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- EMDB-71205: ExoSloNano proof of principle labeling the ribosome in intact and... -

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Entry
Database: EMDB / ID: EMD-71205
TitleExoSloNano proof of principle labeling the ribosome in intact and vitreous cells with 5 nm NG
Map dataExoSloNano: 5nm-NG labeled ribosomes from vitrified cells cryo-FIB-milled.
Sample
  • Complex: ribosome labeled with 1.4 nm nanogold inside cell
KeywordsExoSloNano / RIBOSOME
Biological speciesHomo sapiens (human)
Methodsubtomogram averaging / cryo EM / Resolution: 16.0 Å
AuthorsYoung L / Villa E
Funding support United States, 1 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Nat Methods / Year: 2025
Title: ExoSloNano: multimodal nanogold labels for identification of macromolecules in live cells and cryo-electron tomograms.
Authors: Lindsey N Young / Alice Sherrard / Huabin Zhou / Farhaz Shaikh / Joshua Hutchings / Margot Riggi / Mythreyi Narasimhan / W Alexander Flaherty / Eric J Bennett / Michael K Rosen / Antonio J ...Authors: Lindsey N Young / Alice Sherrard / Huabin Zhou / Farhaz Shaikh / Joshua Hutchings / Margot Riggi / Mythreyi Narasimhan / W Alexander Flaherty / Eric J Bennett / Michael K Rosen / Antonio J Giraldez / Elizabeth Villa /
Abstract: In situ cryo-electron microscopy (cryo-EM) enables the direct interrogation of structure-function relationships by resolving macromolecular structures in their native cellular environment. Recent ...In situ cryo-electron microscopy (cryo-EM) enables the direct interrogation of structure-function relationships by resolving macromolecular structures in their native cellular environment. Recent progress in sample preparation, imaging and data processing has enabled the identification and determination of large biomolecular complexes. However, the majority of proteins are of a size that still eludes identification in cellular cryo-EM data, and most proteins exist in low copy numbers. Therefore, novel tools are needed for cryo-EM to identify macromolecules across multiple size scales (from microns to nanometers). Here we introduce nanogold probes for detecting specific proteins using correlative light and electron microscopy, cryo-electron tomography (cryo-ET) and resin-embedded electron microscopy. These nanogold probes can be introduced into live cells, in a manner that preserves intact molecular networks and cell viability. We use this ExoSloNano system to identify both cytoplasmic and nuclear proteins by room-temperature electron microscopy, and resolve associated structures by cryo-ET. By providing high-efficiency protein labeling in live cells and molecular specificity within cryo-ET tomograms, ExoSloNano expands the proteome available to electron microscopy.
History
DepositionJun 12, 2025-
Header (metadata) releaseNov 19, 2025-
Map releaseNov 19, 2025-
UpdateDec 10, 2025-
Current statusDec 10, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_71205.png

Downloads & links

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Map

FileDownload / File: emd_71205.map.gz / Format: CCP4 / Size: 1.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationExoSloNano: 5nm-NG labeled ribosomes from vitrified cells cryo-FIB-milled.
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
8 Å/pix.
x 72 pix.
= 576. Å		8 Å/pix.
x 72 pix.
= 576. Å		8 Å/pix.
x 72 pix.
= 576. Å

Surface

Projections

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Images are generated by Spider.

Voxel sizeX=Y=Z: 8 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 2.84
Minimum - Maximum-0.960064 - 5.4500794
Average (Standard dev.)0.12108097 (±0.57594544)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions727272
Spacing727272
CellA=B=C: 576.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_71205_msk_1.map
Projections & Slices
AxesZYX

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Histogram

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Half map: Unfiltered half-map (half 1).

Fileemd_71205_half_map_1.map
AnnotationUnfiltered half-map (half 1).
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: Unfiltered half-map (half 2).

Fileemd_71205_half_map_2.map
AnnotationUnfiltered half-map (half 2).
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : ribosome labeled with 1.4 nm nanogold inside cell

EntireName: ribosome labeled with 1.4 nm nanogold inside cell
Components
  • Complex: ribosome labeled with 1.4 nm nanogold inside cell

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Supramolecule #1: ribosome labeled with 1.4 nm nanogold inside cell

SupramoleculeName: ribosome labeled with 1.4 nm nanogold inside cell / type: complex / ID: 1 / Parent: 0
Details: ExoSloNano proof of principle, labeling ribosomes with 1.4 nm nanogold inside cells and performing subtomogram analysis from vitrified cryo-FIB-thinned cells.
Source (natural)Organism: Homo sapiens (human) / Strain: Human Embryonic Kidney cells

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 7.3
GridModel: Quantifoil R1/4 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS
VitrificationCryogen name: ETHANE-PROPANE

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOCONTINUUM (6k x 4k) / Average electron dose: 4.8 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 81000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 16.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1.4) / Number subtomograms used: 1743
ExtractionNumber tomograms: 41 / Number images used: 13748 / Software - Name: RELION (ver. 3.1.4)
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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