Hamer R J, Koedam J A, Beeser-Visser N H, Sixma J J
Biochim Biophys Acta. 1986 Oct 17;873(3):356-66. doi: 10.1016/0167-4838(86)90084-1.
Human factor VIII was purified from commercial factor VIII concentrate with a 12% yield. The specific coagulant activity of purified factor VIII was 8,000 units/mg. In the presence of SDS the purified factor VIII consisted of a variety of polypeptides on polyacrylamide gels, ranging between Mr 80,000 and Mr 208,000. In the absence of SDS the purified factor VIII showed an apparent molecular weight of 270,000 upon Sephadex G200 gel-filtration. The purified factor VIII could be activated by thrombin, which resulted in the disappearance of Mr 108,000-208,000 polypeptides in favor of an Mr 92,000 polypeptide. Treatment with factor Xa also activated factor VIII, whereas treatment with activated protein C resulted in the inactivation of coagulant activity. Coagulant-active 125I-factor VIII was prepared using a lactoperoxidase radioiodination procedure. This 125I-factor had the same characteristics as unlabeled factor VIII. All polypeptides could be precipitated with monoclonal antibodies directed against factor VIII. With 125I-factor VIII a pIapp of 5.7 was found in the presence of urea.
从市售凝血因子 VIII 浓缩物中纯化出人凝血因子 VIII，产率为 12%。纯化后的凝血因子 VIII 的比凝血活性为 8000 单位/毫克。在十二烷基硫酸钠（SDS）存在的情况下，纯化后的凝血因子 VIII 在聚丙烯酰胺凝胶上由多种多肽组成，分子量在 80000 至 208000 之间。在没有 SDS 的情况下，经葡聚糖 G200 凝胶过滤，纯化后的凝血因子 VIII 的表观分子量为 270000。纯化后的凝血因子 VIII 可被凝血酶激活，这导致分子量为 108000 - 208000 的多肽消失，转而形成分子量为 92000 的多肽。用因子 Xa 处理也可激活凝血因子 VIII，而用活化蛋白 C 处理则导致凝血活性丧失。采用乳过氧化物酶放射性碘化方法制备了具有凝血活性的 125I - 凝血因子 VIII。这种 125I - 凝血因子与未标记的凝血因子 VIII具有相同的特性。所有多肽都可用针对凝血因子 VIII 的单克隆抗体沉淀。对于 125I - 凝血因子 VIII，在有尿素存在的情况下发现其表观等电点为 5.7。
