Preparation of DNA-loaded chitosan nanoparticle vaccine
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摘要: 目的 制备负载抗龋DNA疫苗pVAX1-wapA质粒的壳聚糖和季铵化壳聚糖纳米粒,优化其制备工艺,测定其细胞转染效率。 方法 以包封率和粒径为主要指标,单因素法考察载体浓度、pH值、N/P、TPP浓度等因素的影响,Realtime-PCR检测细胞对质粒编码蛋白的转录表达水平以评价载质粒纳米粒的促转染作用。 结果 制得的载DNA疫苗纳米粒粒径均一,形态圆整。壳聚糖(CS)纳米粒粒径为(219.2±18.2) nm,Zeta电位为(24.7±3.5) mV,包封率为91.24%。季铵化壳聚糖(CSTM)纳米粒粒径为(222.5±15.6) nm,Zeta电位为(19.6±1.2) mV,包封率为87.66%。纳米粒可以促进pVAX1-wapA进入细胞,并成功被转录。 结论 制备的包载pVAX1-wapA的季铵化壳聚糖纳米粒可用于重组基因疫苗的运送。Abstract: Objective To study and optimize the preparation condition of pVAX1-wapA-loaded nanoparticles and determine the transfection efficiency. Methods The related effects of the crucial factors for the formation of nanoparticles: concentration of chitosan and TPP, pH value, N/P ratio were studied by single-factor experiment, with nanoparticles size and zeta potential as index. Cell transfection test was carried out to indicate that enhancement of cell transfection efficiency of nano-carrier. Results Nanoparticles loaded DNA vaccine were nearly spherical shape with uniform particle size chitosan nanoparticle(CS),(219.2±18.2) nm;quaternary ammonium chitosan nanoparticles(CSTM),(222.5±15.6) nm. Zeta potential of CS and CSTM was (24.7±3.5) mV, (19.6±1.2) mV and encapsulation efficiency was 91.24%, 87.66%,respectively.CSTM nanoparticle could enhance cellular uptake of pVAX1-wapA obviously. Conclusion CSTM nanoparticle was proved to be an efficient DNA vaccine delivery vector.
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Key words:
- DNA vaccine /
- chitosan nanoparticle /
- wapA protein /
- transfection /
- quaternary ammonium chitosan
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