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  High-throughput ultrastructure screening using electron microscopy and fluorescent barcoding

Bykov, Y. S., Cohen, N., Gabrielli, N., Manenschijn, H., Welsch, S., Chlanda, P., et al. (2019). High-throughput ultrastructure screening using electron microscopy and fluorescent barcoding. Journal of Cell Biology, 218(8), 2797-2811. doi:10.1083/jcb.201812081.

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 Creators:
Bykov, Y. S., Author
Cohen, N., Author
Gabrielli, N., Author
Manenschijn, H., Author
Welsch, S., Author
Chlanda, P., Author
Kukulski, W., Author
Patil, K. R., Author
Schuldiner, M., Author
Briggs, John A. G.1, 2, Author           
Affiliations:
1European Molecular Biology Laboratory, External Organizations, ou_3346677              
2MRC Laboratory of Molecular Biology, External Organizations, Cambridge, GB, ou_3346673              

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Free keywords: correlated fluorescence yeast cell localization protein gene tomography libraries light form Cell Biology
 Abstract: Genetic screens using high-throughput fluorescent microscopes have generated large datasets, contributing many cell biological insights. Such approaches cannot tackle questions requiring knowledge of ultrastructure below the resolution limit of fluorescent microscopy. Electron microscopy (EM) reveals detailed cellular ultrastructure but requires time-consuming sample preparation, limiting throughput. Here we describe a robust method for screening by high-throughput EM. Our approach uses combinations of fluorophores as barcodes to uniquely mark each cell type in mixed populations and correlative light and EM (CLEM) to read the barcode of each cell before it is imaged by EM. Coupled with an easy-to-use software workflow for correlation, segmentation, and computer image analysis, our method, called "MultiCLEM," allows us to extract and analyze multiple cell populations from each EM sample preparation. We demonstrate several uses for MultiCLEM with 15 different yeast variants. The methodology is not restricted to yeast, can be scaled to higher throughput, and can be used in multiple ways to enable EM to become a powerful screening technique.

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Language(s): eng - English
 Dates: 2019
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Type: -
 Identifiers: Other: WOS:000478788200024
DOI: 10.1083/jcb.201812081
ISSN: 0021-9525
 Degree: -

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Title: Journal of Cell Biology
  Alternative Title : J. Cell Biol.
Source Genre: Journal
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Publ. Info: -
Pages: - Volume / Issue: 218 (8) Sequence Number: - Start / End Page: 2797 - 2811 Identifier: -