title: Quantification of intracellular payload release from polymersome nanoparticles.
abstract: Polymersome nanoparticles (PMs) are attractive candidates for spatio-temporal controlled delivery of therapeutic agents. Although many studies have addressed cellular uptake of solid nanoparticles, there is very little data available on intracellular release of molecules encapsulated in membranous carriers, such as polymersomes. Here, we addressed this by developing a quantitative assay based on the hydrophilic dye, fluorescein. Fluorescein was encapsulated stably in PMs of mean diameter 85 nm, with minimal leakage after sustained dialysis. No fluorescence was detectable from fluorescein PMs, indicating quenching. Following incubation of L929 cells with fluorescein PMs, there was a gradual increase in intracellular fluorescence, indicating PM disruption and cytosolic release of fluorescein. By combining absorbance measurements with flow cytometry, we quantified the real-time intracellular release of a fluorescein at a single-cell resolution. We found that 173 ± 38 polymersomes released their payload per cell, with significant heterogeneity in uptake, despite controlled synchronisation of cell cycle. This novel method for quantification of the release of compounds from nanoparticles provides fundamental information on cellular uptake of nanoparticle-encapsulated compounds. It also illustrates the stochastic nature of population distribution in homogeneous cell populations, a factor that must be taken into account in clinical use of this technology.
authors.affiliation: Centre for Human Development, Stem Cells and Regeneration, University of Southampton Faculty of Medicine, Tremona Road, Southampton, SO16 6YD, United Kingdom.|Institute for Life Sciences, Centre for Biological Sciences, B85, University Road, University of Southampton, United Kingdom.
authors.forename: Edoardo
authors.initials: E
authors.lastname: Scarpa
authors.affiliation: Institute for Life Sciences, Centre for Biological Sciences, B85, University Road, University of Southampton, United Kingdom.
authors.forename: Joanne L
authors.initials: JL
authors.lastname: Bailey
authors.affiliation: Centre for Human Development, Stem Cells and Regeneration, University of Southampton Faculty of Medicine, Tremona Road, Southampton, SO16 6YD, United Kingdom.|Institute for Life Sciences, Centre for Biological Sciences, B85, University Road, University of Southampton, United Kingdom.
authors.forename: Agnieszka A
authors.initials: AA
authors.lastname: Janeczek
authors.affiliation: Centre for Human Development, Stem Cells and Regeneration, University of Southampton Faculty of Medicine, Tremona Road, Southampton, SO16 6YD, United Kingdom.|Institute for Life Sciences, Centre for Biological Sciences, B85, University Road, University of Southampton, United Kingdom.
authors.forename: Patrick S
authors.initials: PS
authors.lastname: Stumpf
authors.affiliation: Institute for Life Sciences, Centre for Biological Sciences, B85, University Road, University of Southampton, United Kingdom.
authors.forename: Alexander H
authors.initials: AH
authors.lastname: Johnston
authors.affiliation: Centre for Human Development, Stem Cells and Regeneration, University of Southampton Faculty of Medicine, Tremona Road, Southampton, SO16 6YD, United Kingdom.|Institute for Life Sciences, Centre for Biological Sciences, B85, University Road, University of Southampton, United Kingdom.
authors.forename: Richard O C
authors.initials: RO
authors.lastname: Oreffo
authors.affiliation: Department of Obstetrics and Gynaecology, Faculty of Medicine, University of Malaya, Kuala Lumpur, 50603, Malaysia.|University of Malaya Cancer Research Institute (UMCRI), University of Malaya, Kuala Lumpur, 50603, Malaysia.
authors.forename: Yin L
authors.initials: YL
authors.lastname: Woo
authors.affiliation: Centre for Human Development, Stem Cells and Regeneration, University of Southampton Faculty of Medicine, Tremona Road, Southampton, SO16 6YD, United Kingdom.
authors.forename: Ying C
authors.initials: YC
authors.lastname: Cheong
authors.affiliation: Centre for Human Development, Stem Cells and Regeneration, University of Southampton Faculty of Medicine, Tremona Road, Southampton, SO16 6YD, United Kingdom.|Institute for Life Sciences, Centre for Biological Sciences, B85, University Road, University of Southampton, United Kingdom.|Bioengineering Sciences Group, Faculty of Engineering and the Environment, University of Southampton, Highfield, Southampton, SO17 1BJ, United Kingdom.
authors.forename: Nicholas D
authors.initials: ND
authors.lastname: Evans
authors.affiliation: Institute for Life Sciences, Centre for Biological Sciences, B85, University Road, University of Southampton, United Kingdom.|Clinical and Experimental Sciences, Medicine, University of Southampton, SO17 1BJ, United Kingdom.
authors.forename: Tracey A
authors.initials: TA
authors.lastname: Newman