Stimulated emission depletion (STED) nanoscopy of a fluorescent protein-labeled organelle inside a living cell
We demonstrate far-field optical imaging with subdiffraction resolution of the endoplasmic reticulum (ER) in the interior of a living mammalian cell. The diffraction barrier is overcome by applying stimulated emission depletion (STED) on a yellow fluorescent protein tag. Imaging individual structura...
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| Veröffentlicht in: | Proceedings of the National Academy of Sciences - PNAS 2008-09, Vol.105 (38), p.14271-14276 |
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| container_issue | 38 |
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| container_title | Proceedings of the National Academy of Sciences - PNAS |
| container_volume | 105 |
| creator | Hein, Birka Willig, Katrin I Hell, Stefan W |
| description | We demonstrate far-field optical imaging with subdiffraction resolution of the endoplasmic reticulum (ER) in the interior of a living mammalian cell. The diffraction barrier is overcome by applying stimulated emission depletion (STED) on a yellow fluorescent protein tag. Imaging individual structural elements of the ER revealed a focal plane (x, y) resolution of |
| doi_str_mv | 10.1073/pnas.0807705105 |
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The diffraction barrier is overcome by applying stimulated emission depletion (STED) on a yellow fluorescent protein tag. Imaging individual structural elements of the ER revealed a focal plane (x, y) resolution of <50 nm inside the living cell, corresponding to a 4-fold improvement over that of a confocal microscope and a 16-fold reduction in the focal-spot cross-sectional area. A similar gain in resolution is realized with both pulsed- and continuous-wave laser illumination. Images of highly convoluted parts of the ER reveal a similar resolution improvement in 3D optical sectioning by a factor of 3 along the optic axis (z). Time-lapse STED recordings document morphological changes of the ER over time. 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Thus, nanoscale 3D imaging of organelles in the interior of living cells greatly expands the scope of light microscopy in cell biology.</description><subject>Animals</subject><subject>Bacterial Proteins - genetics</subject><subject>Bacterial Proteins - metabolism</subject><subject>Cell Line</subject><subject>Cell Survival</subject><subject>Cells</subject><subject>Cells - cytology</subject><subject>Cellular biology</subject><subject>Diffraction</subject><subject>Endoplasmic Reticulum - metabolism</subject><subject>Endoplasmic Reticulum - ultrastructure</subject><subject>Fluorescence</subject><subject>Image resolution</subject><subject>Imaging</subject><subject>Laser beams</subject><subject>Lasers</subject><subject>Luminescent Proteins - genetics</subject><subject>Luminescent Proteins - metabolism</subject><subject>Mammals</subject><subject>Microscopes</subject><subject>Microscopy</subject><subject>Microscopy, Confocal</subject><subject>Microscopy, Fluorescence - economics</subject><subject>Microscopy, Fluorescence - instrumentation</subject><subject>Microscopy, Fluorescence - methods</subject><subject>Microscopy, Fluorescence - standards</subject><subject>Molecules</subject><subject>Physical Sciences</subject><subject>Potoroidae</subject><subject>Proteins</subject><subject>Stimulated emission</subject><subject>Wave diffraction</subject><issn>0027-8424</issn><issn>1091-6490</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2008</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkc1v1DAUxCNERZfCmRNgcUD0kPb5K7YvSFUpBakSh23PljdxFq-8drCTqv3vcbSrLnDhZMvze6M3nqp6g-EMg6DnQzD5DCQIARwDf1YtMChcN0zB82oBQEQtGWHH1cucNwCguIQX1TGWQjUNsEUVlqPbTt6MtkN263J2MaDODt6O8-3T8vbqyykKJsTcxuERxR4Z1PspJptbG0Y0pDhaF2pvVtYXk5jWJljvLXIhu84W3Lt7F9aoLa-vqqPe-Gxf78-T6u7r1e3lt_rmx_X3y4ubuuVSjTVtYSV60lOMleUKK96Z1lLFKaUNlQRD27WKyB5TkF2LVcelFKJvWAOy4YKeVJ93vsO02tpu3jQZr4fktiY96mic_lsJ7qdex3tNOJWM42LwcW-Q4q_J5lGXz5kTlHBxyrpRXDHKVQE__ANu4pRCCacJYFo4AQU630Ftijkn2z9tgkHPReq5SH0osky8-zPAgd83V4DTPTBPHuy4plJjRgTW_eT9aB_GwqL_sAV5u0M2eYzpiSGcNYxgWfT3O703UZt1clnfLeeAgDkjtAH6G9QDxV8</recordid><startdate>20080923</startdate><enddate>20080923</enddate><creator>Hein, Birka</creator><creator>Willig, Katrin I</creator><creator>Hell, Stefan W</creator><general>National Academy of Sciences</general><general>National Acad Sciences</general><scope>FBQ</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QG</scope><scope>7QL</scope><scope>7QP</scope><scope>7QR</scope><scope>7SN</scope><scope>7SS</scope><scope>7T5</scope><scope>7TK</scope><scope>7TM</scope><scope>7TO</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20080923</creationdate><title>Stimulated emission depletion (STED) nanoscopy of a fluorescent protein-labeled organelle inside a living cell</title><author>Hein, Birka ; 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| subjects | Animals Bacterial Proteins - genetics Bacterial Proteins - metabolism Cell Line Cell Survival Cells Cells - cytology Cellular biology Diffraction Endoplasmic Reticulum - metabolism Endoplasmic Reticulum - ultrastructure Fluorescence Image resolution Imaging Laser beams Lasers Luminescent Proteins - genetics Luminescent Proteins - metabolism Mammals Microscopes Microscopy Microscopy, Confocal Microscopy, Fluorescence - economics Microscopy, Fluorescence - instrumentation Microscopy, Fluorescence - methods Microscopy, Fluorescence - standards Molecules Physical Sciences Potoroidae Proteins Stimulated emission Wave diffraction |
| title | Stimulated emission depletion (STED) nanoscopy of a fluorescent protein-labeled organelle inside a living cell |
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