Localization-based super-resolution imaging meets high-content screening.

Abstract:

:Single-molecule localization microscopy techniques have proven to be essential tools for quantitatively monitoring biological processes at unprecedented spatial resolution. However, these techniques are very low throughput and are not yet compatible with fully automated, multiparametric cellular assays. This shortcoming is primarily due to the huge amount of data generated during imaging and the lack of software for automation and dedicated data mining. We describe an automated quantitative single-molecule-based super-resolution methodology that operates in standard multiwell plates and uses analysis based on high-content screening and data-mining software. The workflow is compatible with fixed- and live-cell imaging and allows extraction of quantitative data like fluorophore photophysics, protein clustering or dynamic behavior of biomolecules. We demonstrate that the method is compatible with high-content screening using 3D dSTORM and DNA-PAINT based super-resolution microscopy as well as single-particle tracking.

journal_name

Nat Methods

journal_title

Nature methods

authors

Beghin A,Kechkar A,Butler C,Levet F,Cabillic M,Rossier O,Giannone G,Galland R,Choquet D,Sibarita JB

doi

10.1038/nmeth.4486

subject

Has Abstract

pub_date

2017-12-01 00:00:00

pages

1184-1190

issue

12

eissn

1548-7091

issn

1548-7105

pii

nmeth.4486

journal_volume

14

pub_type

杂志文章
  • Fluorescent indicators for simultaneous reporting of all four cell cycle phases.

    abstract::A robust method for simultaneous visualization of all four cell cycle phases in living cells is highly desirable. We developed an intensiometric reporter of the transition from S to G2 phase and engineered a far-red fluorescent protein, mMaroon1, to visualize chromatin condensation in mitosis. We combined these new re...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.4045

    authors: Bajar BT,Lam AJ,Badiee RK,Oh YH,Chu J,Zhou XX,Kim N,Kim BB,Chung M,Yablonovitch AL,Cruz BF,Kulalert K,Tao JJ,Meyer T,Su XD,Lin MZ

    更新日期:2016-12-01 00:00:00

  • Anna Moroni.

    abstract:: ...

    journal_title:Nature methods

    pub_type: 评论,新闻

    doi:10.1038/s41592-018-0192-y

    authors: Marx V

    更新日期:2018-11-01 00:00:00

  • Tracking transmitter-gated P2X cation channel activation in vitro and in vivo.

    abstract::We present a noninvasive approach to track activation of ATP-gated P2X receptors and potentially other transmitter-gated cation channels that show calcium fluxes. We genetically engineered rat P2X receptors to carry calcium sensors near the channel pore and tested this as a reporter for P2X(2) receptor opening. The me...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth1144

    authors: Richler E,Chaumont S,Shigetomi E,Sagasti A,Khakh BS

    更新日期:2008-01-01 00:00:00

  • Nanoscale resolution in GFP-based microscopy.

    abstract::We report attainment of subdiffraction resolution using stimulated emission depletion (STED) microscopy with GFP-labeled samples. The approximately 70 nm lateral resolution attained in this study is demonstrated by imaging GFP-labeled viruses and the endoplasmic reticulum (ER) of a mammalian cell. Our results mark the...

    journal_title:Nature methods

    pub_type: 杂志文章,评审

    doi:10.1038/nmeth922

    authors: Willig KI,Kellner RR,Medda R,Hein B,Jakobs S,Hell SW

    更新日期:2006-09-01 00:00:00

  • Wirelessly powered, fully internal optogenetics for brain, spinal and peripheral circuits in mice.

    abstract::To enable sophisticated optogenetic manipulation of neural circuits throughout the nervous system with limited disruption of animal behavior, light-delivery systems beyond fiber optic tethering and large, head-mounted wireless receivers are desirable. We report the development of an easy-to-construct, implantable wire...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.3536

    authors: Montgomery KL,Yeh AJ,Ho JS,Tsao V,Mohan Iyer S,Grosenick L,Ferenczi EA,Tanabe Y,Deisseroth K,Delp SL,Poon AS

    更新日期:2015-10-01 00:00:00

  • A large-scale evaluation of computational protein function prediction.

    abstract::Automated annotation of protein function is challenging. As the number of sequenced genomes rapidly grows, the overwhelming majority of protein products can only be annotated computationally. If computational predictions are to be relied upon, it is crucial that the accuracy of these methods be high. Here we report th...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.2340

    authors: Radivojac P,Clark WT,Oron TR,Schnoes AM,Wittkop T,Sokolov A,Graim K,Funk C,Verspoor K,Ben-Hur A,Pandey G,Yunes JM,Talwalkar AS,Repo S,Souza ML,Piovesan D,Casadio R,Wang Z,Cheng J,Fang H,Gough J,Koskinen P,Törö

