Simultaneous analysis by capillary electrophoresis of five amyloid peptides as potential biomarkers of Alzheimer's disease.
We report here a CE method for the separation and quantitation of five amyloid peptides (Abeta1-42, 1-40, 1-39, 1-38, and 1-37) considered as potential biomarkers of Alzheimer's disease. These amyloid peptides have very similar structures. Sample preparation and storage conditions are critical parameters to ensure their solubility and to avoid the aggregation process in particular for Abeta1-42. Their solubility was found fully dependent on the NH(4)OH concentration that was employed initially to dissolve the lyophilized amyloid peptides. Conditions to achieve a full separation of these peptides were found using a dynamic coating with 1,4-diaminobutane (DAB). The linear decrease of their electrophoretic mobility highlighted an ion-pairing phenomenon between the peptides and DAB. The optimal background electrolyte was a 40 mM borate buffer, pH 9 containing 3 mM of DAB. Under these conditions, resolutions ranged from 1.3 to 2.4 with theoretical plates reaching 300,000. Under the retained conditions, we showed that adsorption of peptides to silica was negligible (recovery over 94.5%) and depletion effect of the background electrolyte was overcome. The method was finally validated in terms of linearity and repeatability and the limits of detection for the five Abeta peptides were estimated. The inter-day repeatability of the migration times was very satisfactory with RSDs less than 1.55%. The RSDs of the peak areas were below 5%. With this CE-UV method, limits of detection of the peptides ranged from 300 to 500 nM. We finally demonstrated that this method can be applied to real biological samples such as CSF.
Verpillot
Romain
R
Univ Paris-Sud, JE 2495, Protéines et Nanotechnologies en Sciences Séparatives, Faculté de Pharmacie, 92296 Châtenay-Malabry, France.
Otto
Markus
M
Klafki
Hans
H
Taverna
Myriam
M
D000544:Alzheimer Disease; D000595:Amino Acid Sequence; D064753:Ammonium Hydroxide; D016229:Amyloid beta-Peptides / Q000032:analysis* / Q000134:cerebrospinal fluid; D015415:Biomarkers / Q000032:analysis / Q000134:cerebrospinal fluid; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006878:Hydroxides / Q000737:chemistry; D016018:Least-Squares Analysis; D008969:Molecular Sequence Data; D009994:Osmolar Concentration; D010446:Peptide Fragments / Q000032:analysis* / Q000134:cerebrospinal fluid; D011700:Putrescine / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D016229:Amyloid beta-Peptides; D015415:Biomarkers; D006878:Hydroxides; D010446:Peptide Fragments; C078639:amyloid beta-protein (1-38); C078592:amyloid beta-protein (1-40); C075222:amyloid beta-protein (1-42); D064753:Ammonium Hydroxide; D011700:Putrescine