Nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction

Trisha Eustaquio, James F LearyWeldon School of Biomedical Engineering, Purdue University, West Lafayette, IN, USABackground: Determination of the fate of nanoparticles (NPs) in a biological system, or NP biodistribution, is critical in evaluating an NP formulation for nanomedicine. Current methods...

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Autores principales:	Eustaquio T, Leary JF
Formato:	article
Lenguaje:	EN
Publicado:	Dove Medical Press 2012
Materias:	Medicine (General) R5-920
Acceso en línea:	https://doaj.org/article/a5fabaacdd1c4440909deb53ba9b7486
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spelling	oai:doaj.org-article:a5fabaacdd1c4440909deb53ba9b74862021-12-02T01:43:42ZNanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction1176-91141178-2013https://doaj.org/article/a5fabaacdd1c4440909deb53ba9b74862012-11-01T00:00:00Zhttp://www.dovepress.com/nanobarcoding-detecting-nanoparticles-in-biological-samples-using-in-s-a11427https://doaj.org/toc/1176-9114https://doaj.org/toc/1178-2013Trisha Eustaquio, James F LearyWeldon School of Biomedical Engineering, Purdue University, West Lafayette, IN, USABackground: Determination of the fate of nanoparticles (NPs) in a biological system, or NP biodistribution, is critical in evaluating an NP formulation for nanomedicine. Current methods to determine NP biodistribution are greatly inadequate, due to their limited detection thresholds. Herein, proof of concept of a novel method for improved NP detection based on in situ polymerase chain reaction (ISPCR), coined &ldquo;nanobarcoding,&rdquo; is demonstrated.Methods: Nanobarcoded superparamagnetic iron oxide nanoparticles (NB-SPIONs) were characterized by dynamic light scattering, zeta potential, and hyperspectral imaging measurements. Cellular uptake of Cy5-labeled NB-SPIONs (Cy5-NB-SPIONs) was imaged by confocal microscopy. The feasibility of the nanobarcoding method was first validated by solution-phase PCR and &ldquo;pseudo&rdquo;-ISPCR before implementation in the model in vitro system of HeLa human cervical adenocarcinoma cells, a cell line commonly used for ISPCR-mediated detection of human papilloma virus (HPV).Results: Dynamic light-scattering measurements showed that NB conjugation stabilized SPION size in different dispersion media compared to that of its precursor, carboxylated SPIONs (COOH-SPIONs), while the zeta potential became more positive after NB conjugation. Hyperspectral imaging confirmed NB conjugation and showed that the NB completely covered the SPION surface. Solution-phase PCR and pseudo-ISPCR showed that the expected amplicons were exclusively generated from the NB-SPIONs in a dose-dependent manner. Although confocal microscopy revealed minimal cellular uptake of Cy5-NB-SPIONs at 50 nM over 24 hours in individual cells, ISPCR detected definitive NB-SPION signals inside HeLa cells over large sample areas.Conclusion: Proof of concept of the nanobarcoding method has been demonstrated in in vitro systems, but the technique needs further development before its widespread use as a standardized assay.Keywords: nanomedicine, in situ PCR, single cell, DNA-functionalized nanoparticlesEustaquio TLeary JFDove Medical PressarticleMedicine (General)R5-920ENInternational Journal of Nanomedicine, Vol 2012, Iss default, Pp 5625-5639 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine (General) R5-920
spellingShingle	Medicine (General) R5-920 Eustaquio T Leary JF Nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction
description	Trisha Eustaquio, James F LearyWeldon School of Biomedical Engineering, Purdue University, West Lafayette, IN, USABackground: Determination of the fate of nanoparticles (NPs) in a biological system, or NP biodistribution, is critical in evaluating an NP formulation for nanomedicine. Current methods to determine NP biodistribution are greatly inadequate, due to their limited detection thresholds. Herein, proof of concept of a novel method for improved NP detection based on in situ polymerase chain reaction (ISPCR), coined &ldquo;nanobarcoding,&rdquo; is demonstrated.Methods: Nanobarcoded superparamagnetic iron oxide nanoparticles (NB-SPIONs) were characterized by dynamic light scattering, zeta potential, and hyperspectral imaging measurements. Cellular uptake of Cy5-labeled NB-SPIONs (Cy5-NB-SPIONs) was imaged by confocal microscopy. The feasibility of the nanobarcoding method was first validated by solution-phase PCR and &ldquo;pseudo&rdquo;-ISPCR before implementation in the model in vitro system of HeLa human cervical adenocarcinoma cells, a cell line commonly used for ISPCR-mediated detection of human papilloma virus (HPV).Results: Dynamic light-scattering measurements showed that NB conjugation stabilized SPION size in different dispersion media compared to that of its precursor, carboxylated SPIONs (COOH-SPIONs), while the zeta potential became more positive after NB conjugation. Hyperspectral imaging confirmed NB conjugation and showed that the NB completely covered the SPION surface. Solution-phase PCR and pseudo-ISPCR showed that the expected amplicons were exclusively generated from the NB-SPIONs in a dose-dependent manner. Although confocal microscopy revealed minimal cellular uptake of Cy5-NB-SPIONs at 50 nM over 24 hours in individual cells, ISPCR detected definitive NB-SPION signals inside HeLa cells over large sample areas.Conclusion: Proof of concept of the nanobarcoding method has been demonstrated in in vitro systems, but the technique needs further development before its widespread use as a standardized assay.Keywords: nanomedicine, in situ PCR, single cell, DNA-functionalized nanoparticles
format	article
author	Eustaquio T Leary JF
author_facet	Eustaquio T Leary JF
author_sort	Eustaquio T
title	Nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction
title_short	Nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction
title_full	Nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction
title_fullStr	Nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction
title_full_unstemmed	Nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction
title_sort	nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction
publisher	Dove Medical Press
publishDate	2012
url	https://doaj.org/article/a5fabaacdd1c4440909deb53ba9b7486
work_keys_str_mv	AT eustaquiot nanobarcodingdetectingnanoparticlesinbiologicalsamplesusinginsitupolymerasechainreaction AT learyjf nanobarcodingdetectingnanoparticlesinbiologicalsamplesusinginsitupolymerasechainreaction
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Nanobarcoding: detecting nanoparticles in biological samples using in situ polymerase chain reaction

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