Intravascular thrombosis is a main cause of multiple cardiovascular diseases. A high thrombolytic activity of the microbial fibrinolytic enzyme Subtilisin QK-2, which is highly homologous to Nattokinase, shows great exploitable potential in thrombolytic therapy. However, the lack of a sensitive detection method limits the further analysis of Subtilisin QK-2 in vivo. We prepared a polyclonal antibody and four monoclonal antibodies (IgG1, titers of 1:500,000) to establish a sensitive sandwich ELISA for Subtilisin QK-2 detection. The limit of detection (LOD) of this ELISA was 1.160 ng/mL. The linear range of the standard curve was 1.96-250 ng/mL (R2 = 0.9912). The cut-off value was 0.236. Subsequently, a pharmacokinetic dose (IV bolus) was administered and analyzed with the established ELISA. The concentration-time profiles were best fitted to a two-compartment model. T1/2α values for doses of 2 mg/kg, 4 mg/kg, and 8 mg/kg were 29.90 ± 10.02 min, 27.17 ± 1.96 min, and 21.83 ± 9.95 min, and T1/2β values were 144.43 ± 49.73 min, 173.46 ± 52.58 min, and 159.49 ± 48.75 min, respectively. Subtilisin QK-2 was eliminated through a mechanism with first-order kinetics. In conclusion, this study provides useful data for further research and clinical applications of Subtilisin QK-2 in the treatment of cardiovascular diseases.

中文翻译：

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使用新型夹心酶联免疫吸附测定法在大鼠模型中推注静脉内注射后，对枯草杆菌蛋白酶QK-2进行定量药代动力学评估。

血管内血栓形成是多种心血管疾病的主要原因。与纳豆激酶高度同源的微生物纤溶酶Subtilisin QK-2具有很高的溶栓活性，在溶栓治疗中显示出巨大的开发潜力。但是，缺乏灵敏的检测方法限制了枯草杆菌蛋白酶QK-2在体内的进一步分析。我们准备了多克隆抗体和四种单克隆抗体（IgG1，效价为1：500,000）来建立用于枯草杆菌蛋白酶QK-2检测的灵敏夹心ELISA。该ELISA的检出限（LOD）为1.160 ng / mL。标准曲线的线性范围为1.96-250 ng / mL（R2 = 0.9912）。截止值为0.236。随后，给药药代动力学剂量（静脉推注）并用已建立的ELISA分析。浓度-时间曲线最适合两室模型。2 mg / kg，4 mg / kg和8 mg / kg剂量的T1 /2α值为29.90±10.02分钟，27.17±1.96分钟和21.83±9.95分钟，T1 /2β值为144.43±49.73分钟，分别为173.46±52.58分钟和159.49±48.75分钟。枯草杆菌蛋白酶QK-2是通过一级动力学机制消除的。总之，这项研究为枯草杆菌蛋白酶QK-2在心血管疾病治疗中的进一步研究和临床应用提供了有用的数据。