TY - JOUR
T1 - Fast electrophoretic separation optimization using gradient micro free-flow electrophoresis
AU - Fonslow, Bryan R.
AU - Bowser, Michael T.
PY - 2008/5/1
Y1 - 2008/5/1
N2 - The continuous nature of micro free-flow electrophoresis (μ-FFE) was used to monitor the effect of a gradient of buffer conditions on the separation. This unique application has great potential for fast optimization of separation conditions and estimation of equilibrium constants. COMSOL was used to model pressure profiles in the development of a new μ-FFE design that allowed even application of a buffer gradient across the separation channel. The new design was fabricated in an all glass device using our previously published multiple-depth etch method (Fonslow, B. R.; Barocas, V. H.; Bowser, M. T. Anal. Chem. 2006, 78, 5369-5374, ref 1). Fluorescein solutions were used to characterize the applied gradients in the separation channel. Linear gradients were observed when buffer conditions were varied over a period of 5-10 min. The effect of a gradient of 0-50 mM hydroxypropyl-β-cyclodextrin (HP-β-CD) on the separation of a group of 4-fluoro7-nitro-2,1,3-benzoxadiazole (NBD-F) labeled primary amines was monitored as a proof of concept experiment. Direct comparisons to capillary electrophoresis (CE) separations performed under the same conditions were made. Gradient μ-FFE recorded 60 separations during a 5 min gradient allowing nearly complete coverage across a range of HP-β-CD concentrations. In comparison, 4 h were required to assess 15 sets of conditions across the same range of HP-β-CD concentrations using CE. Qualitatively, μ-FFE separations were predictive of the migration order and spacing of peaks in CE electropherograms measured under the same conditions. Data were fit to equations describing 1:1 analyte-additive binding to allow a more quantitative comparison between gradient μ-FFE and CE.
AB - The continuous nature of micro free-flow electrophoresis (μ-FFE) was used to monitor the effect of a gradient of buffer conditions on the separation. This unique application has great potential for fast optimization of separation conditions and estimation of equilibrium constants. COMSOL was used to model pressure profiles in the development of a new μ-FFE design that allowed even application of a buffer gradient across the separation channel. The new design was fabricated in an all glass device using our previously published multiple-depth etch method (Fonslow, B. R.; Barocas, V. H.; Bowser, M. T. Anal. Chem. 2006, 78, 5369-5374, ref 1). Fluorescein solutions were used to characterize the applied gradients in the separation channel. Linear gradients were observed when buffer conditions were varied over a period of 5-10 min. The effect of a gradient of 0-50 mM hydroxypropyl-β-cyclodextrin (HP-β-CD) on the separation of a group of 4-fluoro7-nitro-2,1,3-benzoxadiazole (NBD-F) labeled primary amines was monitored as a proof of concept experiment. Direct comparisons to capillary electrophoresis (CE) separations performed under the same conditions were made. Gradient μ-FFE recorded 60 separations during a 5 min gradient allowing nearly complete coverage across a range of HP-β-CD concentrations. In comparison, 4 h were required to assess 15 sets of conditions across the same range of HP-β-CD concentrations using CE. Qualitatively, μ-FFE separations were predictive of the migration order and spacing of peaks in CE electropherograms measured under the same conditions. Data were fit to equations describing 1:1 analyte-additive binding to allow a more quantitative comparison between gradient μ-FFE and CE.
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U2 - 10.1021/ac702367m
DO - 10.1021/ac702367m
M3 - Article
C2 - 18351751
AN - SCOPUS:42949086973
SN - 0003-2700
VL - 80
SP - 3182
EP - 3189
JO - Analytical Chemistry
JF - Analytical Chemistry
IS - 9
ER -