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Pierson J, et al.  (2010) Improving the technique of vitreous cryo-sectioning for cryo-electron tomography: Electrostatic charging for section attachment and implementation of an anti-contamination glove box. J Struct Biol 169(2):219-225

Abstract: Cryo-electron tomography of vitreous cryo-sections is the most suitable method for exploring the 3D organization of biological samples that are too large to be imaged in an intact state. Producing good quality vitreous cryo-sections, however, is challenging. Here, we focused on the major obstacles to success: contamination in and around the microtome, and attachment of the ribbon of sections to an electron microscopic grid support film. The conventional method for attaching sections to the grid has involved mechanical force generated by a crude stamping or pressing device, but this disrupts the integrity of vitreous cryo-sections. Furthermore, attachment is poor, and parts of the ribbon of sections are often far from the support film. This results in specimen instability during image acquisition and subsequent difficulty with aligning projection images. Here, we have implemented a protective glove box surrounding the cryo-ultramicrotome that reduces the humidity around and within the microtome during sectioning. We also introduce a novel way to attach vitreous cryo-sections to an EM grid support film using electrostatic charging. The ribbon of vitreous cryo-sections remains in place during transfer and storage and is devoid of stamping related artefacts. We illustrate these improvements by exploring the structure of putative cellular 80S ribosomes within 50nm, vitreous cryo-sections of Saccharomyces cerevisiae.

Status: Published Type: Journal Article PubMed ID: 19822214

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ASC1 FUN12 NIP7 NMD3 PAT1 RDN18-1 RDN18-2 RDN25-1 RDN25-2 RDN5-1
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RDN5-2 RDN5-3 RDN5-4 RDN5-5 RDN5-6 RDN58-1 RDN58-2 RPL10 RPL11A RPL11B
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Topics Genes linked to topics (#91 - 100 )
RPL8B RPL9A RPL9B RPP0 RPP1A RPP1B RPP2A RPP2B RPS0A RPS0B
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Topics Genes linked to topics (#101 - 110 )
RPS10A RPS10B RPS11A RPS11B RPS12 RPS13 RPS14A RPS14B RPS15 RPS16A
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RPS16B RPS17A RPS17B RPS18A RPS18B RPS19A RPS19B RPS1A RPS1B RPS2
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RPS20 RPS21A RPS21B RPS22A RPS22B RPS23A RPS23B RPS24A RPS24B RPS25A
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Topics Genes linked to topics (#131 - 140 )
RPS25B RPS26A RPS26B RPS27A RPS27B RPS28A RPS28B RPS29A RPS29B RPS3
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Protein/Nucleic Acid Structure blue ball blue ball blue ball blue ball blue ball blue ball blue ball blue ball blue ball blue ball

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RPS30A RPS30B RPS31 RPS4A RPS4B RPS5 RPS6A RPS6B RPS7A RPS7B
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Protein/Nucleic Acid Structure blue ball blue ball blue ball blue ball blue ball blue ball blue ball blue ball blue ball blue ball

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RPS8A RPS8B RPS9A RPS9B SIS1 TIF5 YGR054W
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