基于核酸适体的比色技术检测肿瘤标志物血管内皮生长因子

发布时间：2018-04-21 22:33

本文选题：核酸适体 + 磁珠　；参考：《南方医科大学学报》2017年02期

【摘要】：目的基于核酸适体和磁珠,提出一种检测肿瘤标志物血管内皮生长因子(VEGF)的比色方法。方法核酸适体与标记脲酶的单链结合,通过生物素-亲和素的特殊识别作用固定在链霉亲和素修饰的磁珠上;待检测的肿瘤VEGF标志物与适体结合形成茎环结构,使标记脲酶的单链脱落下来;经磁分离,转移上清液,加入到含一定量尿素和酚红试剂的溶液中。利用上清液中的脲酶水解尿素产生氨,使得溶液的p H值升高,导致溶液的颜色变化,通过肉眼或紫外分光光度计分析显色结果。结果该检测体系在最佳条件下,检测VEGF的线性范围为0.1~10 pmol/L,检测限达0.06 pmol/L。并利用此体系,检测人肺癌患者血清中的VEGF浓度,其结果与ELISA方法检测出的结果基本一致。将本方法与ELISA方法同步盲法测定10例血清样本,结果显示本方法的准确率为90%,表明其具有较高的有效性和敏感性。结论该检测方法操作简单方便,同时具有较高的灵敏度和选择性,在临床VEGF检测中具有潜在的应用前景。
[Abstract]:Objective to develop a colorimetric method for the detection of vascular endothelial growth factor (VEGF), a tumor marker, based on aptamer and magnetic beads. Methods Nucleic acid aptamer combined with a single strand of urease labeled urease was immobilized on the magnetic beads modified by streptavidin through the special recognition of biotin-avidin, and the tumor VEGF markers to be detected combined with the aptamer to form the stem ring structure. After magnetic separation the supernatant was transferred and added to the solution containing a certain amount of urea and phenolic red reagent. Urea was hydrolyzed by urease in supernatant to produce ammonia. The pH value of the solution was increased and the color of the solution was changed. The result of color development was analyzed by naked eye or ultraviolet spectrophotometer. Results under the optimal conditions, the linear range of detection of VEGF was 0.1 渭 mol / L and the detection limit was 0.06 pmol / L. The concentration of VEGF in serum of human lung cancer patients was detected by this system, and the results were consistent with those detected by ELISA method. The results showed that the accuracy of the method was 90, which indicated that the method was effective and sensitive. Conclusion this method is easy to operate and has high sensitivity and selectivity. It has potential application prospect in clinical VEGF detection.
【作者单位】：福州市第一医院中西医结合肿瘤科;福建医科大学药学院;
【基金】：国家自然科学基金(21375017) 福州市科技计划项目(2013-S-122-4) 福建省科技厅重点项目(2013Y0045) 福建省自然科学基金(2013J06003) 福建省高等学校新世纪优秀人才支持计划(JA13130)~~
【分类号】：R730.43

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