A quantitative CE method to analyse tau protein isoforms using coated fused silica capillaries.
We evaluated the potential of CE to analyse different isoforms of unphosphorylated recombinant tau protein and for separating one phosphorylated tau from the respective unphosphorylated protein. Different capillary coatings such as polyacrylamide, poly-(ethylene oxide) and polybrene (PB) were evaluated to overcome the poor efficiencies obtained with fused-silica capillary. Although peak asymmetry values were quite similar for the three investigated coatings, the peak efficiencies were 35-fold and 5-fold higher with PB coating than with polyacrylamide and poly(ethylene oxide) coatings, respectively. The recovery percentage (over 97%) was satisfactory and confirmed the efficacy of PB coating to limit the adsorption of tau protein to capillary walls. Moreover, PB coating produced higher repeatability for migration times (RSD values <1.2%) in comparison to the neutral coatings. The potential of PB-modified capillary in producing high resolutive separations of one phosphorylated tau isoform from its unphosphorylated counterpart and of a mixture of phosphorylated and unphosphorylated tau peptides was demonstrated with 50 mM phosphate buffer pH 3.0. The separation of unphosphorylated tau isoform 352 (Tau-352) from Tau-352 phosphorylated in vitro by the mitogen-activated protein kinase ERK2, was accomplished in less than 15 min.
Bohoyo
Diego
D
Univ Paris-Sud, Laboratoire des Protéines et Nanotechnologies en Sciences Séparatives, Faculté de Pharmacie, Châtenay-Malabry, France.
Le Potier
Isabelle
I
Rivière
Céline
C
Klafki
Hans
H
Wiltfang
Jens
J
Taverna
Myriam
M
D000180:Acrylic Resins / Q000737:chemistry*; D019075:Electrophoresis, Capillary; D006583:Hexadimethrine Bromide / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000737:chemistry*; D020033:Protein Isoforms / Q000032:analysis*; D012822:Silicon Dioxide / Q000737:chemistry*; D013499:Surface Properties; D013997:Time Factors; D016875:tau Proteins / Q000032:analysis*; D000099166:Polyacrylamides
D000180:Acrylic Resins; D006583:Hexadimethrine Bromide; D011092:Polyethylene Glycols; D020033:Protein Isoforms; D012822:Silicon Dioxide; D016875:tau Proteins; D000099166:Polyacrylamides