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Counterbalancing hydrodynamic sample distortion effects increases resolution of free‐flow zone electrophoresis
Author(s) -
Weber Gerhard,
Bauer Johann
Publication year - 1998
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.1150190709
Subject(s) - distortion (music) , electrophoresis , flow (mathematics) , sample (material) , resolution (logic) , chemistry , materials science , biological system , chromatography , mechanics , physics , biology , computer science , optoelectronics , amplifier , cmos , artificial intelligence
On fractionation of highly heterogeneous protein mixtures, optimal resolution was achieved by forcing proteins to migrate through a preestablished pH gradient, until they entered a medium with a pH similar but not equal to their p I s. For this purpose, up to seven different media were pumped through the electrophoresis chamber so that they were flowing adjacently to each other, forming a pH gradient declining stepwise from the cathode to the anode. This gradient had a sufficiently strong band‐focusing effect to counterbalance sample distortion effects of the flowing medium as proteins approached their isoelectric medium closer than 0.5 pH units. Continuous free‐flow zone electrophoresis (FFZE) with high throughput capability was applicable if proteins did not precipitate or aggregate in these media. If components of heterogeneous protein mixtures had already started to precipitate or aggregate, in a medium with a pH exceeding their p I by more than 0.5 pH units, the application of interval modus and media forming flat pH gradients appeared advantageous.