverage, consolidating research activities in all areas of nanoscience and nanotechnology into a single and unique reference source. JNN is the first cross-disciplinary journal to publish original full research articles, rapid communications of important new scientific and technological findings, timely state-of-the-art reviews with author's photo and short biography, and current research news encompassing the fundamental and applied research in all disciplines of science, engineering and medicine.
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