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目的:探讨聚乙烯亚胺(polyethylene im ine,PEI)作为生存素(survivin)反义核酸(anti-sense oligodeoxynuc leotides,ASODN)的载体投递肝癌SMMC-7721细胞的条件。方法:凝胶阻滞实验筛选PEI与survivin ASODN形成静电复合物的最佳质量比;W ST-8法检测PEI对肝癌细胞的毒性;流式细胞仪及W ST-8法筛选荧光标记的PEI与ASODN在不同质量比时的瞬时转染效率及48 h转染效率;流式细胞仪及荧光显微镜分别检测荧光标记的ASODN以及PEI-ASODN复合物进入细胞的情况;W ST-8法检测血清对PEI转染效率的影响。结果:m(PEI)∶m(ASODN)为0.625∶1~2.5∶1时可以形成电中性状态的静电复合物;PEI终质量浓度≤4μg/mL对肝癌细胞的毒性较小;m(PEI)∶m(ASODN)为0.75∶1时转染效率最高;血清存在对PEI转染效率无明显影响。结论:PEI是一种低毒的ASODN投递载体,其终质量浓度≤4μg/mL可作为肝癌细胞的转染载体,血清条件下m(PEI)∶m(ASODN)为0.75∶1时,PEI携带survivin ASODN投递肝癌SMMC-7721细胞的效率最高。
OBJECTIVE: To investigate the conditions of delivering polyethylene imine (PEI) as a carrier of survivin anti-sense oligodeoxynucleases (SMOD-7721). METHODS: The best mass ratio of PEI to survivin ASODN was screened by gel blocking assay. The toxicity of PEI to hepatocarcinoma cells was detected by WST-8 method. Fluorescence-labeled PEI was screened by flow cytometry and WST-8 method And transfection efficiency of ASODN at different mass ratios and transfection efficiency at 48 h were detected by flow cytometry and fluorescence microscope respectively.WS-8 method was used to detect the expression of ASODN and PEI-ASODN complex Effect on PEI transfection efficiency. Results: The electroneutral complex could be formed in the condition of m (PEI): m (ASODN) of 0.625:1 ~ 2.5:1. The PEI final concentration of 4μg / mL was less toxic to liver cancer cells. ): M (ASODN) was 0.75:1, the transfection efficiency was the highest; presence of serum had no significant effect on the transfection efficiency of PEI. Conclusion: PEI is a low-toxic ASODN delivery vector. The final concentration of PEI is less than 4μg / mL, which can be used as a transfection vector for liver cancer cells. When PEI: m (ASODN) is 0.75:1, Survivin ASODN delivers the highest efficiency of hepatoma SMMC-7721 cells.