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Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS)
Since the introduction of super-resolution microscopy, there has been growing interest in quantifying the nanoscale spatial distributions of fluorescent probes to better understand cellular processes and their interactions. One way to check if distributions are correlated or not is to perform coloca...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8001887/ https://www.ncbi.nlm.nih.gov/pubmed/33809144 http://dx.doi.org/10.3390/s21062010 |
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author | Cainero, Isotta Cerutti, Elena Faretta, Mario Dellino, Gaetano Ivan Pelicci, Pier Giuseppe Diaspro, Alberto Lanzanò, Luca |
author_facet | Cainero, Isotta Cerutti, Elena Faretta, Mario Dellino, Gaetano Ivan Pelicci, Pier Giuseppe Diaspro, Alberto Lanzanò, Luca |
author_sort | Cainero, Isotta |
collection | PubMed |
description | Since the introduction of super-resolution microscopy, there has been growing interest in quantifying the nanoscale spatial distributions of fluorescent probes to better understand cellular processes and their interactions. One way to check if distributions are correlated or not is to perform colocalization analysis of multi-color acquisitions. Among all the possible methods available to study and quantify the colocalization between multicolor images, there is image cross-correlation spectroscopy (ICCS). The main advantage of ICCS, in comparison with other co-localization techniques, is that it does not require pre-segmentation of the sample into single objects. Here we show that the combination of structured illumination microscopy (SIM) with ICCS (SIM-ICCS) is a simple approach to quantify colocalization and measure nanoscale distances from multi-color SIM images. We validate the SIM-ICCS analysis on SIM images of optical nanorulers, DNA-origami-based model samples containing fluorophores of different colors at a distance of 80 nm. The SIM-ICCS analysis is compared with an object-based analysis performed on the same samples. Finally, we show that SIM-ICCS can be used to quantify the nanoscale spatial distribution of functional nuclear sites in fixed cells. |
format | Online Article Text |
id | pubmed-8001887 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-80018872021-03-28 Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS) Cainero, Isotta Cerutti, Elena Faretta, Mario Dellino, Gaetano Ivan Pelicci, Pier Giuseppe Diaspro, Alberto Lanzanò, Luca Sensors (Basel) Article Since the introduction of super-resolution microscopy, there has been growing interest in quantifying the nanoscale spatial distributions of fluorescent probes to better understand cellular processes and their interactions. One way to check if distributions are correlated or not is to perform colocalization analysis of multi-color acquisitions. Among all the possible methods available to study and quantify the colocalization between multicolor images, there is image cross-correlation spectroscopy (ICCS). The main advantage of ICCS, in comparison with other co-localization techniques, is that it does not require pre-segmentation of the sample into single objects. Here we show that the combination of structured illumination microscopy (SIM) with ICCS (SIM-ICCS) is a simple approach to quantify colocalization and measure nanoscale distances from multi-color SIM images. We validate the SIM-ICCS analysis on SIM images of optical nanorulers, DNA-origami-based model samples containing fluorophores of different colors at a distance of 80 nm. The SIM-ICCS analysis is compared with an object-based analysis performed on the same samples. Finally, we show that SIM-ICCS can be used to quantify the nanoscale spatial distribution of functional nuclear sites in fixed cells. MDPI 2021-03-12 /pmc/articles/PMC8001887/ /pubmed/33809144 http://dx.doi.org/10.3390/s21062010 Text en © 2021 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 Cainero, Isotta Cerutti, Elena Faretta, Mario Dellino, Gaetano Ivan Pelicci, Pier Giuseppe Diaspro, Alberto Lanzanò, Luca Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS) |
title | Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS) |
title_full | Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS) |
title_fullStr | Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS) |
title_full_unstemmed | Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS) |
title_short | Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS) |
title_sort | measuring nanoscale distances by structured illumination microscopy and image cross-correlation spectroscopy (sim-iccs) |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8001887/ https://www.ncbi.nlm.nih.gov/pubmed/33809144 http://dx.doi.org/10.3390/s21062010 |
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