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Effective preparation of a monoclonal antibody against human chromogranin A for immunohistochemical diagnosis
BACKGROUND: Human chromogranin A (CgA) is a ~ 49 kDa secreted protein mainly from neuroendocrine cells and endocrine cells. The CgA values in the diagnosis of tumor, and in the potential role in prognostic and predictive tumor as a biomarker. RESULTS: The synthesized gene of CgA coding area was clon...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5935939/ https://www.ncbi.nlm.nih.gov/pubmed/29728076 http://dx.doi.org/10.1186/s12896-018-0436-z |
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author | Zhang, Danping Xie, Chengjie Wang, Rongzhi Yang, Qinghai Chen, Huiling Ling, Sumei Wang, Shihua Jia, Kunzhi |
author_facet | Zhang, Danping Xie, Chengjie Wang, Rongzhi Yang, Qinghai Chen, Huiling Ling, Sumei Wang, Shihua Jia, Kunzhi |
author_sort | Zhang, Danping |
collection | PubMed |
description | BACKGROUND: Human chromogranin A (CgA) is a ~ 49 kDa secreted protein mainly from neuroendocrine cells and endocrine cells. The CgA values in the diagnosis of tumor, and in the potential role in prognostic and predictive tumor as a biomarker. RESULTS: The synthesized gene of CgA coding area was cloned and expressed as fusion protein CgA-His in procaryotic system. Then the purified CgA-His protein was mixed with QuickAntibody-Mouse5W adjuvant, and injected into mice. The CgA-His protein was also used as coating antigen to determine the antiserum titer. By screening, a stable cell line named 4E5, which can generate anti-CgA monoclonal antibody (mAb), was obtained. The isotype of 4E5 mAb was IgG(2b), and the chromosome number was 102 ± 4. Anti-CgA mAb was purified from ascites fluid, and the affinity constant reached 9.23 × 10(9) L/mol. Furthermore, the specificity of the mAb was determined with ELISA, western blot and immunohistochemistry. Results indicated that the mAb 4E5 was able to detect chromogranin A specifically and sensitively. CONCLUSIONS: A sensitive and reliable method was successfully developed for rapid production of anti-CgA mAb for immunohistochemistry diagnosis in this study, and the current study also provides conclusive guidelines for preparation of mAbs and implements in immunohistochemistry diagnosis. |
format | Online Article Text |
id | pubmed-5935939 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-59359392018-05-11 Effective preparation of a monoclonal antibody against human chromogranin A for immunohistochemical diagnosis Zhang, Danping Xie, Chengjie Wang, Rongzhi Yang, Qinghai Chen, Huiling Ling, Sumei Wang, Shihua Jia, Kunzhi BMC Biotechnol Research Article BACKGROUND: Human chromogranin A (CgA) is a ~ 49 kDa secreted protein mainly from neuroendocrine cells and endocrine cells. The CgA values in the diagnosis of tumor, and in the potential role in prognostic and predictive tumor as a biomarker. RESULTS: The synthesized gene of CgA coding area was cloned and expressed as fusion protein CgA-His in procaryotic system. Then the purified CgA-His protein was mixed with QuickAntibody-Mouse5W adjuvant, and injected into mice. The CgA-His protein was also used as coating antigen to determine the antiserum titer. By screening, a stable cell line named 4E5, which can generate anti-CgA monoclonal antibody (mAb), was obtained. The isotype of 4E5 mAb was IgG(2b), and the chromosome number was 102 ± 4. Anti-CgA mAb was purified from ascites fluid, and the affinity constant reached 9.23 × 10(9) L/mol. Furthermore, the specificity of the mAb was determined with ELISA, western blot and immunohistochemistry. Results indicated that the mAb 4E5 was able to detect chromogranin A specifically and sensitively. CONCLUSIONS: A sensitive and reliable method was successfully developed for rapid production of anti-CgA mAb for immunohistochemistry diagnosis in this study, and the current study also provides conclusive guidelines for preparation of mAbs and implements in immunohistochemistry diagnosis. BioMed Central 2018-05-04 /pmc/articles/PMC5935939/ /pubmed/29728076 http://dx.doi.org/10.1186/s12896-018-0436-z Text en © The Author(s). 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Article Zhang, Danping Xie, Chengjie Wang, Rongzhi Yang, Qinghai Chen, Huiling Ling, Sumei Wang, Shihua Jia, Kunzhi Effective preparation of a monoclonal antibody against human chromogranin A for immunohistochemical diagnosis |
title | Effective preparation of a monoclonal antibody against human chromogranin A for immunohistochemical diagnosis |
title_full | Effective preparation of a monoclonal antibody against human chromogranin A for immunohistochemical diagnosis |
title_fullStr | Effective preparation of a monoclonal antibody against human chromogranin A for immunohistochemical diagnosis |
title_full_unstemmed | Effective preparation of a monoclonal antibody against human chromogranin A for immunohistochemical diagnosis |
title_short | Effective preparation of a monoclonal antibody against human chromogranin A for immunohistochemical diagnosis |
title_sort | effective preparation of a monoclonal antibody against human chromogranin a for immunohistochemical diagnosis |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5935939/ https://www.ncbi.nlm.nih.gov/pubmed/29728076 http://dx.doi.org/10.1186/s12896-018-0436-z |
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