Zimmerman T S, Voss R, Edgington T S
J Clin Invest. 1979 Nov;64(5):1298-302. doi: 10.1172/JCI109585.
We have examined the plasma Factor VIII/von Willebrand factor (FVIII/vWF) molecule from 16 patients with von Willebrand's disease, and have found no evidence of a significant decrease of carbohydrate content in 15 of these patients. FVIII/vWF was isolated by preparative counter immunoelectrophoresis directly from plasma using antibody to Factor VIII-related antigen, reduced in sodium dodecyl sulfate in the presence of urea, and electrophoresed in 5% polyacrylamide gels to separate the FVIII/vWF subunit from other proteins. Duplicate gels were stained by either the periodic acid-Schiff (PAS) reaction or by Coomassie Brilliant Blue G250. The ratio of Coomassie: PAS was determined by spectrophotometric scanning of the gels. Transferrin was used as an internal reference standard. The ratio for 23 normal individuals was 2.4+/-0.38 and the observed range was 1.8-3.8. 15 patients with von Willebrand's disease fell within this range. One patient independently reported as having decreased FVIII/vWF carbohydrate was also studied by this technique. A ratio of 6.8 was found, indicative of decreased, though not absent, carbohydrate. Cold insoluble globulin did not represent a significant contaminant in these analyses. 11 of the von Willebrand's disease patients with normal FVIII/vWF carbohydrate had abnormal crossed immunoelectrophoretic patterns characterized by absence of the less anodic forms of Factor VIII-related antigen. Four patients had normal patterns. These studies indicate that an absence or decrease of PAS reactive FVIII/vWF carbohydrate is not a consistent abnormality in von Willebrand's disease.
我们检测了16例血管性血友病患者的血浆因子VIII/血管性血友病因子（FVIII/vWF）分子，发现其中15例患者没有明显的碳水化合物含量降低的证据。通过制备性对流免疫电泳，使用针对因子VIII相关抗原的抗体直接从血浆中分离FVIII/vWF，在尿素存在的情况下于十二烷基硫酸钠中还原，并在5%聚丙烯酰胺凝胶中电泳，以将FVIII/vWF亚基与其他蛋白质分离。重复的凝胶用高碘酸-希夫（PAS）反应或考马斯亮蓝G250染色。考马斯亮蓝与PAS的比例通过凝胶的分光光度扫描来确定。转铁蛋白用作内部参考标准。23名正常个体的比例为2.4±0.38，观察到的范围为1.8 - 3.8。15例血管性血友病患者的比例在此范围内。对一名独立报告为FVIII/vWF碳水化合物减少的患者也采用该技术进行了研究。发现比例为6.8，表明碳水化合物减少，但并非没有。冷不溶性球蛋白在这些分析中不是显著的污染物。11例FVIII/vWF碳水化合物正常的血管性血友病患者交叉免疫电泳图谱异常，其特征是缺乏因子VIII相关抗原的阳极较弱形式。4例患者图谱正常。这些研究表明，PAS反应性FVIII/vWF碳水化合物的缺失或减少在血管性血友病中并非一致的异常表现。
