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Fluorescence Microscopy-Based Quantitation of GLUT4 Translocation: High Throughput or High Content?
Due to the global rise of type 2 diabetes mellitus (T2DM) in combination with insulin resistance, novel compounds to efficiently treat this pandemic disease are needed. Screening for compounds that induce the translocation of glucose transporter 4 (GLUT4) from the intracellular compartments to the p...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7662403/ https://www.ncbi.nlm.nih.gov/pubmed/33120934 http://dx.doi.org/10.3390/ijms21217964 |
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author | Stadlbauer, Verena Lanzerstorfer, Peter Neuhauser, Cathrina Weber, Florian Stübl, Flora Weber, Petra Wagner, Michael Plochberger, Birgit Wieser, Stefan Schneckenburger, Herbert Weghuber, Julian |
author_facet | Stadlbauer, Verena Lanzerstorfer, Peter Neuhauser, Cathrina Weber, Florian Stübl, Flora Weber, Petra Wagner, Michael Plochberger, Birgit Wieser, Stefan Schneckenburger, Herbert Weghuber, Julian |
author_sort | Stadlbauer, Verena |
collection | PubMed |
description | Due to the global rise of type 2 diabetes mellitus (T2DM) in combination with insulin resistance, novel compounds to efficiently treat this pandemic disease are needed. Screening for compounds that induce the translocation of glucose transporter 4 (GLUT4) from the intracellular compartments to the plasma membrane in insulin-sensitive tissues is an innovative strategy. Here, we compared the applicability of three fluorescence microscopy-based assays optimized for the quantitation of GLUT4 translocation in simple cell systems. An objective-type scanning total internal reflection fluorescence (TIRF) microscopy approach was shown to have high sensitivity but only moderate throughput. Therefore, we implemented a prism-type TIR reader for the simultaneous analysis of large cell populations grown in adapted microtiter plates. This approach was found to be high throughput and have sufficient sensitivity for the characterization of insulin mimetic compounds in live cells. Finally, we applied confocal microscopy to giant plasma membrane vesicles (GPMVs) formed from GLUT4-expressing cells. While this assay has only limited throughput, it offers the advantage of being less sensitive to insulin mimetic compounds with high autofluorescence. In summary, the combined implementation of different fluorescence microscopy-based approaches enables the quantitation of GLUT4 translocation with high throughput and high content. |
format | Online Article Text |
id | pubmed-7662403 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-76624032020-11-14 Fluorescence Microscopy-Based Quantitation of GLUT4 Translocation: High Throughput or High Content? Stadlbauer, Verena Lanzerstorfer, Peter Neuhauser, Cathrina Weber, Florian Stübl, Flora Weber, Petra Wagner, Michael Plochberger, Birgit Wieser, Stefan Schneckenburger, Herbert Weghuber, Julian Int J Mol Sci Article Due to the global rise of type 2 diabetes mellitus (T2DM) in combination with insulin resistance, novel compounds to efficiently treat this pandemic disease are needed. Screening for compounds that induce the translocation of glucose transporter 4 (GLUT4) from the intracellular compartments to the plasma membrane in insulin-sensitive tissues is an innovative strategy. Here, we compared the applicability of three fluorescence microscopy-based assays optimized for the quantitation of GLUT4 translocation in simple cell systems. An objective-type scanning total internal reflection fluorescence (TIRF) microscopy approach was shown to have high sensitivity but only moderate throughput. Therefore, we implemented a prism-type TIR reader for the simultaneous analysis of large cell populations grown in adapted microtiter plates. This approach was found to be high throughput and have sufficient sensitivity for the characterization of insulin mimetic compounds in live cells. Finally, we applied confocal microscopy to giant plasma membrane vesicles (GPMVs) formed from GLUT4-expressing cells. While this assay has only limited throughput, it offers the advantage of being less sensitive to insulin mimetic compounds with high autofluorescence. In summary, the combined implementation of different fluorescence microscopy-based approaches enables the quantitation of GLUT4 translocation with high throughput and high content. MDPI 2020-10-27 /pmc/articles/PMC7662403/ /pubmed/33120934 http://dx.doi.org/10.3390/ijms21217964 Text en © 2020 by the authors. 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 (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Stadlbauer, Verena Lanzerstorfer, Peter Neuhauser, Cathrina Weber, Florian Stübl, Flora Weber, Petra Wagner, Michael Plochberger, Birgit Wieser, Stefan Schneckenburger, Herbert Weghuber, Julian Fluorescence Microscopy-Based Quantitation of GLUT4 Translocation: High Throughput or High Content? |
title | Fluorescence Microscopy-Based Quantitation of GLUT4 Translocation: High Throughput or High Content? |
title_full | Fluorescence Microscopy-Based Quantitation of GLUT4 Translocation: High Throughput or High Content? |
title_fullStr | Fluorescence Microscopy-Based Quantitation of GLUT4 Translocation: High Throughput or High Content? |
title_full_unstemmed | Fluorescence Microscopy-Based Quantitation of GLUT4 Translocation: High Throughput or High Content? |
title_short | Fluorescence Microscopy-Based Quantitation of GLUT4 Translocation: High Throughput or High Content? |
title_sort | fluorescence microscopy-based quantitation of glut4 translocation: high throughput or high content? |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7662403/ https://www.ncbi.nlm.nih.gov/pubmed/33120934 http://dx.doi.org/10.3390/ijms21217964 |
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