CAS Key Lab of Health Informatics, Institute of Biomedical and Health Engineering, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen 518055, Guangdong, PR China.
Nanoscale. 2012 Apr 7;4(7):2454-62. doi: 10.1039/c2nr12005d. Epub 2012 Feb 28.
This work presents a method to visualize the degradation of exogenous DNA in living cells using a novel type of activatable fluorescence imaging probe. Deoxyribonuclease (DNase)-activatable fluorescence probes (DFProbes) are composed of double strands deoxyribonucleic acid (dsDNA) which is labeled with fluorophore (ROX or Cy3) and quencher on the end of one of its strands, and stained with SYBR Green I. In the absence of DNase, DFProbes produce the green fluorescence signal of SYBR Green I. In the presence of DNase, SYBR Green I is removed from the DFProbes and the labeled fluorophore is separated from the quencher owing to the degradation of DFProbes by DNase, resulting in the decrease of the green fluorescence signal and the occurrence of a red fluorescence signal due to fluorescence resonance energy transfer (FRET). DNase in biological samples was detected using DFProbes and the fluorescence imaging in living cells was performed using DFprobe-modified Au nanoparticles. The results show that DFProbes have good responses to DNase, and can clearly visualize the degradation of exogenous DNA in cells in real time. The well-designed probes might be useful in tracing the dynamic changes of exogenous DNA and nanocarriers in vitro and in vivo.
本工作提出了一种使用新型的可激活荧光成像探针可视化活细胞中外源 DNA 降解的方法。脱氧核糖核酸酶(DNase)可激活荧光探针(DFProbes)由双链脱氧核糖核酸(dsDNA)组成,dsDNA 的一端标记有荧光团(ROX 或 Cy3)和猝灭剂,并用 SYBR Green I 染色。在没有 DNase 的情况下,DFProbes 产生 SYBR Green I 的绿色荧光信号。在存在 DNase 的情况下,由于 DNase 对 DFProbes 的降解,SYBR Green I 从 DFProbes 中被去除,并且标记的荧光团与猝灭剂分离,导致绿色荧光信号的降低以及由于荧光共振能量转移(FRET)而发生红色荧光信号。使用 DFProbes 检测生物样品中的 DNase,并使用修饰有 DFprobe 的金纳米粒子进行活细胞中的荧光成像。结果表明,DFProbes 对 DNase 具有良好的响应,可以实时清晰地可视化细胞中外源 DNA 的降解。设计良好的探针可能有助于追踪体外和体内外源 DNA 和纳米载体的动态变化。