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Coceano, Giovanna
Publications (4 of 4) Show all publications
Dreier, J., Castello, M., Coceano, G., Caceres, R., Plastino, J., Vicidomini, G. & Testa, I. (2019). Smart scanning for low-illumination and fast RESOLFT nanoscopy in vivo. Nature Communications, 10, Article ID 556.
Open this publication in new window or tab >>Smart scanning for low-illumination and fast RESOLFT nanoscopy in vivo
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2019 (English)In: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 10, article id 556Article in journal (Refereed) Published
Abstract [en]

RESOLFT fluorescence nanoscopy can nowadays image details far beyond the diffraction limit. However, signal to noise ratio (SNR) and temporal resolution are still a concern, especially deep inside living cells and organisms. In this work, we developed a non-deterministic scanning approach based on a real-time feedback system which speeds up the acquisition up to 6-fold and decreases the light dose by 70-90% for in vivo imaging. Also, we extended the information content of the images by acquiring the complete temporal evolution of the fluorescence generated by reversible switchable fluorescent proteins. This generates a series of images with different spatial resolution and SNR, from conventional to RESOLFT images, which combined through a multi-image deconvolution algorithm further enhances the effective resolution. We reported nanoscale imaging of organelles up to 35 Hz and actin dynamics during an invasion process at a depth of 20-30 mu m inside a living Caenorhabditis elegans worm.

Place, publisher, year, edition, pages
NATURE PUBLISHING GROUP, 2019
National Category
Physical Sciences
Identifiers
urn:nbn:se:kth:diva-244106 (URN)10.1038/s41467-019-08442-4 (DOI)000457443400001 ()30710076 (PubMedID)2-s2.0-85060941002 (Scopus ID)
Funder
Swedish Research Council
Note

QC 20190219

Available from: 2019-02-19 Created: 2019-02-19 Last updated: 2019-02-19Bibliographically approved
Masullo, L. A., Boden, A., Pennacchietti, F., Coceano, G., Ratz, M. & Testa, I. (2018). Enhanced photon collection enables four dimensional fluorescence nanoscopy of living systems. Nature Communications, 9, Article ID 3281.
Open this publication in new window or tab >>Enhanced photon collection enables four dimensional fluorescence nanoscopy of living systems
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2018 (English)In: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 9, article id 3281Article in journal (Refereed) Published
Abstract [en]

The theoretically unlimited spatial resolution of fluorescence nanoscopy often comes at the expense of time, contrast and increased dose of energy for recording. Here, we developed MoNaLISA, for Molecular Nanoscale Live Imaging with Sectioning Ability, a nanoscope capable of imaging structures at a scale of 45-65 nm within the entire cell volume at low light intensities (W-kW cm(-2)). Our approach, based on reversibly switchable fluorescent proteins, features three distinctly modulated illumination patterns crafted and combined to gain fluorescence ON-OFF switching cycles and image contrast. By maximizing the detected photon flux, MoNaLISA enables prolonged (40-50 frames) and large (50 x 50 mu m(2)) recordings at 0.3-1.3 Hz with enhanced optical sectioning ability. We demonstrate the general use of our approach by 4D imaging of organelles and fine structures in epithelial human cells, colonies of mouse embryonic stem cells, brain cells, and organotypic tissues.

Place, publisher, year, edition, pages
NATURE PUBLISHING GROUP, 2018
National Category
Other Physics Topics
Identifiers
urn:nbn:se:kth:diva-234173 (URN)10.1038/s41467-018-05799-w (DOI)000441768300012 ()30115928 (PubMedID)2-s2.0-85051531804 (Scopus ID)
Note

QC 20181017

Available from: 2018-10-17 Created: 2018-10-17 Last updated: 2018-12-10Bibliographically approved
Richter, K. N., Revelo, N. H., Seitz, K. J., Helm, M. S., Sarkar, D., Saleeb, R. S., . . . Rizzoli, S. O. (2018). Glyoxal as an alternative fixative to formaldehyde in immunostaining and super-resolution microscopy. EMBO Journal, 37(1), 139-159
Open this publication in new window or tab >>Glyoxal as an alternative fixative to formaldehyde in immunostaining and super-resolution microscopy
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2018 (English)In: EMBO Journal, ISSN 0261-4189, E-ISSN 1460-2075, Vol. 37, no 1, p. 139-159Article in journal (Refereed) Published
Abstract [en]

