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Delivery of siRNA from lyophilized polymeric surfaces

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Standard in vitro gene silencing protocols are performed using aqueous formulations of transfection reagents and small interfering RNAs (siRNA) reconstituted immediately prior to use. In this study, we describe a method for producing gene silencing-active lyophilized cationic polymer (chitosan) or lipid (TransIT-TKO) siRNA formulations. We demonstrate specific and efficient knockdown of enhanced green fluorescent protein (EGFP) in H1299 human lung carcinoma cells transfected in plates pre-coated with both TransIT-TKO/siRNA ( approximately 85%) and a chitosan/siRNA formulation containing sucrose as lyoprotectant ( approximately 70%). This method removes the necessity for both siRNA reconstitution immediately prior to use and addition onto cells. Furthermore, silencing activity of the chitosan/siRNA formulation was shown over the period studied ( approximately 2 months) when stored at room temperature. Higher cell viability was observed using the chitosan system compared to the lipid formulation. Silencing of the proinflammatory cytokine tumour necrosis factor (TNF-alpha) was also demonstrated in the RAW macrophage cell line using the lyophilized chitosan/siRNA system suggesting that the coating can improve the biocompatibility of medical implants. This work describes an efficient gene silencing methodology using freeze-dried formulations with potential applications as a high throughput screening tool for gene function, biocompatible medical implant components and longer shelf-life therapeutics.
Original languageEnglish
Pages (from-to)506-512
Number of pages7
Publication statusPublished - 2008

    Research areas

  • Animals, Cell Line, Chitosan, Green Fluorescent Proteins, Humans, Lipids, Macrophages, Mice, Nanoparticles, RNA, Small Interfering, Surface Properties, Tumor Necrosis Factor-alpha, Water

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