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目的建立肿瘤基因疫苗的纯化工艺,为肿瘤基因疫苗的工业化生产奠定基础。方法采用碱裂解法粗提质粒DNA,利用凝胶过滤层析、亲和层析及离子交换层析分离纯化质粒DNA,并对所纯化的质粒DNA进行全面检定。结果经3种层析柱纯化所获得的超螺旋质粒DNA,其A260与A280的比值在1.85～1.92之间;超螺旋质粒DNA比例不低于94%;内毒素含量远低于10EU/mg;5批纯化的质粒DNA经全面检定,均符合国家质量标准。结论该纯化工艺制备的肿瘤基因疫苗纯度高,且有效去除了内毒素。
Objective To establish a purification process of tumor gene vaccines and lay a foundation for industrialized production of tumor gene vaccines. Methods Plasmid DNA was crudely extracted by alkaline lysis method, and the plasmid DNA was purified by gel filtration chromatography, affinity chromatography and ion exchange chromatography. The purified plasmid DNA was tested comprehensively. Results The ratio of A260 to A280 was between 1.85 and 1.92. The ratio of supercoiled plasmid DNA was not less than 94%. The content of endotoxin was much lower than 10EU / mg. 5 batches of purified plasmid DNA by a comprehensive test, are in line with national quality standards. Conclusion The tumor gene vaccine prepared by the purification process has high purity and effectively removes endotoxin.