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Studying the uptake of cell-penetrating peptides

Nature Protocols volume 1, pages 1001–1005 (2006)Cite this article

Abstract

More than a decade ago, it was discovered that cationic peptides could traverse the cellular plasma membrane without specific transporter proteins or membrane damage. Subsequently, it was found that these peptides, known as cell-penetrating peptides (CPPs), were also capable of delivering cargos into cells, hence the great potential of these vectors was acknowledged. Today, many different research groups are working with CPPs, which necessitates efforts to develop unified assays enabling the comparison of data. Here we contribute three protocols for evaluation of CPPs which, if used in conjunction, provide complementary data about the amount and mechanism of uptake (fluorometric analysis and confocal microscopy, respectively), as well as the extent of degradation (HPLC analysis of cell lysates). All three protocols are based on the use of fluorescently labeled peptides and can be performed on the same workday.

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Figure 1: Quantitative uptake of the cell-penetrating peptides TP10 and pVEC in HeLa cells after treatment with 5 μM peptide for 1 h.
Figure 2: Detection of cell-penetrating peptides in living cells by confocal laser scanning microscopy.
Figure 3: Uptake of the cell-penetrating peptide (KFF)3K in Saccharomyces cerevisiae.

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Authors and Affiliations

  1. Department of Neurochemistry, Stockholm University, S. Arrheniusv. 21A, Stockholm, SE-106 91, Sweden

    Tina Holm, Henrik Johansson, Pontus Lundberg, Maria Lindgren & Ülo Langel

  2. Estonian Biocentre, Riia 23, Tartu, EE2400, Estonia

    Margus Pooga

Authors
  1. Tina Holm
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  2. Henrik Johansson
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  3. Pontus Lundberg
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  4. Margus Pooga
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  5. Maria Lindgren
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  6. Ülo Langel
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Correspondence to Ülo Langel.

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The authors declare no competing financial interests.

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Holm, T., Johansson, H., Lundberg, P. et al. Studying the uptake of cell-penetrating peptides. Nat Protoc 1, 1001–1005 (2006). https://doi.org/10.1038/nprot.2006.174

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