Protein mapping of peanut extract with capillary electrophoresis

Abstract

Protein separation can be achieved with different modes of capillary electrophoresis, such as with capillary gel electroporesis (CGE) or with capillary zone electrophoresis (CZE). CZE protein mapping of peanut extract was approached in four different ways, combining neutral-coated or multilayer-coated capillaries with pHs well over or under the isoelectric point range of the proteins of interest. At acidic pHs, the mobility ranges of the major peanut allergens Ara h1, Ara h2, Ara h3, and Ara h6 were identified. Although the pH is a major factor in CZE separation, buffers with different compositions but with the same pH and ionic strength showed significantly different resolutions. Different components of the electrolyte were studied in a multifactorial design of experiment. CE-SDS and CZE proved to be suitable for protein mapping and we were able to distinguish different batches of peanut extract and burned peanut extract.

Polyelectrolyte Multilayers in Capillary Electrophoresis

“…Other techniques were used to assess properties of the SMIL coating, notably AFM and topography and recognition imaging (TREC) which showed the homogeneous surface, charge distribution, and reconstruction ability of the SMIL‐55 % PAMAMPS. Similarly, protein mapping of peanut extract was conducted with CZE by Villemet et al., [33] resulting in the characterization of major peanut allergens (Ara h1, Ara h2, Ara h3, Ara h6). A (PB/DS) 1.5 coating was used with a BGE made up of 100 mM phosphoric acid, 70 mM tris, and 3 g/L PS at pH 2.5, and the peaks were identified by comparison with Ara h standards.…”

“…3 not mentioned not mentioned 1.5 M acetic acid [43] Citric acid, lactic acid, 3-hydroxybutyric acid, pyroglutamic acid 3 > 200 not mentioned 6 (200 mM sodium phosphate) [33] Peanut allergens (Ara h 1, 2, 3, 6) RSD 0.16 7.0 (100 mM phosphate) [16] α-chymo A, Cyt C, Lyz, RNAse A RSD 17 % 2.5 (0.5 M acetic acid) PGLU [15] β-lac A, Lyz, Myo, RNAse A, TI 5 not mentioned À 54.9 RSD 1.5 % 2.5 (0.5 M acetic acid) P(lys,ser) HA [15] β-lac A, Lyz, Myo, RNAse A, TI…”

“…Likewise, Villemet et al [33] . investigated the effect of BGE pH on the separation on peanut allergens with a (PB/DS) 1.5 coated capillary and a BGE composed of 100 mM phosphoric acid, 70 mM Tris, and 3 g/L polysorbate 80 (PS) at pH 2.5.…”

“…Likewise, Villemet et al [33] investigated the effect of BGE pH on the separation on peanut allergens with a (PB/DS) 1.5 coated capillary and a BGE composed of 100 mM phosphoric acid, 70 mM Tris, and 3 g/L polysorbate 80 (PS) at pH 2.5. They observed that lowering the pH led to increased electrophoretic mobilities of the proteins and shorter migration times, but peak resolution was lost at pH 2.2.…”

Method development for mono‐ and disaccharides monitoring in cell culture medium by capillary and microchip electrophoresis

“…This could indicate adsorption to the capillary wall, which would be a problem, even more so for the dirtier cell culture medium samples. Neutral surfactants such as poloxamer 188, polysorbate 20, and polysorbate 80 could prevent adsorption [45]. The addition of these three surfactants in different concentrations to the BGE was tested.…”