Paraformaldehyde (PFA) is the most commonly used fixative for immunostaining of cells, but has been associated with various problems, ranging from loss of antigenicity to changes in morphology during fixation. We show here that the small dialdehyde glyoxal can successfully replace PFA. Despite being less toxic than PFA, and, as most aldehydes, likely usable as a fixative, glyoxal has not yet been systematically tried in modern fluorescence microscopy. Here, we tested and optimized glyoxal fixation and surprisingly found it to be more efficient than PFA-based protocols. Glyoxal acted faster than PFA, cross-linked proteins more effectively, and improved the preservation of cellular morphology. We validated glyoxal fixation in multiple laboratories against different PFA-based protocols and confirmed that it enabled better immunostainings for a majority of the targets. Our data therefore support that glyoxal can be a valuable alternative to PFA for immunostaining.

Place, publisher, year, edition, pages
WILEY, 2018
Keywords
fixation, glyoxal, immunocytochemistry, PFA, super-resolution Microscopy
National Category
Biological Sciences
Identifiers
urn:nbn:se:kth:diva-221945 (URN)10.15252/embj.201695709 (DOI)000419328600010 ()29146773 (PubMedID)2-s2.0-85034102627 (Scopus ID)
Note

QC 20180130

Available from: 2018-01-30 Created: 2018-01-30 Last updated: 2018-01-30Bibliographically approved
Terriac, E., Coceano, G., Mavajian, Z., Hageman, T. A. G., Christ, A. F., Testa, I., . . . Gad, A. K. B. (2017). Vimentin Levels and Serine 71 Phosphorylation in the Control of Cell-Matrix Adhesions, Migration Speed, and Shape of Transformed Human Fibroblasts. CELLS, 6(1), Article ID 2.
Open this publication in new window or tab >>Vimentin Levels and Serine 71 Phosphorylation in the Control of Cell-Matrix Adhesions, Migration Speed, and Shape of Transformed Human Fibroblasts
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2017 (English)In: CELLS, ISSN 2073-4409, Vol. 6, no 1, article id 2Article in journal (Refereed) Published
Abstract [en]

Metastasizing tumor cells show increased expression of the intermediate filament (IF) protein vimentin, which has been used to diagnose invasive tumors for decades. Recent observations indicate that vimentin is not only a passive marker for carcinoma, but may also induce tumor cell invasion. To clarify how vimentin IFs control cell adhesions and migration, we analyzed the nanoscale (30-50 nm) spatial organization of vimentin IFs and cell-matrix adhesions in metastatic fibroblast cells, using three-color stimulated emission depletion (STED) microscopy. We also studied whether wild-type and phospho-deficient or -mimicking mutants of vimentin changed the size and lifetime of focal adhesions (FAs), cell shape, and cell migration, using live-cell total internal reflection imaging and confocal microscopy. We observed that vimentin exists in fragments of different lengths. Short fragments were mostly the size of a unit-length filament and were mainly localized close to small cell-matrix adhesions. Long vimentin filaments were found in the proximity of large FAs. Vimentin expression in these cells caused a reduction in FAs size and an elongated cell shape, but did not affect FA lifetime, or the speed or directionality of cell migration. Expression of a phospho-mimicking mutant (S71D) of vimentin increased the speed of cell migration. Taken together, our results suggest that in highly migratory, transformed mesenchymal cells, vimentin levels control the cell shape and FA size, but not cell migration, which instead is linked to the phosphorylation status of S71 vimentin. These observations are consistent with the possibility that not only levels, but also the assembly status of vimentin control cell migration.

Place, publisher, year, edition, pages
MDPI AG, 2017
Keywords
focal adhesions, vimentin, cell migration, total internal reflection (TIRF) microscopy, stimulated emission depletion (STED) microscopy
National Category
Cell Biology
Identifiers
urn:nbn:se:kth:diva-221038 (URN)10.3390/cells6010002 (DOI)000418787000001 ()2-s2.0-85050579240 (Scopus ID)
Funder
Swedish Cancer Society
Note

QC 20180112

Available from: 2018-01-12 Created: 2018-01-12 Last updated: 2018-10-16Bibliographically approved
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