title: GFP-complementation assay to detect functional CPP and protein delivery into living cells.
abstract: Efficient cargo uptake is essential for cell-penetrating peptide (CPP) therapeutics, which deliver widely diverse cargoes by exploiting natural cell processes to penetrate the cell's membranes. Yet most current CPP activity assays are hampered by limitations in assessing uptake, including confounding effects of conjugated fluorophores or ligands, indirect read-outs requiring secondary processing, and difficulty in discriminating internalization from endosomally trapped cargo. Split-complementation Endosomal Escape (SEE) provides the first direct assay visualizing true cytoplasmic-delivery of proteins at biologically relevant concentrations. The SEE assay has minimal background, is amenable to high-throughput processes, and adaptable to different transient and stable cell lines. This split-GFP-based platform can be useful to study transduction mechanisms, cellular imaging, and characterizing novel CPPs as pharmaceutical delivery agents in the treatment of disease.
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Nadia
authors.initials: N
authors.lastname: Milech
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Brooke A C
authors.initials: BA
authors.lastname: Longville
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Paula T
authors.initials: PT
authors.lastname: Cunningham
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Marie N
authors.initials: MN
authors.lastname: Scobie
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Heique M
authors.initials: HM
authors.lastname: Bogdawa
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Scott
authors.initials: S
authors.lastname: Winslow
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Mark
authors.initials: M
authors.lastname: Anastasas
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Theresa
authors.initials: T
authors.lastname: Connor
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Ferrer
authors.initials: F
authors.lastname: Ong
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Shane R
authors.initials: SR
authors.lastname: Stone
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Maria
authors.initials: M
authors.lastname: Kerfoot
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Tatjana
authors.initials: T
authors.lastname: Heinrich
authors.affiliation: Phylogica. Subiaco East, Western Australia, 6008, Australia.
authors.forename: Karen M
authors.initials: KM
authors.lastname: Kroeger
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Yew-Foon
authors.initials: YF
authors.lastname: Tan
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Katrin
authors.initials: K
authors.lastname: Hoffmann
authors.affiliation: Telethon Kids Institute &Centre for Child Health Research, The University of Western Australia, Drug Discovery Group. West Perth, 6872, Australia.
authors.forename: Wayne R
authors.initials: WR
authors.lastname: Thomas
authors.affiliation: Phylogica. Subiaco East, Western Australia, 6008, Australia.
authors.forename: Paul M
authors.initials: PM
authors.lastname: Watt
authors.affiliation: Phylogica. Subiaco East, Western Australia, 6008, Australia.
authors.forename: Richard M
authors.initials: RM
authors.lastname: Hopkins
mesh_terms: D000818:Animals; D016466:CHO Cells; D057846:Cell-Penetrating Peptides / Q000737:chemistry / Q000493:pharmacokinetics / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D016503:Drug Delivery Systems / Q000379:methods*; D011992:Endosomes / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000493:pharmacokinetics / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans
chemical_list: D057846:Cell-Penetrating Peptides; D049452:Green Fluorescent Proteins