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...
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doaj-69acfd63a8ea41d2a412270dded1382e2020-11-25T01:28:27ZengMDPI AGMicroarrays2076-39052014-12-013428230110.3390/microarrays3040282microarrays3040282Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine DetectionIngeborg Hospach0Yvonne Joseph1Michaela Kathrin Mai2Nadejda Krasteva3Gabriele Nelles4Materials Science Laboratory, Sony Deutschland GmbH, Hedelfinger Strasse 61, 70327 Stuttgart, GermanyInstitute of Electronic and Sensor Materials, Technische Universität Bergakademie Freiberg, Gustav-Zeuner-Strasse 3, 09599 Freiberg, GermanyMaterials Science Laboratory, Sony Deutschland GmbH, Hedelfinger Strasse 61, 70327 Stuttgart, GermanyMaterials Science Laboratory, Sony Deutschland GmbH, Hedelfinger Strasse 61, 70327 Stuttgart, GermanyMaterials Science Laboratory, Sony Deutschland GmbH, Hedelfinger Strasse 61, 70327 Stuttgart, GermanyCytokine 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.http://www.mdpi.com/2076-3905/3/4/282microarrayproteinantibodycytokinesspottinghydrogelbiosensorfluorescence readout |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Ingeborg Hospach Yvonne Joseph Michaela Kathrin Mai Nadejda Krasteva Gabriele Nelles |
spellingShingle |
Ingeborg Hospach Yvonne Joseph Michaela Kathrin Mai Nadejda Krasteva Gabriele Nelles Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection Microarrays microarray protein antibody cytokines spotting hydrogel biosensor fluorescence readout |
author_facet |
Ingeborg Hospach Yvonne Joseph Michaela Kathrin Mai Nadejda Krasteva Gabriele Nelles |
author_sort |
Ingeborg Hospach |
title |
Fabrication of Homogeneous High-Density Antibody Microarrays for Cytokine Detection |
title_short |
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_sort |
fabrication of homogeneous high-density antibody microarrays for cytokine detection |
publisher |
MDPI AG |
series |
Microarrays |
issn |
2076-3905 |
publishDate |
2014-12-01 |
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. |
topic |
microarray protein antibody cytokines spotting hydrogel biosensor fluorescence readout |
url |
http://www.mdpi.com/2076-3905/3/4/282 |
work_keys_str_mv |
AT ingeborghospach fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection AT yvonnejoseph fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection AT michaelakathrinmai fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection AT nadejdakrasteva fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection AT gabrielenelles fabricationofhomogeneoushighdensityantibodymicroarraysforcytokinedetection |
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1725101557670739968 |