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目的:分析转移性大肠癌LoVo细胞特异性核酸适配子W14的生物活性和血浆稳定性。方法:采用荧光显微镜流式细胞术分析W14在25℃和37℃下与靶细胞LoVo的特异结合活性;流式细胞术检测W14在血浆环境下对LoVo细胞的特异识别能力;37℃下W14分别在完全培养基和人血浆中孵育0.5、1、1.5、2、3、4、6 h,琼脂糖凝胶电泳检测W14的生物稳定性;用2、4、8、10μmol/L的W14处理LoVo细胞24、48、72 h,MTS实验测定W14的细胞毒性。结果:W14在不同温度和血浆环境中以及不同温度条件下都能特异性地识别和结合靶细胞LoVo,能稳定地存在于血浆中6 h不发生降解,且对细胞没有明显的毒性作用。结论:转移性大肠癌特异性核酸适配子W14具有良好的适应能力和特异性结合能力,有较好的生物稳定性和低毒性,适应于人体内应用。
Objective: To analyze the biological activity and plasma stability of LoVo cell-specific aptamer W14 in metastatic colorectal cancer. Methods: The specific binding activity of W14 to LoVo at 25 ℃ and 37 ℃ was analyzed by fluorescence microscope. The specific recognition of W14 in LoVo cells was detected by flow cytometry. 0.5, 1, 1.5, 2, 3, 4 and 6 h in complete medium and human plasma. The biological stability of W14 was detected by agarose gel electrophoresis. LoVo was treated with 2,4,8 and 10 μmol / L of W14 Cells 24, 48, 72 h, MTS assay determined the cytotoxicity of W14. Results: W14 could specifically recognize and bind to target cell LoVo under different temperature and plasma conditions and under different temperature conditions. It could stably exist in plasma for 6 h without degradation and had no obvious cytotoxic effect on cells. Conclusion: The specific nucleic acid aptamer W14 of metastatic colorectal cancer has good adaptability and specific binding ability, has good biological stability and low toxicity and is suitable for human application.