A label-free assay for high sensitive detection of RNase based on two near IR fluorescence probes
A label-free assay for high sensitive detection of RNase based on two near IR fluorescence probes
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基于两个近红外荧光探针的无标记高灵敏检测 RNase
DOI：
10.1016/j.jlumin.2018.07.048
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发表时间：
2018-12
影响因子：
3.6
通讯作者：
Changying Yang
中科院分区：
文献类型：
--
作者：
Jinya Du;Na Huang;Zhouxuan Xiang;Yuzhi Dong;Qingyun Gao;Wei Yanga;Changying Yang
RNase的功能是切割ssRNA、dsRNA或DNA-RNA杂合链中的RNA，可直接用荧光探针辅助RNA进行分析。本文建立了一种基于荧光探针的非标记RNase检测方法，具有较高的灵敏度和特异性。两种TICT表征探针（H2和L2）在与RNA结合时表现出强发光。然后RNA酶水解底物RNA，使探针暴露于缓冲液中，导致荧光猝灭，引起“OFF-ON-OFF”荧光开关。我们成功地应用该方法检测了两种RNase（RNase A和RNase H），RNase A和RNase H的检测限分别为1.67×10−5 U/mL和1.06×10−4 U/mL。酶动力学再次证实了特异性裂解过程。此外，该方法还可用于筛选RNase抑制剂。RNaseOUT对RNase A有活性，IC 50值为0.88 U/mL，但对RNase H无活性。
RNase, whose function was cleaving RNA in ssRNA, dsRNA or DNA-RNA hybrid chain, can be analyzed directly by fluorescence probe assisted with RNA. In this paper, we constructed a none-labeled RNase assay based on fluorescence probe with high sensitivity and specificity. Two TICT characterize probes (H2 and L2) exhibited strong luminescence when bound with RNA. Then RNase hydrolysis substrate RNA exposing probe into buffer and resulted in fluorescence quench, causing “OFF-ON-OFF” fluorescence switch. We successfully applied the assay to detect two kind of RNase (RNase A and RNase H) with the detection limit of 1.67×10−5 U/mL for RNase A and 1.06×10−4 U/mL for RNase H, respectively. Enzyme kinetics confirmed again the specific cleaving process. Moreover, this approach could also be used for the screening of RNase inhibitors. RNaseOUT was active against RNase A with the IC50 values of 0.88 U/mL, but not against RNase H.
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