Design and simple assembly of gold nanostar bioconjugates for surface-enhanced raman spectroscopy immunoassays

Maria João Oliveira, Miguel P. de Almeida, Daniela Nunes, Elvira Fortunato, Rodrigo Martins, Eulália Pereira, Hugh J. Byrne, Hugo Águas, Ricardo Franco

Research output: Contribution to journalArticle

Abstract

Immunoassays using Surface-Enhanced Raman Spectroscopy are especially interesting on account not only of their increased sensitivity, but also due to its easy translation to point-of-care formats. The bases for these assays are bioconjugates of polyclonal antibodies and anisotropic gold nanoparticles functionalized with a Raman reporter. These bioconjugates, once loaded with the antigen analyte, can react on a sandwich format with the same antibodies immobilized on a surface. This surface can then be used for detection, on a microfluidics or immunochromatographic platform. Here, we have assembled bioconjugates of gold nanostars functionalized with 4-mercaptobenzoic acid, and anti-horseradish peroxidase antibodies. The assembly was by simple incubation, and agarose gel electrophoresis determined a high gold nanostar to antibody binding constant. The functionality of the bioconjugates is easy to determine since the respective antigen presents peroxidase enzymatic activity. Furthermore, the chosen antibody is a generic immunoglobulin G (IgG) antibody, opening the application of these principles to other antibody-antigen systems. Surface-Enhanced Raman Spectroscopy analysis of these bioconjugates indicated antigen detection down to 50 _U of peroxidase activity. All steps of conjugation were fully characterized by ultraviolet-visible spectroscopy, dynamic light scattering, ζ-Potential, scanning electron microscopy, and agarose gel electrophoresis. Based on the latter technique, a proof-of-concept was established for the proposed immunoassay.

Original languageEnglish
Article number1561
JournalNanomaterials
Volume9
Issue number11
DOIs
Publication statusPublished - 1 Nov 2019

Fingerprint

Antibodies
Gold
Raman spectroscopy
Antigens
Electrophoresis
Sepharose
Peroxidase
Gels
Immobilized Antibodies
Ultraviolet visible spectroscopy
Dynamic light scattering
Horseradish Peroxidase
Microfluidics
Assays
Immunoglobulin G
Nanoparticles
Scanning electron microscopy
Acids

Keywords

  • Agarose gel electrophoresis
  • Dynamic light scattering
  • Enhanced Raman Spectroscopy
  • Gold nanoparticles
  • Gold nanostars
  • Horseradish peroxidase
  • Immunoassay
  • Immunoconjugates
  • Raman reporter
  • Surface enhanced Raman spectroscopy Surface

Cite this

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title = "Design and simple assembly of gold nanostar bioconjugates for surface-enhanced raman spectroscopy immunoassays",
abstract = "Immunoassays using Surface-Enhanced Raman Spectroscopy are especially interesting on account not only of their increased sensitivity, but also due to its easy translation to point-of-care formats. The bases for these assays are bioconjugates of polyclonal antibodies and anisotropic gold nanoparticles functionalized with a Raman reporter. These bioconjugates, once loaded with the antigen analyte, can react on a sandwich format with the same antibodies immobilized on a surface. This surface can then be used for detection, on a microfluidics or immunochromatographic platform. Here, we have assembled bioconjugates of gold nanostars functionalized with 4-mercaptobenzoic acid, and anti-horseradish peroxidase antibodies. The assembly was by simple incubation, and agarose gel electrophoresis determined a high gold nanostar to antibody binding constant. The functionality of the bioconjugates is easy to determine since the respective antigen presents peroxidase enzymatic activity. Furthermore, the chosen antibody is a generic immunoglobulin G (IgG) antibody, opening the application of these principles to other antibody-antigen systems. Surface-Enhanced Raman Spectroscopy analysis of these bioconjugates indicated antigen detection down to 50 _U of peroxidase activity. All steps of conjugation were fully characterized by ultraviolet-visible spectroscopy, dynamic light scattering, ζ-Potential, scanning electron microscopy, and agarose gel electrophoresis. Based on the latter technique, a proof-of-concept was established for the proposed immunoassay.",
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Design and simple assembly of gold nanostar bioconjugates for surface-enhanced raman spectroscopy immunoassays. / Oliveira, Maria João; de Almeida, Miguel P.; Nunes, Daniela; Fortunato, Elvira; Martins, Rodrigo; Pereira, Eulália; Byrne, Hugh J.; Águas, Hugo; Franco, Ricardo.

