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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 quality standards for the establishment of appropriate testing methods. Methods: Peptide mapping, HPLC, electrophoresis and enzymatic activity assay were used to identify and purify tPA and its mutants. Results: RP-HPLC and peptide mapping could distinguish two tPA mutants with 3 amino acids difference; RP-HPLC, SDS-PAGE and SDS-polyacrylamide gel electrophoresis Can effectively separate the degradation products; different structures of tPA show different activity. Conclusion: This method is highly specific and actionable.