Detection Of Breast Cancer And Related Molecules By Fluorescence-labled Functional Nucleic Acids

Breast cancer is one of the most commonly diagnosed cancers among females worldwide. Early detection of breast cancer is of vital importance to the reduction of the mortality rate. However, the lack of specific biomarkers that can effectively identify breast cancer cells limits the ability for early diagnosis of breast cancer. RNA-cleaving fluorogenic DNAzymes(RFDs), which can be produced through the Systematic Evolution of Ligands by Exponential enrichment(SELEX) process, are catalytic DNA molecules capable of generating a fluorescent signal when the appropriate target is bound. In this study, new methods were established for breast cancer and related molecules detection by applying functional nucleic acids. The results are as follows:A SELEX experiment was carried out to select for RFDs that are active in the cell lysate of MDA-MB-231, a model breast cancer cell line. We obtained a RFD probe, named AAI2-5, that can detect MDA-MB-231 at a concentration of cell lysate proteins as low as 0.5 μg/mL(which is equivalent to ~5,000 cell/m L). AAI2-5 is capable of distinguishing MDA-MB-231 cells from normal cells as well as other types of tumor cells, including other subtypes of breast cancer cells. Moreover, AAI2-5 responded positively to more than 90% of breast tumors. This report is the first study to explore the RFD system for the detection of cancer cells. The results suggest that RFD can be potentially applied for the diagnosis and treatment of breast cancer in the future.ER-3, the first RFD probe selected for targeting protein, was obtained through SELEX procedure for specific detection of eatrogen receptor(ER). Its performance was improved by a series of optimization experiments, e.g. different ions, concentrations of the ion, pH, temperature. The probe could detect ER protein at the concentration of 0.8 μg/m L. It is expected that ER-3 would be applied for clinical diagnosis of breast cancer as well as distinguishing different sybtypes of breast cancer by advanced research.Highly sensitive ATP detection assay was demonstrated based on a fluorescent aptameric probe and graphene oxide. Improved sensitivity was achieved by the careful design of the allosteric probe and exonuclease III digestion product-triggered signal amplification. This strategy was lower than that of the previous unamplified GO-based fluorescence method and comparable or better than previously reported aptamer-based amplified fluorescent sensors for ATP detection. The method with highly specificity was confirmed barely influenced by the proteins, which could be used to detect ATP in real biological samples.