+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-10533 | |||||||||
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Title | Horse spleen apoferritin deposited with a microfluidic sprayer | |||||||||
Map data | Final sharpened map | |||||||||
Sample |
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Function / homology | Function and homology information : / intracellular sequestering of iron ion / ferric iron binding / ferrous iron binding / iron ion transport / iron ion binding / cytoplasm Similarity search - Function | |||||||||
Biological species | Equus caballus (horse) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.5 Å | |||||||||
Authors | Klebl DP / Muench SP | |||||||||
Funding support | United Kingdom, 1 items
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Citation | Journal: Acta Crystallogr D Struct Biol / Year: 2020 Title: Sample deposition onto cryo-EM grids: from sprays to jets and back. Authors: David P Klebl / Diana C F Monteiro / Dimitrios Kontziampasis / Florian Kopf / Frank Sobott / Howard D White / Martin Trebbin / Stephen P Muench / Abstract: Despite the great strides made in the field of single-particle cryogenic electron microscopy (cryo-EM) in microscope design, direct electron detectors and new processing suites, the area of sample ...Despite the great strides made in the field of single-particle cryogenic electron microscopy (cryo-EM) in microscope design, direct electron detectors and new processing suites, the area of sample preparation is still far from ideal. Traditionally, sample preparation involves blotting, which has been used to achieve high resolution, particularly for well behaved samples such as apoferritin. However, this approach is flawed since the blotting process can have adverse effects on some proteins and protein complexes, and the long blot time increases exposure to the damaging air-water interface. To overcome these problems, new blotless approaches have been designed for the direct deposition of the sample on the grid. Here, different methods of producing droplets for sample deposition are compared. Using gas dynamic virtual nozzles, small and high-velocity droplets were deposited on cryo-EM grids, which spread sufficiently for high-resolution cryo-EM imaging. For those wishing to pursue a similar approach, an overview is given of the current use of spray technology for cryo-EM grid preparation and areas for enhancement are pointed out. It is further shown how the broad aspects of sprayer design and operation conditions can be utilized to improve grid quality reproducibly. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_10533.map.gz | 6.2 MB | EMDB map data format | |
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Header (meta data) | emd-10533-v30.xml emd-10533.xml | 17.6 KB 17.6 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_10533_fsc.xml | 7.2 KB | Display | FSC data file |
Images | emd_10533.png | 82.6 KB | ||
Masks | emd_10533_msk_1.map | 30.5 MB | Mask map | |
Others | emd_10533_additional.map.gz emd_10533_half_map_1.map.gz emd_10533_half_map_2.map.gz | 20.1 MB 20.5 MB 20.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-10533 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-10533 | HTTPS FTP |
-Validation report
Summary document | emd_10533_validation.pdf.gz | 383.9 KB | Display | EMDB validaton report |
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Full document | emd_10533_full_validation.pdf.gz | 383 KB | Display | |
Data in XML | emd_10533_validation.xml.gz | 13.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10533 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10533 | HTTPS FTP |
-Related structure data
Similar structure data |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_10533.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Final sharpened map | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.07 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_10533_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Unsharpened map
File | emd_10533_additional.map | ||||||||||||
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Annotation | Unsharpened map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Halfmap 1
File | emd_10533_half_map_1.map | ||||||||||||
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Annotation | Halfmap 1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Halfmap 2
File | emd_10533_half_map_2.map | ||||||||||||
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Annotation | Halfmap 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : apoferritin from equine spleen
Entire | Name: apoferritin from equine spleen |
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Components |
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-Supramolecule #1: apoferritin from equine spleen
Supramolecule | Name: apoferritin from equine spleen / type: complex / ID: 1 / Parent: 0 |
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Source (natural) | Organism: Equus caballus (horse) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 9 mg/mL | |||||||||
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Buffer | pH: 7.5 Component:
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Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.01 kPa | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 295 K / Instrument: HOMEMADE PLUNGER Details: sample sprayed onto grid using microfluidic nozzle and frozen 36 ms after spraying. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 690 / Average exposure time: 8.0 sec. / Average electron dose: 60.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.8 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |