Ratiometric fluorescence resonance energy transfer for reliable and sensitive detection of intracellular telomerase RNA via strand displacement reaction amplification

Human telomerase RNA (hTR) is one essential component of telomerase and is overexpressed in tumor cells. Therefore, the reliable and sensitive detection of hTR is essential for the early cancer diagnosis. Herein, to avoid the false positive signals caused by co-existing components in the cell, a ratiometric fluorescence resonance energy transfer (FRET) strategy was developed to achieve reliable detection of intracellular hTR. Manganese dioxide nanosheets (MnO2NS) with good biocompatibility carry two fluorophore-labelled hairpin DNA probes into the cancer cell and then release the probes via decomposition of MnO2NS by intracellular L-glutathione reduced (GSH). Then, hTR triggered the cyclic strand displacement reaction (SDR) between two hairpin DNA probes to continuously form DNA duplexes, which made two fluorophores close to each other and led to an effective FRET. Fluorescence imaging demonstrated a higher expression level of hTR in HeLa cells than that in normal HL-7702 cells. The high specificity of hairpin DNA probes and SDR make it easy to discriminate the single-base mutation. Therefore, it provides a highly sensitive, simple and reliable method for the extracellular and intracellular detection of hTR. (C) 2022 Elsevier B.V. All rights reserved.

Automated summary

In this study, a reliable and sensitive method for the detection of intracellular hTR was developed using a ratiometric fluorescence resonance energy transfer (FRET) strategy. The method utilized manganese dioxide nanosheets and hairpin DNA probes to achieve FRET-based detection. The method exhibited high specificity and the ability to discriminate single-base mutations.