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MicroRNA靶向的放射性标记探针的制备及条件优化
引用本文:马 超,康 磊,王荣福. MicroRNA靶向的放射性标记探针的制备及条件优化[J]. 肿瘤学杂志, 2013, 19(12): 918-924
作者姓名:马 超  康 磊  王荣福
作者单位:北京大学第一医院;北京大学第一医院;北京大学第一医院
基金项目:国家自然科学基金(81101065、81071183、31100604);国家重大科学仪器设备开发专项(2011YQ03011409);高等学校博士学科点新教师专项科研基金(20110001120043)
摘    要:[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。

关 键 词:锝[99mTc]  microRNA(miRNA)  放射性标记
收稿时间:2013-11-20

Preparation and Optimization of 99mTc-radiolabeled Antisense Oligonucleotide Targeting microRNA
Affiliation:Peking University First Hospital
Abstract:[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.
Keywords:technium-99m  microRNA  radiolabeling
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