    更新日期:2013-03-01 00:00:00

  • Streaming fragment assignment for real-time analysis of sequencing experiments.

    abstract::We present eXpress, a software package for efficient probabilistic assignment of ambiguously mapping sequenced fragments. eXpress uses a streaming algorithm with linear run time and constant memory use. It can determine abundances of sequenced molecules in real time and can be applied to ChIP-seq, metagenomics and oth...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.2251

    authors: Roberts A,Pachter L

    更新日期:2013-01-01 00:00:00

  • High-performance probes for light and electron microscopy.

    abstract::We describe an engineered family of highly antigenic molecules based on GFP-like fluorescent proteins. These molecules contain numerous copies of peptide epitopes and simultaneously bind IgG antibodies at each location. These 'spaghetti monster' fluorescent proteins (smFPs) distributed well in neurons, notably into sm...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.3365

    authors: Viswanathan S,Williams ME,Bloss EB,Stasevich TJ,Speer CM,Nern A,Pfeiffer BD,Hooks BM,Li WP,English BP,Tian T,Henry GL,Macklin JJ,Patel R,Gerfen CR,Zhuang X,Wang Y,Rubin GM,Looger LL

    更新日期:2015-06-01 00:00:00

  • Tracking protein aggregation and mislocalization in cells with flow cytometry.

    abstract::We applied pulse-shape analysis (PulSA) to monitor protein localization changes in mammalian cells by flow cytometry. PulSA enabled high-throughput tracking of protein aggregation, translocation from the cytoplasm to the nucleus and trafficking from the plasma membrane to the Golgi as well as stress-granule formation....

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.1930

    authors: Ramdzan YM,Polling S,Chia CP,Ng IH,Ormsby AR,Croft NP,Purcell AW,Bogoyevitch MA,Ng DC,Gleeson PA,Hatters DM

    更新日期:2012-03-18 00:00:00

  • Designer receptors for every body.

    abstract::Using a directed evolution approach, researchers demonstrate a way of creating 'designer' receptors that are specifically activated by a ligand with no other biological activity in the cell. ...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth0507-382b

    authors: Kaganman I

    更新日期:2007-05-01 00:00:00

  • Improved and expanded Q-system reagents for genetic manipulations.

    abstract::The Q system is a repressible binary expression system for transgenic manipulations in living organisms. Through protein engineering and in vivo functional tests, we report here variants of the Q-system transcriptional activator, including QF2, for driving strong and ubiquitous expression in all Drosophila tissues. Ou...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.3250

    authors: Riabinina O,Luginbuhl D,Marr E,Liu S,Wu MN,Luo L,Potter CJ

    更新日期:2015-03-01 00:00:00

  • RESA identifies mRNA-regulatory sequences at high resolution.

    abstract::Gene expression is extensively regulated at the levels of mRNA stability, localization and translation. However, decoding functional RNA-regulatory features remains a limitation to understanding post-transcriptional regulation in vivo. Here, we developed RNA-element selection assay (RESA), a method that selects RNA el...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.4121

    authors: Yartseva V,Takacs CM,Vejnar CE,Lee MT,Giraldez AJ

    更新日期:2017-02-01 00:00:00

  • The mammalian-membrane two-hybrid assay (MaMTH) for probing membrane-protein interactions in human cells.

    abstract::Cell signaling, one of key processes in both normal cellular function and disease, is coordinated by numerous interactions between membrane proteins that change in response to stimuli. We present a split ubiquitin-based method for detection of integral membrane protein-protein interactions (PPIs) in human cells, terme...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.2895

    authors: Petschnigg J,Groisman B,Kotlyar M,Taipale M,Zheng Y,Kurat CF,Sayad A,Sierra JR,Mattiazzi Usaj M,Snider J,Nachman A,Krykbaeva I,Tsao MS,Moffat J,Pawson T,Lindquist S,Jurisica I,Stagljar I

    更新日期:2014-05-01 00:00:00

  • NeuroGPS-Tree: automatic reconstruction of large-scale neuronal populations with dense neurites.

    abstract::The reconstruction of neuronal populations, a key step in understanding neural circuits, remains a challenge in the presence of densely packed neurites. Here we achieved automatic reconstruction of neuronal populations by partially mimicking human strategies to separate individual neurons. For populations not resolvab...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.3662

    authors: Quan T,Zhou H,Li J,Li S,Li A,Li Y,Lv X,Luo Q,Gong H,Zeng S

    更新日期:2016-01-01 00:00:00

  • Deciphering laminar-specific neural inputs with line-scanning fMRI.

