Chemiluminometric Immuno-Analysis of Innate Immune Response against Repetitive Bacterial Stimulations for the Same Mammalian Cells
For monitoring of human cellular response to repetitive bacterial stimulations (e.g., Pseudomonas aeruginosa in a lysate form), we devised a chemiluminescent immuno-analytical system for toll-like receptor 1 (TLR1) as marker present on cell surfaces (e.g., A549). Upon stimulation, TLR1 recognizes pa...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4127502/ https://www.ncbi.nlm.nih.gov/pubmed/25109895 http://dx.doi.org/10.1038/srep06011 |
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author | Jeon, Jin-Woo Cho, Il-Hoon Ha, Un-Hwan Seo, Sung-Kyu Paek, Se-Hwan |
author_facet | Jeon, Jin-Woo Cho, Il-Hoon Ha, Un-Hwan Seo, Sung-Kyu Paek, Se-Hwan |
author_sort | Jeon, Jin-Woo |
collection | PubMed |
description | For monitoring of human cellular response to repetitive bacterial stimulations (e.g., Pseudomonas aeruginosa in a lysate form), we devised a chemiluminescent immuno-analytical system for toll-like receptor 1 (TLR1) as marker present on cell surfaces (e.g., A549). Upon stimulation, TLR1 recognizes pathogen-associated molecular patterns of the infectious agent and are then up-regulated via activation of the nuclear factor-κB (NF-κB) pathway. In this study, the receptor density was quantified by employing an antibody specific to the target receptor and by producing a chemiluminometric signal from an enzyme labeled to the binder. The activated status was then switched back to normal down-regulated stage, by changing the culture medium to one containing animal serum. The major factors affecting activation were the stimulation dose of the bacterial lysate, stimulation timing during starvation, and up- and down-regulation time intervals. Reiterative TLR regulation switching up to three times was not affected by either antibody remained after immunoassay or enzyme substrate (e.g., hydrogen peroxide) in solution. This immuno-analysis for TLRs could be unique to acquire accumulated response of the human cells to repeated stimulations and, therefore, can eventually apply to persistency testing of the cellular regulation in screening of anti-inflammatory substances. |
format | Online Article Text |
id | pubmed-4127502 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | Nature Publishing Group |
record_format | MEDLINE/PubMed |
spelling | pubmed-41275022014-08-14 Chemiluminometric Immuno-Analysis of Innate Immune Response against Repetitive Bacterial Stimulations for the Same Mammalian Cells Jeon, Jin-Woo Cho, Il-Hoon Ha, Un-Hwan Seo, Sung-Kyu Paek, Se-Hwan Sci Rep Article For monitoring of human cellular response to repetitive bacterial stimulations (e.g., Pseudomonas aeruginosa in a lysate form), we devised a chemiluminescent immuno-analytical system for toll-like receptor 1 (TLR1) as marker present on cell surfaces (e.g., A549). Upon stimulation, TLR1 recognizes pathogen-associated molecular patterns of the infectious agent and are then up-regulated via activation of the nuclear factor-κB (NF-κB) pathway. In this study, the receptor density was quantified by employing an antibody specific to the target receptor and by producing a chemiluminometric signal from an enzyme labeled to the binder. The activated status was then switched back to normal down-regulated stage, by changing the culture medium to one containing animal serum. The major factors affecting activation were the stimulation dose of the bacterial lysate, stimulation timing during starvation, and up- and down-regulation time intervals. Reiterative TLR regulation switching up to three times was not affected by either antibody remained after immunoassay or enzyme substrate (e.g., hydrogen peroxide) in solution. This immuno-analysis for TLRs could be unique to acquire accumulated response of the human cells to repeated stimulations and, therefore, can eventually apply to persistency testing of the cellular regulation in screening of anti-inflammatory substances. Nature Publishing Group 2014-08-11 /pmc/articles/PMC4127502/ /pubmed/25109895 http://dx.doi.org/10.1038/srep06011 Text en Copyright © 2014, Macmillan Publishers Limited. All rights reserved http://creativecommons.org/licenses/by-nc-nd/4.0/ This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder in order to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-nd/4.0/ |
spellingShingle | Article Jeon, Jin-Woo Cho, Il-Hoon Ha, Un-Hwan Seo, Sung-Kyu Paek, Se-Hwan Chemiluminometric Immuno-Analysis of Innate Immune Response against Repetitive Bacterial Stimulations for the Same Mammalian Cells |
title | Chemiluminometric Immuno-Analysis of Innate Immune Response against Repetitive Bacterial Stimulations for the Same Mammalian Cells |
title_full | Chemiluminometric Immuno-Analysis of Innate Immune Response against Repetitive Bacterial Stimulations for the Same Mammalian Cells |
title_fullStr | Chemiluminometric Immuno-Analysis of Innate Immune Response against Repetitive Bacterial Stimulations for the Same Mammalian Cells |
title_full_unstemmed | Chemiluminometric Immuno-Analysis of Innate Immune Response against Repetitive Bacterial Stimulations for the Same Mammalian Cells |
title_short | Chemiluminometric Immuno-Analysis of Innate Immune Response against Repetitive Bacterial Stimulations for the Same Mammalian Cells |
title_sort | chemiluminometric immuno-analysis of innate immune response against repetitive bacterial stimulations for the same mammalian cells |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4127502/ https://www.ncbi.nlm.nih.gov/pubmed/25109895 http://dx.doi.org/10.1038/srep06011 |
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