Highly sensitive monitoring of telomerase activity in living cells based on rapidly triggered cascade amplification reaction

Abstract

Telomerase is an important potential biomarker for the study of tumor progression. Herein, we designed a cascade-amplification-reaction-based nanoprobe for intracellular telomerase detection based on the integration of rolling circle amplification (RCA) and catalytic hairpin assembly (CHA) onto MnO2 nanosheets. Firstly, MnO2 nanosheets rapidly delivered and released signal amplification units into cells, and very short telomerase extension products formed RCA circular templates and initiated the exponential RCA, producing enriched telomere sequence amplification products. Then the amplification products specifically triggered the CHA process and numerous H1/H2 complexes were formed, realizing the exponential amplification of fluorescence signals. The detection limit is as low as 1 LoVo cell for telomerase activity in cell extract. We further designed a microfluidic chip with six independent cell culture regions for in situ fluorescence imaging. Simultaneous detection of six types of cells was realized on the chip, and only 1–2 μL of cell suspension and reagents are needed. Our detection method features faster response speed and stronger fluorescence signal. Telomerase in living cells showed strong fluorescence signal within 1.5 h, and tumor cells were effectively distinguished from normal cells. Telomerase activities of different types of tumor cells and activity changes were both monitored conveniently. These results demonstrate that this method holds the potential for the sensitive detection of low abundance biomarkers in living cells, and will contribute to cancer diagnosis, cancer treatment and telomerase-related drug screening.