    abstract::Using a line-scanning method during functional magnetic resonance imaging (fMRI), we obtained high temporal (50-ms) and spatial (50-μm) resolution information along the cortical thickness and showed that the laminar position of fMRI onset coincides with distinct neural inputs in rat somatosensory and motor cortices. T...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.2730

    authors: Yu X,Qian C,Chen DY,Dodd SJ,Koretsky AP

    更新日期:2014-01-01 00:00:00

  • Conditional genome engineering in Toxoplasma gondii uncovers alternative invasion mechanisms.

    abstract::We established a conditional site-specific recombination system based on dimerizable Cre recombinase-mediated recombination in the apicomplexan parasite Toxoplasma gondii. Using a new single-vector strategy that allows ligand-dependent, efficient removal of a gene of interest, we generated three knockouts of apicomple...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.2301

    authors: Andenmatten N,Egarter S,Jackson AJ,Jullien N,Herman JP,Meissner M

    更新日期:2013-02-01 00:00:00

  • Colony filtration blot: a new screening method for soluble protein expression in Escherichia coli.

    abstract::The implementation of efficient technologies for the production of recombinant mammalian proteins remains an outstanding challenge in many structural and functional genomics programs. We have developed a new method for rapid identification of soluble protein expression in E. coli, based on a separation of soluble prot...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth767

    authors: Cornvik T,Dahlroth SL,Magnusdottir A,Herman MD,Knaust R,Ekberg M,Nordlund P

    更新日期:2005-07-01 00:00:00

  • Strategy for the fine characterization of glycosyltransferase specificity using isotopomer assembly.

    abstract::Glycosylation, which represents the most complex posttranslational modification (PTM) event during protein maturation, has a vital role in biological processes. Glycan biosynthesis is orchestrated by numerous glycosyltransferases, each displaying different selectivities for multiple reaction sites. The precise specifi...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth1050

    authors: Ito H,Kameyama A,Sato T,Sukegawa M,Ishida HK,Narimatsu H

    更新日期:2007-07-01 00:00:00

  • An FDR metric for top-down proteomics.

    abstract:: ...

    journal_title:Nature methods

    pub_type: 评论,杂志文章

    doi:10.1038/s41592-019-0343-9

    authors: Doerr A

    更新日期:2019-03-01 00:00:00

  • Fast, high-contrast imaging of animal development with scanned light sheet-based structured-illumination microscopy.

    abstract::Recording light-microscopy images of large, nontransparent specimens, such as developing multicellular organisms, is complicated by decreased contrast resulting from light scattering. Early zebrafish development can be captured by standard light-sheet microscopy, but new imaging strategies are required to obtain high-...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.1476

    authors: Keller PJ,Schmidt AD,Santella A,Khairy K,Bao Z,Wittbrodt J,Stelzer EH

    更新日期:2010-08-01 00:00:00

  • fMRIPrep: a robust preprocessing pipeline for functional MRI.

    abstract::Preprocessing of functional magnetic resonance imaging (fMRI) involves numerous steps to clean and standardize the data before statistical analysis. Generally, researchers create ad hoc preprocessing workflows for each dataset, building upon a large inventory of available tools. The complexity of these workflows has s...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/s41592-018-0235-4

    authors: Esteban O,Markiewicz CJ,Blair RW,Moodie CA,Isik AI,Erramuzpe A,Kent JD,Goncalves M,DuPre E,Snyder M,Oya H,Ghosh SS,Wright J,Durnez J,Poldrack RA,Gorgolewski KJ

    更新日期:2019-01-01 00:00:00

  • Nanoscale imaging of RNA with expansion microscopy.

    abstract::The ability to image RNA identity and location with nanoscale precision in intact tissues is of great interest for defining cell types and states in normal and pathological biological settings. Here, we present a strategy for expansion microscopy of RNA. We developed a small-molecule linker that enables RNA to be cova...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.3899

    authors: Chen F,Wassie AT,Cote AJ,Sinha A,Alon S,Asano S,Daugharthy ER,Chang JB,Marblestone A,Church GM,Raj A,Boyden ES

    更新日期:2016-08-01 00:00:00

  • Non-uniform refinement: adaptive regularization improves single-particle cryo-EM reconstruction.

