Davis Pamela B, Kowalczyk Tomasz H
Cold Spring Harb Protoc. 2010 May;2010(5):pdb.prot5419. doi: 10.1101/pdb.prot5419.
PEGylated poly-L-lysine DNA nanoparticles are composed of plasmid DNA compacted with poly-L-lysine conjugated with polyethylene glycol (PEG). They are soluble and stable in saline and tissue fluids, transfect nondividing cells, display minimal toxicity, and are effective in vivo and in humans. Moreover, they are easy to prepare in a reliable and reproducible fashion. These properties represent a substantial advance for nonviral gene transfer. This article describes the conjugation of methoxy-PEG-maleimide with the peptide CK(30) and the compaction of DNA with the resultant PEGylated polylysine. It also describes the analyses used to check the morphology and colloidal stability of the nanoparticles. These assays should be performed each time the nanoparticles are prepared because, although the compaction procedure is very reproducible, variations in product quality do sometimes occur (e.g., the particles are unstable or have an unacceptable morphology). Variations seem to happen most often when the source of plasmid or method of plasmid production is changed.
聚乙二醇化聚-L-赖氨酸DNA纳米颗粒由与聚乙二醇（PEG）共轭的聚-L-赖氨酸压缩的质粒DNA组成。它们在盐水和组织液中可溶且稳定，能转染非分裂细胞，毒性极小，在体内和人体中均有效。此外，它们易于以可靠且可重复的方式制备。这些特性代表了非病毒基因传递的重大进展。本文描述了甲氧基-PEG-马来酰亚胺与肽CK(30)的共轭以及所得聚乙二醇化聚赖氨酸对DNA的压缩。还描述了用于检查纳米颗粒形态和胶体稳定性的分析方法。每次制备纳米颗粒时都应进行这些测定，因为尽管压缩过程具有很高的可重复性，但产品质量有时仍会出现变化（例如，颗粒不稳定或形态不可接受）。当质粒来源或质粒生产方法改变时，变化似乎最常发生。
