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An optimized method for accurate quantification of cell migration using human small intestine cells

Quantifying the ability of a compound to modulate cell migration rate is a crucial part of many studies including those on chemotaxis, wound healing and cancer metastasis. Existing migration assays all have their strengths and weaknesses. The “scratch” assay is the most widely used because it seems...

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Autores principales: Nyegaard, Steffen, Christensen, Brian, Rasmussen, Jan Trige
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5779728/
https://www.ncbi.nlm.nih.gov/pubmed/29468115
http://dx.doi.org/10.1016/j.meteno.2016.03.002
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author Nyegaard, Steffen
Christensen, Brian
Rasmussen, Jan Trige
author_facet Nyegaard, Steffen
Christensen, Brian
Rasmussen, Jan Trige
author_sort Nyegaard, Steffen
collection PubMed
description Quantifying the ability of a compound to modulate cell migration rate is a crucial part of many studies including those on chemotaxis, wound healing and cancer metastasis. Existing migration assays all have their strengths and weaknesses. The “scratch” assay is the most widely used because it seems appealingly simple and inexpensive. However, the scratch assay has some important limitations, as the tool introducing the “wound” might injure/stress the boundary cells and/or harm underlying matrix coatings, which in both cases will affect cell migration. This described method is a Cell Exclusion Zone Assay, in which cell-free areas are created by growing cells around removable silicone stoppers. Upon appropriate staining with fluorescent dyes and microscopically visualizing the monolayers, the migration rate is then quantified by counting the cells (nuclei) intruding the void area left by the silicone insert. In the current study human small intestine epithelial cells were seeded on a physiological substrate matrix to produce collectively migrating monolayers. Different substrates were tested to determine the optimal surface for enterocyte adherence and migration and morphological changes monitored. Recombinant human epidermal growth factor and osteopontin purified from urine were tested to see if the established migration assay produces accurate and reliable migration data with human small intestine cells. The obtained data accurately confirmed that the two bioactive proteins modulate cellular migration in a dose-dependent manner. The presented assay can likely be converted for use with other adherent cell lines or substrate matrices and allows for high throughput, while cost is kept low and versatility high. Co-staining can be applied in order to assay for cell death, different cell types, cell stress and others allowing intricate analysis of migration rate of mixed populations and correction for cell viability.
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spelling pubmed-57797282018-02-21 An optimized method for accurate quantification of cell migration using human small intestine cells Nyegaard, Steffen Christensen, Brian Rasmussen, Jan Trige Metab Eng Commun Article Quantifying the ability of a compound to modulate cell migration rate is a crucial part of many studies including those on chemotaxis, wound healing and cancer metastasis. Existing migration assays all have their strengths and weaknesses. The “scratch” assay is the most widely used because it seems appealingly simple and inexpensive. However, the scratch assay has some important limitations, as the tool introducing the “wound” might injure/stress the boundary cells and/or harm underlying matrix coatings, which in both cases will affect cell migration. This described method is a Cell Exclusion Zone Assay, in which cell-free areas are created by growing cells around removable silicone stoppers. Upon appropriate staining with fluorescent dyes and microscopically visualizing the monolayers, the migration rate is then quantified by counting the cells (nuclei) intruding the void area left by the silicone insert. In the current study human small intestine epithelial cells were seeded on a physiological substrate matrix to produce collectively migrating monolayers. Different substrates were tested to determine the optimal surface for enterocyte adherence and migration and morphological changes monitored. Recombinant human epidermal growth factor and osteopontin purified from urine were tested to see if the established migration assay produces accurate and reliable migration data with human small intestine cells. The obtained data accurately confirmed that the two bioactive proteins modulate cellular migration in a dose-dependent manner. The presented assay can likely be converted for use with other adherent cell lines or substrate matrices and allows for high throughput, while cost is kept low and versatility high. Co-staining can be applied in order to assay for cell death, different cell types, cell stress and others allowing intricate analysis of migration rate of mixed populations and correction for cell viability. Elsevier 2016-03-15 /pmc/articles/PMC5779728/ /pubmed/29468115 http://dx.doi.org/10.1016/j.meteno.2016.03.002 Text en © 2016 The Authors http://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Article
Nyegaard, Steffen
Christensen, Brian
Rasmussen, Jan Trige
An optimized method for accurate quantification of cell migration using human small intestine cells
title An optimized method for accurate quantification of cell migration using human small intestine cells
title_full An optimized method for accurate quantification of cell migration using human small intestine cells
title_fullStr An optimized method for accurate quantification of cell migration using human small intestine cells
title_full_unstemmed An optimized method for accurate quantification of cell migration using human small intestine cells
title_short An optimized method for accurate quantification of cell migration using human small intestine cells
title_sort optimized method for accurate quantification of cell migration using human small intestine cells
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5779728/
https://www.ncbi.nlm.nih.gov/pubmed/29468115
http://dx.doi.org/10.1016/j.meteno.2016.03.002
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