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Correlative super-resolution fluorescence and electron microscopy using conventional fluorescent proteins in vacuo.

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Title: Correlative super-resolution fluorescence and electron microscopy using conventional fluorescent proteins in vacuo.
Authors: Peddie, Christopher J.1, Domart, Marie-Charlotte1, Snetkov, Xenia2, O'Toole, Peter3, Larijani, Banafshe4, Way, Michael2, Cox, Susan5, Collinson, Lucy M.1 lucy.collinson@crick.ac.uk
Source: Journal of Structural Biology. Aug2017, Vol. 199 Issue 2, p120-131. 12p.
Subjects: Microscopy, Electron microscopy, Fluorescent proteins, Vaccinia, Diglycerides
Abstract: Super-resolution light microscopy, correlative light and electron microscopy, and volume electron microscopy are revolutionising the way in which biological samples are examined and understood. Here, we combine these approaches to deliver super-accurate correlation of fluorescent proteins to cellular structures. We show that YFP and GFP have enhanced blinking properties when embedded in acrylic resin and imaged under partial vacuum, enabling in vacuo single molecule localisation microscopy. In conventional section-based correlative microscopy experiments, the specimen must be moved between imaging systems and/or further manipulated for optimal viewing. These steps can introduce undesirable alterations in the specimen, and complicate correlation between imaging modalities. We avoided these issues by using a scanning electron microscope with integrated optical microscope to acquire both localisation and electron microscopy images, which could then be precisely correlated. Collecting data from ultrathin sections also improved the axial resolution and signal-to-noise ratio of the raw localisation microscopy data. Expanding data collection across an array of sections will allow 3-dimensional correlation over unprecedented volumes. The performance of this technique is demonstrated on vaccinia virus (with YFP) and diacylglycerol in cellular membranes (with GFP). [ABSTRACT FROM AUTHOR]
Copyright of Journal of Structural Biology is the property of Academic Press Inc. and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)
Database: Engineering Source
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Header DbId: egs
DbLabel: Engineering Source
An: 124213455
AccessLevel: 6
PubType: Academic Journal
PubTypeId: academicJournal
PreciseRelevancyScore: 0
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  Data: Correlative super-resolution fluorescence and electron microscopy using conventional fluorescent proteins in vacuo.
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  Data: <searchLink fieldCode="AR" term="%22Peddie%2C+Christopher+J%2E%22">Peddie, Christopher J.</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Domart%2C+Marie-Charlotte%22">Domart, Marie-Charlotte</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Snetkov%2C+Xenia%22">Snetkov, Xenia</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22O'Toole%2C+Peter%22">O'Toole, Peter</searchLink><relatesTo>3</relatesTo><br /><searchLink fieldCode="AR" term="%22Larijani%2C+Banafshe%22">Larijani, Banafshe</searchLink><relatesTo>4</relatesTo><br /><searchLink fieldCode="AR" term="%22Way%2C+Michael%22">Way, Michael</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Cox%2C+Susan%22">Cox, Susan</searchLink><relatesTo>5</relatesTo><br /><searchLink fieldCode="AR" term="%22Collinson%2C+Lucy+M%2E%22">Collinson, Lucy M.</searchLink><relatesTo>1</relatesTo><i> lucy.collinson@crick.ac.uk</i>
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  Data: <searchLink fieldCode="JN" term="%22Journal+of+Structural+Biology%22">Journal of Structural Biology</searchLink>. Aug2017, Vol. 199 Issue 2, p120-131. 12p.
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  Data: Super-resolution light microscopy, correlative light and electron microscopy, and volume electron microscopy are revolutionising the way in which biological samples are examined and understood. Here, we combine these approaches to deliver super-accurate correlation of fluorescent proteins to cellular structures. We show that YFP and GFP have enhanced blinking properties when embedded in acrylic resin and imaged under partial vacuum, enabling in vacuo single molecule localisation microscopy. In conventional section-based correlative microscopy experiments, the specimen must be moved between imaging systems and/or further manipulated for optimal viewing. These steps can introduce undesirable alterations in the specimen, and complicate correlation between imaging modalities. We avoided these issues by using a scanning electron microscope with integrated optical microscope to acquire both localisation and electron microscopy images, which could then be precisely correlated. Collecting data from ultrathin sections also improved the axial resolution and signal-to-noise ratio of the raw localisation microscopy data. Expanding data collection across an array of sections will allow 3-dimensional correlation over unprecedented volumes. The performance of this technique is demonstrated on vaccinia virus (with YFP) and diacylglycerol in cellular membranes (with GFP). [ABSTRACT FROM AUTHOR]
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  Data: <i>Copyright of Journal of Structural Biology is the property of Academic Press Inc. and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract.</i> (Copyright applies to all Abstracts.)
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        Value: 10.1016/j.jsb.2017.05.013
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        Text: English
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        PageCount: 12
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        Type: general
      – SubjectFull: Electron microscopy
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      – SubjectFull: Fluorescent proteins
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      – SubjectFull: Vaccinia
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      – SubjectFull: Diglycerides
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      – TitleFull: Correlative super-resolution fluorescence and electron microscopy using conventional fluorescent proteins in vacuo.
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              Text: Aug2017
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