Cargando…

Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection

Cytokine proteins are known as biomarker molecules, characteristic of a disease or specific body condition. Monitoring of the cytokine pattern in body fluids can contribute to the diagnosis of diseases. Here we report on the development of an array comprised of different anti-cytokine antibodies on...

Descripción completa

Detalles Bibliográficos
Autores principales: Hospach, Ingeborg, Joseph, Yvonne, Mai, Michaela Kathrin, Krasteva, Nadejda, Nelles, Gabriele
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4979058/
https://www.ncbi.nlm.nih.gov/pubmed/27600349
http://dx.doi.org/10.3390/microarrays3040282
_version_ 1782447265388953600
author Hospach, Ingeborg
Joseph, Yvonne
Mai, Michaela Kathrin
Krasteva, Nadejda
Nelles, Gabriele
author_facet Hospach, Ingeborg
Joseph, Yvonne
Mai, Michaela Kathrin
Krasteva, Nadejda
Nelles, Gabriele
author_sort Hospach, Ingeborg
collection PubMed
description Cytokine proteins are known as biomarker molecules, characteristic of a disease or specific body condition. Monitoring of the cytokine pattern in body fluids can contribute to the diagnosis of diseases. Here we report on the development of an array comprised of different anti-cytokine antibodies on an activated solid support coupled with a fluorescence readout mechanism. Optimization of the array preparation was done in regard of spot homogeneity and spot size. The proinflammatory cytokines Tumor Necrosis Factor alpha (TNFα) and Interleukin 6 (IL-6) were chosen as the first targets of interest. First, the solid support for covalent antibody immobilization and an adequate fluorescent label were selected. Three differently functionalized glass substrates for spotting were compared: amine and epoxy, both having a two-dimensional structure, and the NHS functionalized hydrogel (NHS-3D). The NHS-hydrogel functionalization of the substrate was best suited to antibody immobilization. Then, the optimization of plotting parameters and geometry as well as buffer media were investigated, considering the ambient analyte theory of Roger Ekins. As a first step towards real sample studies, a proof of principle of cytokine detection has been established.
format Online
Article
Text
id pubmed-4979058
institution National Center for Biotechnology Information
language English
publishDate 2014
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-49790582016-09-06 Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection Hospach, Ingeborg Joseph, Yvonne Mai, Michaela Kathrin Krasteva, Nadejda Nelles, Gabriele Microarrays (Basel) Article Cytokine proteins are known as biomarker molecules, characteristic of a disease or specific body condition. Monitoring of the cytokine pattern in body fluids can contribute to the diagnosis of diseases. Here we report on the development of an array comprised of different anti-cytokine antibodies on an activated solid support coupled with a fluorescence readout mechanism. Optimization of the array preparation was done in regard of spot homogeneity and spot size. The proinflammatory cytokines Tumor Necrosis Factor alpha (TNFα) and Interleukin 6 (IL-6) were chosen as the first targets of interest. First, the solid support for covalent antibody immobilization and an adequate fluorescent label were selected. Three differently functionalized glass substrates for spotting were compared: amine and epoxy, both having a two-dimensional structure, and the NHS functionalized hydrogel (NHS-3D). The NHS-hydrogel functionalization of the substrate was best suited to antibody immobilization. Then, the optimization of plotting parameters and geometry as well as buffer media were investigated, considering the ambient analyte theory of Roger Ekins. As a first step towards real sample studies, a proof of principle of cytokine detection has been established. MDPI 2014-12-09 /pmc/articles/PMC4979058/ /pubmed/27600349 http://dx.doi.org/10.3390/microarrays3040282 Text en © 2014 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Hospach, Ingeborg
Joseph, Yvonne
Mai, Michaela Kathrin
Krasteva, Nadejda
Nelles, Gabriele
Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection
title Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection
title_full Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection
title_fullStr Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection
title_full_unstemmed Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection
title_short Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection
title_sort fabrication of homogeneous high-density antibody microarrays for cytokine detection
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4979058/
https://www.ncbi.nlm.nih.gov/pubmed/27600349
http://dx.doi.org/10.3390/microarrays3040282
work_keys_str_mv AT hospachingeborg fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection
AT josephyvonne fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection
AT maimichaelakathrin fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection
AT krastevanadejda fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection
AT nellesgabriele fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection