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A Semi-Automatic Method for the Quantification of Astrocyte Number and Branching in Bulk Immunohistochemistry Images
Immunohistochemical staining of cell and molecular targets in brain samples is a powerful tool that can provide valuable information on neurological mechanisms. However, post-processing of photomicrographs acquired after 3,3′-Diaminobenzidine (DAB) staining is particularly challenging due to the com...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10003611/ https://www.ncbi.nlm.nih.gov/pubmed/36901939 http://dx.doi.org/10.3390/ijms24054508 |
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author | Marques, Sandra I. Carmo, Helena Carvalho, Félix Sá, Susana I. Silva, João Pedro |
author_facet | Marques, Sandra I. Carmo, Helena Carvalho, Félix Sá, Susana I. Silva, João Pedro |
author_sort | Marques, Sandra I. |
collection | PubMed |
description | Immunohistochemical staining of cell and molecular targets in brain samples is a powerful tool that can provide valuable information on neurological mechanisms. However, post-processing of photomicrographs acquired after 3,3′-Diaminobenzidine (DAB) staining is particularly challenging due to the complexity associated with the size, samples number, analyzed targets, image quality, and even the subjectivity inherent to the analysis by different users. Conventionally, this analysis relies on the manual quantification of distinct parameters (e.g., the number and size of cells and the number and length of cell branching) in a large set of images. These represent extremely time-consuming and complex tasks, defaulting the processing of high amounts of information. Here we describe an improved semi-automatic method to quantify glial fibrillary acidic protein (GFAP)-labelled astrocytes in immunohistochemistry images of rat brains, at magnifications as low as 20×. This method is a straightforward adaptation of the Young & Morrison method, using ImageJ’s plugin Skeletonize, coupled with intuitive data processing in datasheet-based software. It allows swifter and more efficient post-processing of brain tissue samples, regarding astrocyte size and number quantification, the total area occupied, as well as astrocyte branching and branch length (indicators of astrocyte activation), thus contributing to better understand the possible inflammatory response developed by astrocytes. |
format | Online Article Text |
id | pubmed-10003611 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-100036112023-03-11 A Semi-Automatic Method for the Quantification of Astrocyte Number and Branching in Bulk Immunohistochemistry Images Marques, Sandra I. Carmo, Helena Carvalho, Félix Sá, Susana I. Silva, João Pedro Int J Mol Sci Protocol Immunohistochemical staining of cell and molecular targets in brain samples is a powerful tool that can provide valuable information on neurological mechanisms. However, post-processing of photomicrographs acquired after 3,3′-Diaminobenzidine (DAB) staining is particularly challenging due to the complexity associated with the size, samples number, analyzed targets, image quality, and even the subjectivity inherent to the analysis by different users. Conventionally, this analysis relies on the manual quantification of distinct parameters (e.g., the number and size of cells and the number and length of cell branching) in a large set of images. These represent extremely time-consuming and complex tasks, defaulting the processing of high amounts of information. Here we describe an improved semi-automatic method to quantify glial fibrillary acidic protein (GFAP)-labelled astrocytes in immunohistochemistry images of rat brains, at magnifications as low as 20×. This method is a straightforward adaptation of the Young & Morrison method, using ImageJ’s plugin Skeletonize, coupled with intuitive data processing in datasheet-based software. It allows swifter and more efficient post-processing of brain tissue samples, regarding astrocyte size and number quantification, the total area occupied, as well as astrocyte branching and branch length (indicators of astrocyte activation), thus contributing to better understand the possible inflammatory response developed by astrocytes. MDPI 2023-02-24 /pmc/articles/PMC10003611/ /pubmed/36901939 http://dx.doi.org/10.3390/ijms24054508 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Protocol Marques, Sandra I. Carmo, Helena Carvalho, Félix Sá, Susana I. Silva, João Pedro A Semi-Automatic Method for the Quantification of Astrocyte Number and Branching in Bulk Immunohistochemistry Images |
title | A Semi-Automatic Method for the Quantification of Astrocyte Number and Branching in Bulk Immunohistochemistry Images |
title_full | A Semi-Automatic Method for the Quantification of Astrocyte Number and Branching in Bulk Immunohistochemistry Images |
title_fullStr | A Semi-Automatic Method for the Quantification of Astrocyte Number and Branching in Bulk Immunohistochemistry Images |
title_full_unstemmed | A Semi-Automatic Method for the Quantification of Astrocyte Number and Branching in Bulk Immunohistochemistry Images |
title_short | A Semi-Automatic Method for the Quantification of Astrocyte Number and Branching in Bulk Immunohistochemistry Images |
title_sort | semi-automatic method for the quantification of astrocyte number and branching in bulk immunohistochemistry images |
topic | Protocol |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10003611/ https://www.ncbi.nlm.nih.gov/pubmed/36901939 http://dx.doi.org/10.3390/ijms24054508 |
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