人血浆因子V蛋白免疫印迹分析及其临床应用

目的　建立人血浆因子Ｖ（ＦＶ）蛋白免疫印迹分析，对遗传性ＦＶ缺乏症家系成员血浆ＦＶ含量进行分析。方法　以３３％饱和硫酸铰从多克隆兔抗人ＦＶ抗血清中纯化ＩｇＧ，待测血浆行４％～１２％梯度的ＳＤＳ－ＰＡＧＥ，继之电转印至硝酸纤维膜上。结果　在３３０　ｋＤ附近，正常人混合血浆呈现一明显的条带，患者母亲对应条带的显色强度明显弱于正常对照，而患者则完全缺如，且未能检出其他较小的ＦＶ片段产物。结论　我们建立的人血浆ＦＶ蛋白免疫印迹分析法对于遗传性ＦＶ缺乏症的辅助诊断及分型具有重要作用。该家系属于Ｉ型遗传性ＦＶ缺乏。

Western blot of human plasma factor V and its clinical application

Objective To develope a western blot of human plasma factor V (FV) and conduct immunoblot analyses on riasma FV from a family with hereditary FV deficiency. Methods Polyclonal anti - human FV was purified from rabbit serum by precipitation with 33% ammonium sulate. SDS - PAGE analysis was performed on a 4% ~ 12% gradient gel and transfer of the proteins from the gel onto the nitrocellulose was accomplished. Results A single band was detected in plasma from pool normal plasma at position corresponding to a molecular weight of 330 000, while the counterpart was of a lighterstaining intensity for the patient's mother, and undetectable for the patient. Conclusion The western blot is conducive to the diagnosis and classification of hereditary FV deficiency. Analyses showed the presence of type I FV deficiency in the family.