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| Title | The reduction of FIB damage on cryo-lamella by lowering energy of ion beam revealed by a quantitative analysis. |
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| Journal, issue, pages | Structure, Vol. 31, Issue 10, Page 1275-11281.e4, Year 2023 |
| Publish date | Oct 5, 2023 |
 Authors | Qi Yang / Chunling Wu / Dongjie Zhu / Junxi Li / Jing Cheng / Xinzheng Zhang /  |
| PubMed Abstract | Focused ion beam (FIB) is widely used for thinning frozen cells to produce lamellae for cryo-electron microscopy imaging and for protein structures study in vivo. However, FIB damages the lamellae ...Focused ion beam (FIB) is widely used for thinning frozen cells to produce lamellae for cryo-electron microscopy imaging and for protein structures study in vivo. However, FIB damages the lamellae and a quantitative experimental analysis of the damage is lacking. We used a 30-keV gallium FIB to prepare lamellae of a highly concentrated icosahedral virus sample. The viruses were grouped according to their distance from the surface of lamellae and reconstructed. Damage to the approximately 20-nm-thick outermost lamella surface was similar to that from exposure to 16 e/Å in a 300-kV cryo-electron microscope at high-resolution range. The damage was negligible at a depth beyond 50 nm, which was reduced to 30 nm if 8-keV Ga was used during polishing. We designed extra steps in the reconstruction refinement to maximize undamaged signals and increase the resolution. The results demonstrated that low-energy beam polishing was essential for high-quality thinner lamellae. |
 External links | Structure / PubMed:37527655 |
| Methods | EM (single particle) |
| Resolution | 3.09 - 4.05 Å |
| Structure data |  EMDB-36343: The cryo-EM structure of Parvovirus milled by 30 keV gallium FIB at 3.09 Angstrom resolution.  EMDB-36346: The cryo-EM structure of 10-20nm group Parvovirus from lamellae surface milled by 30 keV Ga+ FIB at 4.05 Angstrom resolution.  EMDB-36347: The cryo-EM structure of 20-30nm group Parvovirus from lamellae surface milled by 30 keV Ga+ FIB at 3.88 Angstrom resolution.  EMDB-36348: The cryo-EM structure of 30-40 nm group Parvovirus from lamellae surface milled by 30 keV Ga+ FIB at 3.82 Angstrom resolution.  EMDB-36349: The cryo-EM structure of 40-50 nm group Parvovirus from lamellae surface milled by 30 keV Ga+ FIB at 3.62 Angstrom resolution.  EMDB-36350: The cryo-EM structure of 50-60 nm group Parvovirus from lamellae surface milled by 30 keV Ga+ FIB at 3.60 Angstrom resolution.  EMDB-36351: The cryo-EM structure of 60-70 nm group Parvovirus from lamellae surface milled by 30 keV Ga+ FIB at 3.57 Angstrom resolution.  EMDB-36352: The cryo-EM structure of Parvovirus milled by 8 keV gallium FIB at 3.11 Angstrom resolution.  EMDB-36355: The cryo-EM structure of 10-20 nm group Parvovirus from lamellae surface milled by 8 keV Ga+ FIB at 3.88 Angstrom resolution.  EMDB-36356: The cryo-EM structure of 20-30 nm group Parvovirus from lamellae surface milled by 8 keV Ga+ FIB at 3.70 Angstrom resolution.  EMDB-36357: The cryo-EM structure of 30-40 nm group Parvovirus from lamellae surface milled by 8 keV Ga+ FIB at 3.60 Angstrom resolution.  EMDB-36358: The cryo-EM structure of 40-50 nm group Parvovirus from lamellae surface milled by 8 keV Ga+ FIB at 3.54 Angstrom resolution.  EMDB-36359: The cryo-EM structure of 50-60 nm group Parvovirus from lamellae surface milled by 8 keV Ga+ FIB at 3.57 Angstrom resolution. |
