MicroRNA靶向的放射性标记探针的制备及条件优化

[目的]以microRNA-21（miR-21）为靶点,探索制备放射性核素锝[99mTc]标记的反义寡核苷酸探针的方法与实验条件。[方法]设计化学结构修饰的反义寡核苷酸序列并进行合成,利用双功能螯合剂NHS-MAG3偶联待标记探针和99mTc,优化不同实验条件下的标记率及放化纯,并对标记产物进行凝胶电泳鉴定及稳定性评价。[结果]降低氯化亚锡的用量、减少99mTc的反应体积、适度延长标记反应时间均可显著提高标记率,最高可达97%。标记探针在血清37℃条件中孵育6h其放化纯仍可维持在96%以上。凝胶电泳结果证实核酸条带与放射性分布一致。[结论]通过优化探针设计和标记反应条件能够大大提高标记率和放化纯度,无需纯化即可进一步应用,达到简化制备程序的目的。

Preparation and Optimization of 99mTc-radiolabeled Antisense Oligonucleotide Targeting microRNA

[Purpose] To explore the methods and optimized reaction conditions for the preparation of99mTc radiolabeled antisense oligonucleotide targeted microRNA-21.[Methods] The antisense oligonucleotide probe with chemical and structural modifications was designed and synthesized.This probe was radiolabeled with 99mTc via the conjugation of a bifunctional chelator Sacetylmercaptoacetyltriglycine（NHS-MAG3）.The labeled probe was tested for its radiolabeling efficiency,purity and its stability under different reaction conditions,as well as the identification with gel electrophoresis.[Results] The labeling efficiency reached to 97% under optimized reaction conditions of different volume of SnCl2·2H2O,99mTc and reaction time.The radiochemical purity of 99mTc-anti-miR in fresh human serum at 37℃ could maintained more than 96% after 6h.[Conclusions] The labeling efficiency can be improved greatly by optimizing design and reaction conditions.Therefore,the probe could be prepared easily and simply for further applications without further purification.