Aptamer-based Catalyzed Signal Amplification Method For Detection Of Cancer Cells

Cancer was a serious threat to human life and health. The focus of the currentfield of medical research to be solved was to explore new methods of early diagnosisto achieve patient cure rate and survival improved. Aptamer provides a possible toolfor early diagnosis of tumors as a novel probe of molecular recognition.Catalytic-based amplification methods, such as a horseradish peroxidase (HRP)mimicking DNAzyme and nano-gold particles catalyzed silver enhancement had beensuccessfully applied to the detection of biological molecules. In this thesis, DNAzymeand silver enhanced signal amplification technology were designed to detect leukemiacells (CCRF-CEM) and human non-small cell lung cancer cells (A549). Several workshad been mainly performed as following:1DNAzyme-based colorimetric signal amplification detection of CCRF-CEMBased on SP2.M forming a G-quadruplex structure which can catalyze theoxidation of ABTS2-by H2O2and Sgc8c were able to specifically bind to CCRF-CEMcells, a signal amplification colorimetric method for detection of human leukemiacells were developed. The catalytic product of the ABTS-absorbance changes at418nm were measured to the specific and quantitative detection of CCRF-CEM cells，the detection limit of1000cells/mL.2.DNAzyme-based colorimetric signal amplification detection of A549Based on the first part of the work, the A549adherent cells were in applied to thedetection method. The method could equally well be applied to the detection of theA549. Human cervical cancer cell line (HeLa), human normal liver cell line (L-02),human breast cancer cell line (MCF-07) were selected as control cells or sequence. Ithas a good cell-specific and sequence-specific, detection limit was3000cells/mL.The results showed that cell biological molecules activity was reduced recoveryresulting from digestion after shock culture.It was indicating that this assay had thepotential to differentiate between different types of target and control cells based onthe aptamers.3Silver-enhanced gold nanoparticles assay for A549cell detectionCombination with cell-specific nucleic acid aptamer S6and the silver enhancedcatalyzed by gold nanoparticles assay, A549cells were detected. It was quantitativedetection of A549cells by Gray value. The results showed that A549cells had goodeffects but control cells (human tongue cancer (TCA), human breast cancer cell line (MCF-07) and human cervical cancer cells (HeLa)) imaging signal was low. Thesignal of the target cells and control cells were very different obvious and thedetection limit was7000cells/mL. The method was convenient, simple operation andexpected to be developed into one of the universal detection of adherent cells, evenfurther applied to the tumor tissue biopsy up.