Precise quantification of cellular uptake of cell-penetrating peptides using fluorescence-activated cell sorting and fluorescence correlation spectroscopy

Rachid Rezgui, Katy Blumer, Gilbert Yeoh-Tan, Adam J. Trexler, Mazin Magzoub

Research output: Contribution to journalArticle


Cell-penetrating peptides (CPPs) have emerged as a potentially powerful tool for drug delivery due to their ability to efficiently transport a whole host of biologically active cargoes into cells. Although concerted efforts have shed some light on the cellular internalization pathways of CPPs, quantification of CPP uptake has proved problematic. Here we describe an experimental approach that combines two powerful biophysical techniques, fluorescence-activated cell sorting (FACS) and fluorescence correlation spectroscopy (FCS), to directly, accurately and precisely measure the cellular uptake of fluorescently-labeled molecules. This rapid and technically simple approach is highly versatile and can readily be applied to characterize all major CPP properties that normally require multiple assays, including amount taken up by cells (in moles/cell), uptake efficiency, internalization pathways, intracellular distribution, intracellular degradation and toxicity threshold. The FACS-FCS approach provides a means for quantifying any intracellular biochemical entity, whether expressed in the cell or introduced exogenously and transported across the plasma membrane.

Original languageEnglish (US)
Pages (from-to)1499-1506
Number of pages8
JournalBiochimica et Biophysica Acta - Biomembranes
Issue number7
StatePublished - Jul 1 2016



  • Cellular uptake
  • Drug delivery
  • Internalization pathways
  • Intracellular degradation
  • Intracellular distribution
  • Toxicity threshold

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Cell Biology

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