Western Blot Evaluation of siRNA Delivery by pH-Responsive Peptides

  • Wanling Liang
  • A. James Mason
  • Jenny K. W. Lam
Protocol

Abstract

Gene silencing, via RNA interference (RNAi) technologies using small interfering RNA (siRNA), has been developed as an important tool for target identification and validation in drug discovery and has huge therapeutic potential. However, effective delivery into cells presents a major challenge to the use of siRNA. pH-responsive cell-penetrating peptides have attracted considerable attention in recent years as delivery vectors due to their ability to transport their cargos across the biological membrane and/or to promote endosomal escape and prevent lysosomal degradation. To evaluate the in vitro transfection efficiency of the pH-responsive peptide-based siRNA delivery system, the western blotting technique is commonly employed. This method offers a simple, efficient and economical way to study the gene silencing effect of the siRNA by analysing the protein of interest in a sample with minimum equipment requirement. This chapter provides a description of siRNA delivery and analysis by western blotting protocols for qualitatively and quantitatively assessing gene silencing efficiency and selectivity.

Key words

Transfection siRNA GAPDH Western blotting Densitometry pH-responsive peptide Chemiluminescence 

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Copyright information

© SpringerScience+Business Media New York 2013

Authors and Affiliations

  • Wanling Liang
    • 1
  • A. James Mason
    • 2
  • Jenny K. W. Lam
    • 1
  1. 1.Li Ka Shing Faculty of Medicine, Department of Pharmacology and PharmacyThe University of Hong KongPokfulamHong Kong
  2. 2.Institute of Pharmaceutical Science, King’s College LondonLondonUK

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