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Performance implications of chemical mobilization after microchannel IEF
Author(s) -
Tentori Augusto M.,
Herr Amy E.
Publication year - 2014
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.201400023
Subject(s) - microchannel , mobilization , chemistry , chromatography , nanotechnology , materials science , political science , law
Chemical mobilization following IEF enables single‐point detection of an ideally stationary equilibrium electrophoresis mode. Despite prior studies exploring optimization of chemical mobilization conditions and recent insight from numerical simulations, understanding of both chemical mobilization mechanisms and the implications of mobilization on IEF analytical performance remains limited. In this study, we utilize full‐field imaging of microchannel IEF to assess the performance of a range of canonical chemical mobilization schemes. We empirically demonstrate and characterize key areas where limited understanding of performance implications exists, including: the effects of mobilization solution p H and ion concentration, differences between ionic and zwitterionic mobilization, and diffusion as a source of zone broadening. We utilize S imul 5 simulations to gain insight into the sources of the measured performance differences. Measurements of the location, linearity, and slope of the IEF p H gradient (via fluorescent p H markers imaged before and during mobilization) as well as mobilization‐associated broadening of focused analytes were performed to quantify performance and determine the dominant sources of variability. Our results suggest that nonuniform broadening of the p H gradient and changes in the p H gradient linearity stem from conductivity nonuniformities in the separation channel and not diffusion‐associated band broadening during mobilization.