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- Bohlin, Maria, et al.
(författare)
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Estimation of the amount of β2-glycoprotein I adsorbed at the inner surface of fused silica capillaries after acidic, neutral and alkaline pretreatment
- 2012
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Ingår i: Electrophoresis. - Weinheim, Germany : John Wiley & Sons. - 0173-0835 .- 1522-2683. ; 33:12, s. 1695-1702
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Tidskriftsartikel (refereegranskat)abstract
- Sample adsorption to the inner surface of fused silica capillaries is a common problem inCE when analyzingmacromolecules and is harmful to the analysis. We previously utilizedthe pH hysteresis effect of fused silica to facilitate electrophoresis of the strongly adsorbingprotein β2gpI in plain-fused silica capillaries at neutral pH. In the present paper, theeffect of different pretreatments of the capillary on the adsorption of the β2-glycoproteinI has been investigated using electroosmosis markers, SDS mobilization, and imagingbased on indirect immunofluorescence microscopy for direct visualization. The amountof β2gpI adsorbed on the surface was probed using all these independent techniques afterelectrophoresis at neutral pH on capillaries pretreated with HCl, background electrolyte(BGE), and NaOH. BGE pretreatment was included as a positive control. We found that80% or more of the starting material was adsorbed to the inner surface of the silicacapillaries during electrophoresis after pretreatment with only BGE or with NaOH, butafter acidic pretreatment the loss was consistently less than 20%. NaOH most efficientlyremoves adsorbed protein between runs. A theoretical calculation of the pH change ofthe BGE showed that electrolysis affects the pH more than the deprotonation of silanolsduring electrophoresis. We conclude that acidic pretreatment of fused silica capillariesdiminishes adsorption of β2gpI by decreasing charge-dependent wall adsorption.
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