Monitoring Telomerase Activity in Living Cells with High Sensitivity Using Cascade Amplification Reaction-Based Nanoprobe.
Huanhuan FanHuarong BaiQin LiuHang XingXiao-Bing ZhangWeihong TanPublished in: Analytical chemistry (2019)
Human telomerase has been considered as a promising tumor marker for early cancer diagnosis and tumor progression monitoring. Current methods for detection of telomerase mainly rely on in vitro assays using cell lysate, which cannot provide information on telomerase activities in living systems. Only the few reported intracellular probes possess high telomerase selectivity but involve no signal amplification process, which potentially limits their use in application scenarios requiring high sensitivity. The development of an ultrasensitive intracellular telomerase probe is of high demand but challenging, because of the difficulty in designing a robust amplification process in living cells. Inspired by the mechanism of telomerase primer binding and extension, we introduce a cascade amplification reaction-based nanoprobe for intracellular telomerase detection by incorporating DNAzyme and catalytic hairpin assembly onto MnO2 nanosheets. The MnO2 nanosheets can deliver and release multicomponent signal amplification motifs with designed ratio at the same intracellular position, thereby enabling the cascade process in cells to occur. The released Mn2+ ions from degraded MnO2 nanosheets can activate DNAzyme as a metal cofactor and facilitate endosomal escape, because of the ion sponge effect. We used the nanoprobe to successfully monitor the dynamic change of telomerase activity in the HeLa cell, as well as in three other types of cells. This cascade amplification nanoprobe provides ultrasensitive detection of telomerase activity, indicating its use as a promising bioassay for early cancer diagnosis.
Keyphrases
- living cells
- fluorescent probe
- label free
- single molecule
- nucleic acid
- quantum dots
- gold nanoparticles
- induced apoptosis
- endothelial cells
- single cell
- cell cycle arrest
- cell proliferation
- loop mediated isothermal amplification
- papillary thyroid
- cell therapy
- squamous cell carcinoma
- signaling pathway
- high resolution
- small molecule
- highly efficient
- real time pcr