Optimized preparation of DNA-chitosan nanoparticles with high transfection effi-cency through a central composition design
10.3969/j.issn.1006-0111.2014.06.006
- VernacularTitle:中心组合设计法优化载基因壳聚糖纳米粒的最佳转染制备区域
- Author:
Shuai SHAO
1
;
Guanghua CUI
;
Xu ZHOU
;
Zhonggao GAO
;
Wei HUANG
Author Information
1. 中国医学科学院、北京协和医学院药物研究所药物制剂研究室
- Keywords:
nanoparticles;
plasmid DNA;
central composition design;
chitosan;
transfection efficency
- From:
Journal of Pharmaceutical Practice
2014;(6):419-424
- CountryChina
- Language:Chinese
-
Abstract:
Objective This study aimed to optimize the preparation condition of DNA-chitosan nanoparticles with high transfec-tion efficency through a central composition design .Methods The DNA-chitosan nanoparticles were prepared by complex coacervation between pEGFP and chitosan .We selected the concentrations of chitosan and plasmid as two experimental factors , and a central compos-ite design with two factors and five levels was used to optimize the preparation condition of DNA-chitosan nanoparticles for high transfec-tion efficency .The concentrations of chitosan and plasmid were selected as the independent variables , respectively .The dependent varia-bles included average particle size and transfection efficiency .The morphology of DNA-chitosan nanoparticles was observed using a trans-mission electron microscope .The size and zeta potential of nanoparticles were measured by dynamic light scattering ( DLS) and electro-phoretic light scattering ( ELS ) , respectively .The stability of plasmids in the process of nanoparticles preparation was investigated through the agrose gel electrophoresis .The expression of plasmids delivered by nanoparticles was observed under an inverted fluorescence microscope .The transfection efficiency of DNA-chitosan nanoparticles was assayed by flow cytometry .Results The preparation condition of DNA-chitosan nanoparticles with high transfection efficency was optimized successfully .Under the optimum preparative conditions , the DNA-chitosan nanoparticles were almost spherical .The average size of nanoparticles was 217.6nm, and distributed in a narrow range with a polydispersity index of 0.241.The zeta potential was +22.4 mV, which suggested that a den-sity of positive charge exist onto the surface of nanoparticles and consequently enhanced the stability of nanoparticles suspension .The results of gel electrophoresis showed that plasmids were not destroyed in the process of nanoparticles preparation .The cell transfection of nanoparticles was very highly efficient .The nanoparticles could effectively deliver the pEGFP plasmids into cells to express the green fluorescent protein at a high level.Conclusion The established mathematic models have the good predictive function .Under the optimum preparative condi-tions, the DNA-chitosan nanoparticles have the high potential of cell transfection .
