Design and fabrication of a fluorescence probe for simple, rapid and sensitive detection of telomerase activity as a selective cancer biomarker

Human telomerase is a ribonucleoprotein reverse transcriptasethat catalyzes the addition of the telomeric repeats(TTAGGG)n, onto the end of the humanchromosomes. Even though the vast majority of human cancers express telomerase activity, most human somatic cells lackthe telomerase activity. This rendersthe enzyme a valuable biomarker for cancer diagnosis and an important therapeutictarget. Up to date, the polymerase chain reaction (PCR)-based telomeric repeat amplification protocol (TRAP) is themost widely used because of its high sensitivity.However,TRAPrequires the use of DNA polymerases and is thereforesusceptible to PCR-derived artifacts.Recently, a number of PCRfreeassays have beendeveloped for telomerase activity based on DNAzymes,electrochemical sensors,and nanoparticles. However, each of them still has its shortcomings including low sensitivity, complicated manipulation and time-consuming procedures, or the need for elaborate instruments. In this study, a simple, fast, sensitive and homogeneous off-on fluorometric method is developed for the determination of telomerase activity. For this purpose, the telomeric sequence was labeled with graphene quantum dots (GQDs) upon which the graphene oxide quenched the fluorescence of the GQDs. Addition of the cell extracts containing telomerase will results in the elongation of the telomeric sequence, release of GQDs and regeneration of fluorescence signal. The proposed method allowed the measurement of telomeraseactivity in crude cell extracts equivalent to 5 HeLa cells with a dynamic range of 10-10000 cells. Telomerase activities ofdifferent cell lines were also successfully evaluated

کلیدواژه ها:

Telomerase ، Fluorescence ، Forster resonance energy transfer (FRET) ، Graphene quantum dots ، Graphene oxide