Skip to main content
SpringerLink
Log in
Book cover

Liposomes pp 439–455Cite as

FRET Imaging of Cells Transfected with siRNA/Liposome Complexes

  • Protocol
  • First Online:

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 606))

Abstract

By monitoring the efficiency of fluorescence resonance energy transfer of dyes attached to the different strands of siRNA, the structural integrity of the latter can be traced inside cells. Here, the experimental details of dye-labeled siRNA construction, tissue culture, and transfection with liposomally formulated siRNAs are given, as well as the conditions for confocal microscopy and an algorithm allowing the visualization of intact siRNA after image data treatment. The method allows rapid screening of different liposomal siRNA formulations, obtained by small scale dual asymmetric centrifugation with high entrapping efficiency.

This is a preview of subscription content, log in via an institution.

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Järve A, Müller J, Kim IH, Rohr K, MacLean C, Fricker G, Massing U, Eberle F, Dalpke A, Fischer R, Trendelenburg MF, Helm M (2007) Surveillance of siRNA integrity by FRET imaging. Nucleic Acids Res 35:e124

    Article  PubMed  Google Scholar 

  2. Zimmermann TS, Lee AC, Akinc A, Bramlage B, Bumcrot D, Fedoruk MN, Harborth J, Heyes JA, Jeffs LB, John M, Judge AD, Lam K, McClintock K, Nechev LV, Palmer LR, Racie T, Rohl I, Seiffert S, Shanmugam S, Sood V, Soutschek J, Toudjarska I, Wheat AJ, Yaworski E, Zedalis W, Koteliansky V, Manoharan M, Vornlocher HP, MacLachlan I (2006) RNAi-mediated gene silencing in non-human primates. Nature 441:111-114

    Article  CAS  PubMed  Google Scholar 

  3. Massing U, Cicko S, Ziroli V (2008) Dual asymmetric centrifugation (DAC)-a new technique for liposome preparation. J Control Release 125:16-24

    Article  CAS  PubMed  Google Scholar 

  4. Hirsch M, Ziroli V, Helm M, Massing U (2009) Preparation of small amounts of sterile siRNA-liposomes with high entrapping ­efficiency by dual asymmetric centrifugation (DAC). J Control Release 135:80-88

    Article  CAS  PubMed  Google Scholar 

  5. Elbashir SM, Harborth J, Weber K, Tuschl T (2002) Analysis of gene function in somatic mammalian cells using small interfering RNAs. Methods 26:199-213

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

M.H. gratefully acknowledges the Heidelberg Rotary Club for funding, Heike Einberger for technical assistance, and the Nikon Imaging Center at the University of Heidelberg. We thank Andres Jäschke for generous support.

Author information

Authors and Affiliations

  1. Department of Bioinformatics and Functional Genomics, German Cancer Research Center (DKFZ), Institute of Pharmacy and Molecular Biotechnology, University of Heidelberg, Heidelberg, Germany

    Il-Han Kim & Karl Rohr

  2. Department of Chemistry, Institute of Pharmacy and Molecular Biotechnology, University of Heidelberg, Heidelberg, Germany

    Anne Järve, Markus Hirsch & Mark Helm

  3. German Cancer Research Center (DKFZ), Heidelberg, Germany

    Roger Fischer & Michael F. Trendelenburg

  4. Department of Clinical Research, Tumor Biology Center, Freiburg, Germany

    Ulrich Massing

Authors
  1. Il-Han Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Anne Järve
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Markus Hirsch
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Roger Fischer
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Michael F. Trendelenburg
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ulrich Massing
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Karl Rohr
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Mark Helm
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Mark Helm .

Editor information

Editors and Affiliations

  1. College of Pharmacy, Midwestern University, N. 59th Avenue 19555, Glendale, 85308, U.S.A.

    Volkmar Weissig

Rights and permissions

Reprints and permissions

Copyright information

© 2010 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Kim, IH. et al. (2010). FRET Imaging of Cells Transfected with siRNA/Liposome Complexes. In: Weissig, V. (eds) Liposomes. Methods in Molecular Biology™, vol 606. Humana Press. https://doi.org/10.1007/978-1-60761-447-0_30

Download citation

  • DOI: https://doi.org/10.1007/978-1-60761-447-0_30

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-60761-446-3

  • Online ISBN: 978-1-60761-447-0

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation