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.