特异性结合白色念珠菌（1，3）-β-D-葡聚糖单链DNA适配子的筛选及其应用  

Screening of ssDNA aptamers specifically binding to （1,3）-β-D-glucan from Candida albicans and their applications

作　　者：唐晓磊 华影[2] 袁纯辉 

机构地区：[1]芜湖市第二人民医院检验科,241000 [2]阜阳职业技术学院生化工程学院,236031 [3]武汉大学基础医学院免疫系,430072

出　　处：《国际生物制品学杂志》2016年第1期1-7,共7页International Journal of Biologicals

摘　　要：目的通过指数富集的配基系统进化技术（systematic evolution of ligands by exponentialenrichment, SELEX）筛选能与高亲和力（1，3）-β-D-葡聚糖特异性结合的适配子，并用该适配子建立双适配子夹心酶联寡聚核苷酸吸附试验（enzyme-linked oligonucleotide assay, ELONA）来对深部真菌感染血浆进行辅助诊断。方法提取白色念珠菌ATCC10231株细胞壁（1，3）-β-D-肛葡聚糖，通过SELEX筛选获得能与（1，3）-βD-葡聚糖特异性结合的高亲和力单链DNA适配子，并用该适配子建立双适配子夹心ELONA来检测深部真菌感染血浆中的（1，3）-β-D-葡聚糖。结果从白色念珠菌细胞壁中成功提取了高聚合度（1，3）-β-D-葡聚糖，并通过体外酶解获得可溶性低聚合度（1，3）-β-D-葡聚糖作为筛选靶标。用SELEX进行12轮筛选后，从初始单链DNA文库中获得2个高亲和力适配子Au1和AD1，检测发现它们并非结合于（1，3）-β-D-葡聚糖的同一表位。双适配子夹心ELONA检测深部真菌感染血浆中（1，3）-β-D-葡聚糖的特异性和敏感度分别为91．94％和92．31％。结论从初始单链DNA文库中成功获得可与（1，3）-β-D-葡聚糖特异结合的高亲和力适配子，为深部真菌感染新型诊断试剂的研发奠定了基础。Objective To screen high affinity aptamers binding to （1,3）-β-D-glucan by systematic evolution of ligands by exponential enrichment （SELEX） and establish double aptamers sandwich enzyme-linked oligonucleotide assay （ELONA） to help diagnosis of invasive fungal infection in plasma. Methods （1,3）-β-D-glucan from cell wall of Candida albicans ATCC10231 strain was extracted and purified. The high affinity ssDNA aptarners specifically binding to （ 1,3）-β-D-glucan were selected by SELEX. The aptamers were used to establish double aptamers sandwich ELONA for detecting （1,3）-β-D-glucan in plasma of invasive fungal infection. Results The high polymerization degree （1,3）-β-D-glucan was extracted successfully from Candida albicans and the soluble low polymerization degree （l, 3）-β-D-glucan was obtained as target by enzymolysis in vitro. Two high affinity aptamers AU1 and AD1 were screened from the initial ssDNA library after 12 rounds selection by SELEX, and these two aptmers bound different epitopes. The specificity and sensitivity of the double aptamers sandwich ELONA for detecting （1, 3）-β-D-glucan in plasma of invasive fungal infection were 91. 94% and 92. 31% respectively. Conclusion The selection of high affinity aptamers specifically binding to （1,3）-β-D-glucan from initial ssDNA library is successful, and lays foundation for the development of novel invasive fungal infection diagnostic agents.

关 键 词：适体  核苷 念珠菌  白色 葡聚糖类 SELEX适体技术 

分 类 号：R978.5[医药卫生—药品]