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. 2015 Feb 4;2:14056. doi: 10.1038/mtm.2014.56

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Copyright © 2015 American Society of Gene & Cell Therapy

This work is licensed. under a Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-sa/3.0/

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Annexin V-7AAD apoptosis assay and protein delivery efficiency analysis by flow cytometry. (a) B16, Colo857, HCT116 p53^+/+, and HCT116 p53^−/− cells were incubated with AlexaFluor 488 labeled proteins (10 nmol/l) and stained with Annexin V-APC and 7-AAD. Alive cells show negative staining for Annexin V and 7-AAD. Cells undergoing apoptosis show positive staining for Annexin V and negative for 7-AAD, while cells positive for both Annexin V and 7-AAD are either in the end stage of apoptosis, undergoing necrosis or already dead. (b) Examples of FACS profiles of untreated cells and doxorubicin-treated cells, used as controls for viability/toxicity setting gates. (c) Representative plot of HCT116 p53^−/− cells incubated with 0.3 µmol/l MD₁₁-eGFP, showing no toxic effect associated to the MD₁₁-mediated protein internalization. (d) Evaluation of delivery efficiency of AlexaFluor 488-labeled recombinant proteins in selected cells. Histograms represent percentages of internalized MD₁₁-AN, MD₁₁-NF, MD₁₁-wt, and MD₁₁-K510R proteins in target cells.