A Novel Assay Of Rare Disseminated Tumor Cells In Real Sample By Aptamer-induced Rolling Circle Amplification On Cell Surface

The identification of tumor cells disseminating in body fluids as a new noninvasive way of liquid biopsy plays a critical role in the early diagnosis and treatment guidance,tumor recurrence and metastasis,clinical assessment,individualized treatment and prognosis assessment of cancer.In addition,an invasive procedure limited to such patients who are unable to endure the risks associated with biopsy examination,especially in those tumors where tissue is not easy to obtain,such as breast cancer,lung cancer,colorectal cancer.At present,the biggest challenge for the detection of tumor cell detection is that its contents in peripheral blood and body fluids are very low,and the existing methods can not achieve sensitive detection.Herein,a simple and pragmatic colorimetric strategy has been developed for ultrasensitive detection of tumor cells based on aptamer-induced rolling circle amplification(RCA)on cell surface for identification of tumor cells disseminating in body fluids.In this study,breast carcinoma MCF-7 cells were used as a model,and a designed Epithelial Cell Adhesion Molecule(EpCAM)-primer probe,while targeted aptamer was selected as recognition agent to specifically recognize and bind with tumor cells.Following,the aptamer-induced RCA performed in situ,presenting numerous G-quadruplexes sequences on the cell surface.G-quadruplex/hemin DNAzyme then formed and catalyzed colorimetric reaction for tumor cells detection.The designed colorimetric assay could distinguish as low as 10 cancer cells in 105 of benign cells mixture,which was equal to 0.01%of MCF-7 cells in 105 cells mixture.The proposed colorimetric assay was further applied to directly analyze 10 clinical body fluids specimens including 5 cases of benign pleural fluids and 5 cases of malignant body fluids,showing a high specificity and great capacity in clinical application.Thus this proposed strategy successfully realized the high sensitive detection of low abundance tumor cells in body fluid samples and provided a simple,handy and powerful platform for ultrasensitive detection of tumor cells and had great potential for clinical application in convenient assay of rare disseminated tumor cells in body fluid samples for early cancer screening.