Cell penetration is a major obstacle for developing peptide, protein and nucleic acid therapeutics. Most commonly used techniques for measuring cell penetration are qualitative, and most cannot distinguish cytosolic material from material trapped at the cell surface or in endosomes. We have devised a new technique, the chloroalkane penetration assay (CAPA), that measures penetration to the cytosol in a high-throughput, quantitative manner. Here, we describe the advantages and disadvantages of CAPA and demonstrate some of its applications. These include: determining structure-activity relationships for cell penetration of peptide therapeutics, quantitating penetration into different subcellular organelles and compartments, and measuring cytosolic penetration across a wide range of bioactive molecules and drug delivery systems. With CAPA, we are making rapid progress in more specifically defining the features that promote cytosolic penetration for a variety of biomolecules.

Measuring cytosolic penetration using the chloroalkane penetration assay

256th National Meeting (2018)

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