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...
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MDPI AG
2021-03-01
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| Series: | Sensors |
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| Online Access: | https://www.mdpi.com/1424-8220/21/6/2010 |
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| author | Isotta Cainero Elena Cerutti Mario Faretta Gaetano Ivan Dellino Pier Giuseppe Pelicci Alberto Diaspro Luca Lanzanò |
| author_facet | Isotta Cainero Elena Cerutti Mario Faretta Gaetano Ivan Dellino Pier Giuseppe Pelicci Alberto Diaspro Luca Lanzanò |
| author_sort | Isotta Cainero |
| collection | DOAJ |
| 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. |
| first_indexed | 2024-03-10T13:18:28Z |
| format | Article |
| id | doaj.art-83a6ffc3f8ec45fb8ff1dd4054e7817e |
| institution | Directory Open Access Journal |
| issn | 1424-8220 |
| language | English |
| last_indexed | 2024-03-10T13:18:28Z |
| publishDate | 2021-03-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Sensors |
| spelling | doaj.art-83a6ffc3f8ec45fb8ff1dd4054e7817e2023-11-21T10:14:41ZengMDPI AGSensors1424-82202021-03-01216201010.3390/s21062010Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS)Isotta Cainero0Elena Cerutti1Mario Faretta2Gaetano Ivan Dellino3Pier Giuseppe Pelicci4Alberto Diaspro5Luca Lanzanò6Nanoscopy and NIC@IIT, CHT Erzelli, Istituto Italiano di Tecnologia, Via Enrico Melen 83, Building B, 16152 Genoa, ItalyNanoscopy and NIC@IIT, CHT Erzelli, Istituto Italiano di Tecnologia, Via Enrico Melen 83, Building B, 16152 Genoa, ItalyDepartment of Experimental Oncology, IEO, European Institute of Oncology IRCCS, 20100 Milan, ItalyDepartment of Experimental Oncology, IEO, European Institute of Oncology IRCCS, 20100 Milan, ItalyDepartment of Experimental Oncology, IEO, European Institute of Oncology IRCCS, 20100 Milan, ItalyNanoscopy and NIC@IIT, CHT Erzelli, Istituto Italiano di Tecnologia, Via Enrico Melen 83, Building B, 16152 Genoa, ItalyNanoscopy and NIC@IIT, CHT Erzelli, Istituto Italiano di Tecnologia, Via Enrico Melen 83, Building B, 16152 Genoa, ItalySince 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.https://www.mdpi.com/1424-8220/21/6/2010super-resolution microscopystructured illumination microscopySIMimage correlation spectroscopyimage cross-correlation spectroscopyICCS |
| spellingShingle | Isotta Cainero Elena Cerutti Mario Faretta Gaetano Ivan Dellino Pier Giuseppe Pelicci Alberto Diaspro Luca Lanzanò Measuring Nanoscale Distances by Structured Illumination Microscopy and Image Cross-Correlation Spectroscopy (SIM-ICCS) Sensors super-resolution microscopy structured illumination microscopy SIM image correlation spectroscopy image cross-correlation spectroscopy 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 | super-resolution microscopy structured illumination microscopy SIM image correlation spectroscopy image cross-correlation spectroscopy ICCS |
| url | https://www.mdpi.com/1424-8220/21/6/2010 |
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