Preparative capillary zone electrophoresis using a dynamic coated wide‐bore capillary

Abstract

Preparative capillary zone electrophoresis separations of cytochrome c from bovine and horse heart are performed efficiently in a surfactant-coated capillary. The surfactant, dimethylditetradecylammonium bromide (2C(14)DAB), effectively eliminated protein adsorption from the capillary surface, such that symmetrical peaks with efficiencies of 0.7 million plates/m were observed in 50-microm id capillaries when low concentrations of protein were injected. At protein concentrations greater than 1 g/L, electromigration dispersion became the dominant source of band broadening and the peak shape distorted to triangular fronting. Matching of the mobility of the buffer co-ion to that of the cytochrome c resulted in dramatic improvements in the efficiency and peak shape. Using 100 mM bis(2-hydroxyethyl)imino-tris(hydroxymethyl)methane phosphate buffer at pH 7.0 with a 100-microm id capillary, the maximum sample loading capacity in a single run was 160 pmol (2.0 microg) of each protein.

Protein attachment onto silica surfaces – a survey of molecular fundamentals, resulting effects and novel preventive strategies in CE

“…The efficiency was also reduced if protein concentrations were raised. In parallel, the coating stability decreased with increasing capillary id, since the volume that can be saturated by 2C 12 DAB during detachment increases [201,203]. Furthermore, Yeung et al [204] have applied 2C 14 DAB capillary coating in offline coupling CE with MALDI-MS via fraction collection and the coating which had to be renewed between subsequent separations.…”

From micro to macro: Conversion of capillary electrophoretic separations of biomolecules and bioparticles to preparative free‐flow electrophoresis scale

“…Peptides in collected fractions are later on concentrated by evaporation of the solvent and off‐line characterized by MS or by amino acid and sequence analysis. In cationic surfactant‐coated 100 μm id fused‐silica capillary, effectively eliminating protein adsorption, a complete micropreparative separation of 2 μg (160 pmol) of bovine and horse cytochrome c was achieved 34. CE with replaceable sieving matrix (entangled polymer networks) in 180 μm id capillary was used for isolation of nanomolar quantities of DNA fragments in 5 μL volume fractions of 0.25×TBE buffer for subsequent downstream processing (cloning) 35.…”

Cationic poly(methacryl oxyethyl trimethylammonium) and its poly(ethylene glycol)‐grafted analogue as capillary coating materials in electrophoresis

“…4 Among cationic surfactants, didodecyldimethylammonium bromide performs as a semipermanent coating, [5][6][7] and its applicability in preparative CE was shown. 8 Polybrene TM (hexadimethrine bromide) is a common polyelectrolyte for controlling the electroosmotic flow (EOF); however, in most cases a low concentration of Polybrene TM must also be added to the BGE solution. 9 Over the years, various adsorbed polymer coatings for electrophoresis have been developed to avoid the need for additives.…”

“…The use of long double‐chain cationic surfactants as semipermanent coatings for CE in mixed organic–water solvents was recently demonstrated 4. Among cationic surfactants, didodecyldimethylammonium bromide performs as a semipermanent coating,5–7 and its applicability in preparative CE was shown 8. Polybrene™ (hexadimethrine bromide) is a common polyelectrolyte for controlling the electroosmotic flow (EOF); however, in most cases a low concentration of Polybrene™ must also be added to the BGE solution 9.…”