Synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.

The efficient transport of micron-sized beads into cells, via a non-endocytosis mediated mechanism, has only recently been described. As such there is considerable scope for optimization and exploitation of this procedure to enable imaging and sensing applications to be realized. Herein, we report t...

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Autores principales:	Jonathan M Behrendt, David Nagel, Evita Chundoo, Lois M Alexander, Damien Dupin, Anna V Hine, Mark Bradley, Andrew J Sutherland
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2013
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/f8ee7efd1f304cea85f20c4deb4575ae
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spelling	oai:doaj.org-article:f8ee7efd1f304cea85f20c4deb4575ae2021-11-18T07:53:04ZSynthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.1932-620310.1371/journal.pone.0050713https://doaj.org/article/f8ee7efd1f304cea85f20c4deb4575ae2013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/23526923/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203The efficient transport of micron-sized beads into cells, via a non-endocytosis mediated mechanism, has only recently been described. As such there is considerable scope for optimization and exploitation of this procedure to enable imaging and sensing applications to be realized. Herein, we report the design, synthesis and characterization of fluorescent microsphere-based cellular delivery agents that can also carry biological cargoes. These core-shell polymer microspheres possess two distinct chemical environments; the core is hydrophobic and can be labeled with fluorescent dye, to permit visual tracking of the microsphere during and after cellular delivery, whilst the outer shell renders the external surfaces of the microspheres hydrophilic, thus facilitating both bioconjugation and cellular compatibility. Cross-linked core particles were prepared in a dispersion polymerization reaction employing styrene, divinylbenzene and a thiol-functionalized co-monomer. These core particles were then shelled in a seeded emulsion polymerization reaction, employing styrene, divinylbenzene and methacrylic acid, to generate orthogonally functionalized core-shell microspheres which were internally labeled via the core thiol moieties through reaction with a thiol reactive dye (DY630-maleimide). Following internal labeling, bioconjugation of green fluorescent protein (GFP) to their carboxyl-functionalized surfaces was successfully accomplished using standard coupling protocols. The resultant dual-labeled microspheres were visualized by both of the fully resolvable fluorescence emissions of their cores (DY630) and shells (GFP). In vitro cellular uptake of these microspheres by HeLa cells was demonstrated conventionally by fluorescence-based flow cytometry, whilst MTT assays demonstrated that 92% of HeLa cells remained viable after uptake. Due to their size and surface functionalities, these far-red-labeled microspheres are ideal candidates for in vitro, cellular delivery of proteins.Jonathan M BehrendtDavid NagelEvita ChundooLois M AlexanderDamien DupinAnna V HineMark BradleyAndrew J SutherlandPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 3, p e50713 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Jonathan M Behrendt David Nagel Evita Chundoo Lois M Alexander Damien Dupin Anna V Hine Mark Bradley Andrew J Sutherland Synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.
description	The efficient transport of micron-sized beads into cells, via a non-endocytosis mediated mechanism, has only recently been described. As such there is considerable scope for optimization and exploitation of this procedure to enable imaging and sensing applications to be realized. Herein, we report the design, synthesis and characterization of fluorescent microsphere-based cellular delivery agents that can also carry biological cargoes. These core-shell polymer microspheres possess two distinct chemical environments; the core is hydrophobic and can be labeled with fluorescent dye, to permit visual tracking of the microsphere during and after cellular delivery, whilst the outer shell renders the external surfaces of the microspheres hydrophilic, thus facilitating both bioconjugation and cellular compatibility. Cross-linked core particles were prepared in a dispersion polymerization reaction employing styrene, divinylbenzene and a thiol-functionalized co-monomer. These core particles were then shelled in a seeded emulsion polymerization reaction, employing styrene, divinylbenzene and methacrylic acid, to generate orthogonally functionalized core-shell microspheres which were internally labeled via the core thiol moieties through reaction with a thiol reactive dye (DY630-maleimide). Following internal labeling, bioconjugation of green fluorescent protein (GFP) to their carboxyl-functionalized surfaces was successfully accomplished using standard coupling protocols. The resultant dual-labeled microspheres were visualized by both of the fully resolvable fluorescence emissions of their cores (DY630) and shells (GFP). In vitro cellular uptake of these microspheres by HeLa cells was demonstrated conventionally by fluorescence-based flow cytometry, whilst MTT assays demonstrated that 92% of HeLa cells remained viable after uptake. Due to their size and surface functionalities, these far-red-labeled microspheres are ideal candidates for in vitro, cellular delivery of proteins.
format	article
author	Jonathan M Behrendt David Nagel Evita Chundoo Lois M Alexander Damien Dupin Anna V Hine Mark Bradley Andrew J Sutherland
author_facet	Jonathan M Behrendt David Nagel Evita Chundoo Lois M Alexander Damien Dupin Anna V Hine Mark Bradley Andrew J Sutherland
author_sort	Jonathan M Behrendt
title	Synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.
title_short	Synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.
title_full	Synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.
title_fullStr	Synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.
title_full_unstemmed	Synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.
title_sort	synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.
publisher	Public Library of Science (PLoS)
publishDate	2013
url	https://doaj.org/article/f8ee7efd1f304cea85f20c4deb4575ae
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Synthesis and characterization of dual-functionalized core-shell fluorescent microspheres for bioconjugation and cellular delivery.

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