    abstract::Cryogenic electron microscopy (cryo-EM) is widely used to study biological macromolecules that comprise regions with disorder, flexibility or partial occupancy. For example, membrane proteins are often kept in solution with detergent micelles and lipid nanodiscs that are locally disordered. Such spatial variability ne...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/s41592-020-00990-8

    authors: Punjani A,Zhang H,Fleet DJ

    更新日期:2020-12-01 00:00:00

  • CancerMine: a literature-mined resource for drivers, oncogenes and tumor suppressors in cancer.

    abstract::Tumors from individuals with cancer are frequently genetically profiled for information about the driving forces behind the disease. We present the CancerMine resource, a text-mined and routinely updated database of drivers, oncogenes and tumor suppressors in different types of cancer. All data are available online ( ...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/s41592-019-0422-y

    authors: Lever J,Zhao EY,Grewal J,Jones MR,Jones SJM

    更新日期:2019-06-01 00:00:00

  • Detergent-free mass spectrometry of membrane protein complexes.

    abstract::We developed a method that allows release of intact membrane protein complexes from amphipols, bicelles and nanodiscs in the gas phase for observation by mass spectrometry (MS). Current methods involve release of membrane protein complexes from detergent micelles, which reveals subunit composition and lipid binding. W...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.2691

    authors: Hopper JT,Yu YT,Li D,Raymond A,Bostock M,Liko I,Mikhailov V,Laganowsky A,Benesch JL,Caffrey M,Nietlispach D,Robinson CV

    更新日期:2013-12-01 00:00:00

  • Analysis of the Human Protein Atlas Image Classification competition.

    abstract::Pinpointing subcellular protein localizations from microscopy images is easy to the trained eye, but challenging to automate. Based on the Human Protein Atlas image collection, we held a competition to identify deep learning solutions to solve this task. Challenges included training on highly imbalanced classes and pr...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/s41592-019-0658-6

    authors: Ouyang W,Winsnes CF,Hjelmare M,Cesnik AJ,Åkesson L,Xu H,Sullivan DP,Dai S,Lan J,Jinmo P,Galib SM,Henkel C,Hwang K,Poplavskiy D,Tunguz B,Wolfinger RD,Gu Y,Li C,Xie J,Buslov D,Fironov S,Kiselev A,Panchenko D,C

    更新日期:2019-12-01 00:00:00

  • Non-reversible tissue fixation retains extracellular vesicles for in situ imaging.

    abstract::Extracellular vesicles (EVs) are secreted nanosized particles with many biological functions and pathological associations. The inability to image EVs in fixed tissues has been a major limitation to understanding their role in healthy and diseased tissue microenvironments. Here, we show that crosslinking mammalian tis...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/s41592-019-0623-4

    authors: Gupta MP,Tandalam S,Ostrager S,Lever AS,Fung AR,Hurley DD,Alegre GB,Espinal JE,Remmel HL,Mukherjee S,Levine BM,Robins RP,Molina H,Dill BD,Kenific CM,Tuschl T,Lyden D,D'Amico DJ,Pena JTG

    更新日期:2019-12-01 00:00:00

  • Directed evolution of APEX2 for electron microscopy and proximity labeling.

    abstract::APEX is an engineered peroxidase that functions as an electron microscopy tag and a promiscuous labeling enzyme for live-cell proteomics. Because limited sensitivity precludes applications requiring low APEX expression, we used yeast-display evolution to improve its catalytic efficiency. APEX2 is far more active in ce...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.3179

    authors: Lam SS,Martell JD,Kamer KJ,Deerinck TJ,Ellisman MH,Mootha VK,Ting AY

    更新日期:2015-01-01 00:00:00

  • Dynamic characterization of growth and gene expression using high-throughput automated flow cytometry.

    abstract::Cells adjust to changes in environmental conditions using complex regulatory programs. These cellular programs are the result of an intricate interplay between gene expression, cellular growth and protein degradation. Technologies that enable simultaneous and time-resolved measurements of these variables are necessary...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.2879

    authors: Zuleta IA,Aranda-Díaz A,Li H,El-Samad H

    更新日期:2014-04-01 00:00:00

  • An auxin-based degron system for the rapid depletion of proteins in nonplant cells.

    abstract::Plants have evolved a unique system in which the plant hormone auxin directly induces rapid degradation of the AUX/IAA family of transcription repressors by a specific form of the SCF E3 ubiquitin ligase. Other eukaryotes lack the auxin response but share the SCF degradation pathway, allowing us to transplant the auxi...

    journal_title:Nature methods

    pub_type: 杂志文章

    doi:10.1038/nmeth.1401

    authors: Nishimura K,Fukagawa T,Takisawa H,Kakimoto T,Kanemaki M

    更新日期:2009-12-01 00:00:00