论文部分内容阅读
基于适配体的高特异识别能力和可扩增性,构建了经磁分离后实时定量PCR检测重组人促红细胞生成素-α(rHuEPO-α)的新方法;针对实际血清样品中高丰度蛋白质等的干扰,利用碱基互补配对原理设计合成了分别与适配体两端引物区结合的互补链,通过凝胶迁移阻滞分析(EMSA)筛选出最佳的互补链,并将生物素化的互补链连接到链霉亲和素磁珠上,以此为探针捕获复杂基质中形成的待测复合物。研究结果表明,结合该样品前处理策略,该方法可成功地应用于正常人血清中的rHuEPO-α定量检测,检出限为25pmol/L,线性范围为50 pmol/L～50 nmol/L。
Based on the high specific recognition ability and amplifiability of aptamers, a new method of real-time quantitative PCR for the detection of recombinant human erythropoietin-α (rHuEPO-α) was established. The high abundance protein And so on. Based on the principle of complementary base pairing, the complementary strands that were respectively bound to the primer regions of aptamers were designed and synthesized. The best complementary strand was screened by gel permeation retardation assay (EMSA), and the biotinylated Of the complementary strand to streptavidin beads as a probe to capture complexes formed in a complex matrix. The results showed that the method was successfully applied to the quantitative determination of rHuEPO-α in normal human serum with a detection limit of 25 pmol / L and a linear range of 50 pmol / L to 50 nmol / L.