Analysis of von Willebrand factor multimers by simultaneous high- and low-resolution vertical SDS-agarose gel electrophoresis and Cy5-labeled antibody high-sensitivity fluorescence detection.
Analysis of von Willebrand factor (vWF) multimers allows classification of the subtypes of von Willebrand disease (vWD) in human serum and platelet lysates. A novel method for multimer analysis of vWF by 2-chamber, vertical (sodium dodecyl sulfate), agarose gel electrophoresis, designed for comparing discontinuous high- and low-resolving gels for plasma and platelets, followed by Western blotting and high-sensitivity fluorescence detection (HSFD) of cyanine (Cy)5-labeled vWF multimers is presented. HSFD shows that this method has high discriminatory power for visualization and densitometric analysis of platelets and plasma vWF multimers in various types of vWD and allows rapid classification of vWD types, to separate types 2A and 2B. The described procedures of vWF multimer analysis with high-sensitivity Cy5 fluorescence detection and direct comparison of high- and low-resolving gels for screening and detection of the complete range of high- and low-molecular vWF multimers is efficient and useful for screening, detecting, and classifying vWD subtypes and makes this method diagnostically and clinically relevant.
Ott
Helmut W
HW
Medical University Innsbruck, Anichstr 35, A-6020 Innsbruck, Austria.
Griesmacher
Andrea
A
Schnapka-Koepf
Mirjam
M
Golderer
Georg
G
Sieberer
Andrea
A
Spannagl
Michael
M
Scheibe
Burghardt
B
Perkhofer
Susanne
S
Will
Kerstin
K
Budde
Ulrich
U
D002232:Carbocyanines; D003720:Densitometry; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D005453:Fluorescence; D006801:Humans; D012967:Sodium Dodecyl Sulfate; D014841:von Willebrand Factor / Q000032:analysis*
D002232:Carbocyanines; C085321:cyanine dye 5; D014841:von Willebrand Factor; D012967:Sodium Dodecyl Sulfate