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Improvements for Tissue-Chopping-Based Immunofluorescence Staining Method of Chloroplast Proteins

Immunofluorescence staining is a very common method for the subcellular localization study of proteins. A tissue-chopping-based immunofluorescence staining method for chloroplast proteins overcomes the restriction of plant cell wall, makes the operation simpler, and uses less experimental materials....

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Detalles Bibliográficos
Autores principales: Wang, Lulu, Chen, Yajuan, Niu, Di, Tang, Mingdong, An, Jinjie, Xue, Shanshan, Liu, Xiaomin, Gao, Hongbo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9963192/
https://www.ncbi.nlm.nih.gov/pubmed/36840189
http://dx.doi.org/10.3390/plants12040841
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author Wang, Lulu
Chen, Yajuan
Niu, Di
Tang, Mingdong
An, Jinjie
Xue, Shanshan
Liu, Xiaomin
Gao, Hongbo
author_facet Wang, Lulu
Chen, Yajuan
Niu, Di
Tang, Mingdong
An, Jinjie
Xue, Shanshan
Liu, Xiaomin
Gao, Hongbo
author_sort Wang, Lulu
collection PubMed
description Immunofluorescence staining is a very common method for the subcellular localization study of proteins. A tissue-chopping-based immunofluorescence staining method for chloroplast proteins overcomes the restriction of plant cell wall, makes the operation simpler, and uses less experimental materials. Here we provide some improvements for this method. We found that the stained tissues can be directly observed with a confocal microscope without tissue lysis. Samples maintained at a low temperature (0–4 °C) throughout the process can reduce the intensity of chlorophyll autofluorescence and the background signal. A low temperature is also good for the storage of the sample. Fluorescence signal of the stained samples can be kept for several weeks if they are stored at −20 °C. FtsZ is an essential component of the chloroplast division apparatus. We demonstrated this method with the immunofluorescence staining of FtsZ1 in wildtype Arabidopsis and some chloroplast division mutants. We also successfully tested this method by the immunofluorescence staining of FtsZ1 in many other plants, including woody plants. With these procedures, the performance of tissue-chopping-based immunofluorescence staining method are further improved.
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spelling pubmed-99631922023-02-26 Improvements for Tissue-Chopping-Based Immunofluorescence Staining Method of Chloroplast Proteins Wang, Lulu Chen, Yajuan Niu, Di Tang, Mingdong An, Jinjie Xue, Shanshan Liu, Xiaomin Gao, Hongbo Plants (Basel) Communication Immunofluorescence staining is a very common method for the subcellular localization study of proteins. A tissue-chopping-based immunofluorescence staining method for chloroplast proteins overcomes the restriction of plant cell wall, makes the operation simpler, and uses less experimental materials. Here we provide some improvements for this method. We found that the stained tissues can be directly observed with a confocal microscope without tissue lysis. Samples maintained at a low temperature (0–4 °C) throughout the process can reduce the intensity of chlorophyll autofluorescence and the background signal. A low temperature is also good for the storage of the sample. Fluorescence signal of the stained samples can be kept for several weeks if they are stored at −20 °C. FtsZ is an essential component of the chloroplast division apparatus. We demonstrated this method with the immunofluorescence staining of FtsZ1 in wildtype Arabidopsis and some chloroplast division mutants. We also successfully tested this method by the immunofluorescence staining of FtsZ1 in many other plants, including woody plants. With these procedures, the performance of tissue-chopping-based immunofluorescence staining method are further improved. MDPI 2023-02-13 /pmc/articles/PMC9963192/ /pubmed/36840189 http://dx.doi.org/10.3390/plants12040841 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 Communication
Wang, Lulu
Chen, Yajuan
Niu, Di
Tang, Mingdong
An, Jinjie
Xue, Shanshan
Liu, Xiaomin
Gao, Hongbo
Improvements for Tissue-Chopping-Based Immunofluorescence Staining Method of Chloroplast Proteins
title Improvements for Tissue-Chopping-Based Immunofluorescence Staining Method of Chloroplast Proteins
title_full Improvements for Tissue-Chopping-Based Immunofluorescence Staining Method of Chloroplast Proteins
title_fullStr Improvements for Tissue-Chopping-Based Immunofluorescence Staining Method of Chloroplast Proteins
title_full_unstemmed Improvements for Tissue-Chopping-Based Immunofluorescence Staining Method of Chloroplast Proteins
title_short Improvements for Tissue-Chopping-Based Immunofluorescence Staining Method of Chloroplast Proteins
title_sort improvements for tissue-chopping-based immunofluorescence staining method of chloroplast proteins
topic Communication
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9963192/
https://www.ncbi.nlm.nih.gov/pubmed/36840189
http://dx.doi.org/10.3390/plants12040841
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