Real-time multiple particle tracking of gene nanocarriers in complex biological environments

Samuel K. Lai, Justin Hanes

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Complex biological fluids, such as the vast and molecularly crowded cell cytoplasm and the highly viscoelastic mucus that protects many entry ways to the body, pose significant barriers to efficient gene delivery. Understanding the dynamics of gene carriers in such environments allows insight that leads to rational improvements in gene vector design. Fluorescence techniques that provide only ensemble-averaged transport characteristics do not provide detailed information related to the nature of various barriers to efficient gene vector transport to target cell nuclei. Multiple particle tracking (MPT) allows the tracking of the real-time motion of up to hundreds of individual particles simultaneously with high temporal and spatial resolution. We have adapted MPT to study gene carrier transport in live cells and in fresh, undiluted human mucus. By analyzing the displacements of gene vectors as a function of time scale, this technique provides, on a per particle basis, highly quantitative measurements of the transport rates and transport mechanisms, as well as biophysical information of the complex biological environments. Combining MPT with confocal microscopy (confocal particle tracking) allows dynamic and quantitative co-localization determination of gene carriers with various cellular structures, such as endosomes, lysosomes, the endoplasmic reticulum, and Golgi. We have applied MPT to enhance understanding of critical extracellular and intracellular bottlenecks to gene transfer.

Original languageEnglish (US)
Title of host publicationGene Therapy Protocols
Subtitle of host publicationDesign and Characterization of Gene Transfer Vectors
PublisherHumana Press
Pages81-97
Number of pages17
ISBN (Print)9781603272476
DOIs
StatePublished - 2008

Publication series

NameMethods in Molecular Biology
Volume434
ISSN (Print)1064-3745

Keywords

  • Active transport
  • Diffusion
  • Drug delivery systems
  • Fluorescence
  • Intracellular transport
  • Mucus
  • Polyethylenimine
  • Polymers
  • Polyplexes

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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