重组人组织纤维蛋白溶酶原激活剂及突变体的分析

Analysis of Recombinant Human Tissue Plasminogen Activator and Its Mutants

目的:为制订质量标准建立相应的检测方法。方法:应用肽图分析、高效液相色谱、电泳和酶活性测定等方法对tPA及突变体进行鉴别、纯度检查和活性测定。结果:反相高效液相色谱和肽图分析方法能区分相差3个氨基酸的2种tPA突变体;反相高效液相色谱、分子排阻高效液相色谱和SDS-聚丙烯酰胺凝胶电泳方法能有效分离降解产物;不同结构的tPA表现的活性不同。结论:本文方法专属性强,且可操作。

Objective: To establish the individual methods for the specification of tPA, K2tPA and TNK - tPA. Method: Peptide mapping, HPLC, SDS - PAGE were used for identification, impurities separation and potency assay. Results: Two tPA mutants with 3 amino acid residues differences can be separated by peptide mapping and RP - HPLC; various degraded products can be separated from tPA or its mutants by RP - HPLC, HPSEC and SDS -PAGE; tPA with different structures show different activity units. Conclusion: These methods were specific and available for quality control of tPA and its mutants.