In: Nanomaterials, Vol. 9, No. 11, 1561, 01.11.2019.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Design and simple assembly of gold nanostar bioconjugates for surface-enhanced raman spectroscopy immunoassays

AU - Oliveira, Maria João

AU - de Almeida, Miguel P.

AU - Nunes, Daniela

AU - Fortunato, Elvira

AU - Martins, Rodrigo

AU - Pereira, Eulália

AU - Byrne, Hugh J.

AU - Águas, Hugo

AU - Franco, Ricardo

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N2 - Immunoassays using Surface-Enhanced Raman Spectroscopy are especially interesting on account not only of their increased sensitivity, but also due to its easy translation to point-of-care formats. The bases for these assays are bioconjugates of polyclonal antibodies and anisotropic gold nanoparticles functionalized with a Raman reporter. These bioconjugates, once loaded with the antigen analyte, can react on a sandwich format with the same antibodies immobilized on a surface. This surface can then be used for detection, on a microfluidics or immunochromatographic platform. Here, we have assembled bioconjugates of gold nanostars functionalized with 4-mercaptobenzoic acid, and anti-horseradish peroxidase antibodies. The assembly was by simple incubation, and agarose gel electrophoresis determined a high gold nanostar to antibody binding constant. The functionality of the bioconjugates is easy to determine since the respective antigen presents peroxidase enzymatic activity. Furthermore, the chosen antibody is a generic immunoglobulin G (IgG) antibody, opening the application of these principles to other antibody-antigen systems. Surface-Enhanced Raman Spectroscopy analysis of these bioconjugates indicated antigen detection down to 50 _U of peroxidase activity. All steps of conjugation were fully characterized by ultraviolet-visible spectroscopy, dynamic light scattering, ζ-Potential, scanning electron microscopy, and agarose gel electrophoresis. Based on the latter technique, a proof-of-concept was established for the proposed immunoassay.

AB - Immunoassays using Surface-Enhanced Raman Spectroscopy are especially interesting on account not only of their increased sensitivity, but also due to its easy translation to point-of-care formats. The bases for these assays are bioconjugates of polyclonal antibodies and anisotropic gold nanoparticles functionalized with a Raman reporter. These bioconjugates, once loaded with the antigen analyte, can react on a sandwich format with the same antibodies immobilized on a surface. This surface can then be used for detection, on a microfluidics or immunochromatographic platform. Here, we have assembled bioconjugates of gold nanostars functionalized with 4-mercaptobenzoic acid, and anti-horseradish peroxidase antibodies. The assembly was by simple incubation, and agarose gel electrophoresis determined a high gold nanostar to antibody binding constant. The functionality of the bioconjugates is easy to determine since the respective antigen presents peroxidase enzymatic activity. Furthermore, the chosen antibody is a generic immunoglobulin G (IgG) antibody, opening the application of these principles to other antibody-antigen systems. Surface-Enhanced Raman Spectroscopy analysis of these bioconjugates indicated antigen detection down to 50 _U of peroxidase activity. All steps of conjugation were fully characterized by ultraviolet-visible spectroscopy, dynamic light scattering, ζ-Potential, scanning electron microscopy, and agarose gel electrophoresis. Based on the latter technique, a proof-of-concept was established for the proposed immunoassay.

KW - Agarose gel electrophoresis

KW - Dynamic light scattering

KW - Enhanced Raman Spectroscopy

KW - Gold nanoparticles

KW - Gold nanostars

KW - Horseradish peroxidase

KW - Immunoassay

KW - Immunoconjugates

KW - Raman reporter

KW - Surface enhanced Raman spectroscopy Surface

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SN - 2079-4991

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