Quantum Dots And Functionalized Gold Nanoprobes For Telomerase Activity Analysis

Cancer,also known as malignant tumor,is one of the most important diseases that seriously threaten human health and life.At present,the number of people who die of cancer every year in China reaches nearly one million.The cure rate of skin cancer and cervical cancer with early diagnosis is very high,and if it can be controlled in the early stage,the cure rate may be close to 100%.Small amount of cancer markers will be released into the blood in the early stage,where telomerase is one of the important cancer markers.In normal somatic cells,telomerase activity is repressed or absent.However,the telomerase activity is activated and over-expressed in over 85% of cancer cells.Therefore,the determination of telomerase activity is of great significance for the early diagnosis of human cancers,the study of tumors development,and the screening of anticancer-related drugs.The concentration of telomerase is extremely low in the early stage of cancer,and the current methods of detecting telomerase have the disadvantages such as complicated operation steps and strong background signals,which can not achieve the purpose of early diagnosis of cancer.Concerning the existing problems of detecting telomerase activity in the previous works,in order to achieve the purpose of detecting telomerase activity with high sensitivity,simple operation and low background signal,quantum dots and noble metal nanomaterials were introduced into the analysis system,and the fluorescence analysis methods for detecting telomerase activity were established by using the excellent optical properties of nanomaterials.The main points of our work in this paper are summarized as follows:In Chapter 2,based on the controlled quantum dot aggregation,a new method has been proposed for the detection of telomerase activity with high sensitivity and good selectivity.In the presence of telomerase and biotin-dATP,the extension products contain a large amount of biotin,quantum dots streptavidin conjugates(QDs-SA)will bind to the telomerase elongation products due to the high affinity between the biotin and streptavidin,protecting the QDs-SA from aggregation and retaining their strong fluorescence.In the absence of telomerase,the aggregation and thus the fluorescence quenching of QDs-SA can be observed in buffer solution due to charge shielding effect.The telomerase activity can be detected simply and reliably by the evolution of the QDs-SA fluorescence signal,and the telomerase activity in the cells can also be imaged and analyzed.In chapter 3,we develop a novel method for sensitive detection of telomerase activity by designing a gold nanoparticles/graphene oxide(AuNPs/GO)probe.The AuNPs were functionalized with a telomerase substrate(TS)primer and a FAM-modified complementary DNA(P1).In the absence of telomerase,P1 exists in the random-coiled conformation,and the fluorescence resonance energy transfer(FRET)from FAM to AuNPs and GO results in efficient fluorescence quenching.In the presence of telomerase,the multiple hybridization between TS extension products and P1 leads to the conformation transition of P1 from single-stranded DNA to double-stranded rigid structure.The fluorescent dye(FAM)is far away from AuNPs and GO surface that can prevent the FRET process with the efficient fluorescence recovery.The metal-enhanced fluorescence(MEF)can further enhance the fluorescence signal by changing the length of the P1.It is worth mentioning that the proposed strategy does not need to design complex hairpin structure and allows the measurement of telomerase activity in crude cell extracts within 1 hour.In addition,the present sensing platform can be applied to inhibitor screening,in situ telomerase imaging,and intracellular drug delivery.