Preparation of Biodegradable Nanoparticles and Their Use in Transfection
This protocol was adapted from “Biodegradable Nanoparticles,” Chapter 56, in Gene Transfer: Delivery and Expression of DNA and RNA (eds. Friedmann and Rossi). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2007.INTRODUCTION
Biodegradable nanoparticles (NPs) are colloidal particles with a gene of interest encapsulated inside a polymeric matrix. They are typically 100 nm in diameter, and are formulated using FDA-approved, biodegradable, biocompatible polymers such as poly(D,L-lactide-co-glycolide) (PLGA) or polylactide (PLA). The NPs are taken up by cells via an endocytic process, and the encapsulated plasmid DNA entrapped in NPs is protected from degradation by both extra- and intracellular nucleases. It is released slowly, sustaining gene delivery and gene expression. In contrast, higher but transient gene expression is observed with lipid- or polymer-based complexes where most of the delivered DNA is available quickly for transfection. Thus, unlike other nonviral gene delivery systems, NPs thus constitute a sustained gene expression vector. Sustained gene expression is advantageous, especially when the half-life of the expressed protein is very low or when chronic gene delivery is required for therapeutic efficacy. This protocol describes a method for nanoencapsulation of DNA and the subsequent use of NPs for transfection.
