Ultrasensitive detection of telomerase activity in lung cancer cells with quencher-free molecular beacon-assisted quadratic signal amplification.

Abstract Telomerase is an important biomarker for cancer diagnosis and a valuable target for cancer therapy. Most of the reported telomerase assays suffer from the repeating thermal cycles, laborious protocols, expensive instruments and the limited sensitivity. To solve these issues, we develop a new telomerase assay based on telomerization-driven release of fluorescent 2-aminopurine. We designed a 2-aminopurine molecular beacon in which the fluorescence of 2-aminopurine is quenched due to stacking interaction effect without the use of extra quenchers. The presence of target telomerase can open the 2-aminopurine molecular beacon, triggering a quadratic signal amplification reaction to digest abundant 2-aminopurine molecular beacons, releasing large amounts of free 2-aminopurine molecules with strong fluorescence emission. Due to the inherent low background signal of 2-aminopurine molecular beacon and the highly efficient telomerization-driven quadratic signal amplification, this method exhibits high sensitivity with a limit of detection equivalent to 1 human lung cancer A549 cell and a large dynamic range of 4 orders of magnitude from 1 to 10000 cells. Moreover, this method can be further applied for telomerase inhibition assay and the discrimination of cancer cells from normal cells, holding great potential in telomerase-related clinical diagnosis and drug discovery. Graphical abstract We develop a new method for ultrasensitive detection of telomerase activity in lung cancer cells based on telomerization-driven release of fluorescent 2-aminopurine. Image 1 Highlights • We develop a new fluorescent method for ultrasensitive detection of telomerase activity. • This assay can be performed under isothermal condition without the involvement of any extra quenchers. • This method can measure endogenous telomerase activity equivalent to 1 lung cancer cell. • This method can discriminate cancer cells from normal cells. [ABSTRACT FROM AUTHOR]