Separation of human fibrinopeptides by capillary zone electrophoresis.
We describe a method for the simultaneous determination of the five fibrinopeptide forms derived from the thrombin-promoted activation of human fibrinogen by capillary zone electrophoresis (CZE). The fibrinopeptide mixture was first desalted by a solid-phase extraction (SPE) step. The analysis was performed in reversed polarity in a highly cross-linked polyethylene glycol (PEG)-coated capillary with UV-light absorption detection at 200 nm. Several parameters including buffer concentration and pH, presence of an organic modifier, temperature, and applied voltage, have been tested. The best separations were obtained within 20 min, utilizing a 20 mM sodium phosphate buffer without organic modifier, in the narrow 6.1-6.2 pH range, at 25 degrees C, with an applied voltage of 20 kV. Quantitative analysis is made possible by the use of sheep fibrinopeptide A as an internal standard to correct for both extraction and injection errors.
Profumo
Aldo
A
S.C. Medicina Rigenerativa, Istituto Nazionale per la Ricerca sul Cancro, Genova, Italy. aldo.profumo@istge.it
Cardinali
Barbara
B
Cuniberti
Carla
C
Rocco
Mattia
M
D000818:Animals; D002851:Chromatography, High Pressure Liquid; D019075:Electrophoresis, Capillary / Q000379:methods*; D005344:Fibrinopeptide A / Q000031:analogs & derivatives* / Q000302:isolation & purification*; D005345:Fibrinopeptide B / Q000302:isolation & purification*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012756:Sheep
C500980:fibrinopeptide A, des-Ala(1)-; C500981:fibrinopeptide A, phospho-Ser(3)-; D005344:Fibrinopeptide A; D005345:Fibrinopeptide B; C034958:fibrinopeptide B, des-Arg(14)-