Cryo-EM Imaging of Lassa Virus

An aliquot (3 μl) of purified LASV or VLP sample was mixed with 3 μl of colloidal 6-nm gold particles coupled to bovine serum albumin (Aurion, Wageningen, The Netherlands) and applied on a plasma cleaned EM grids coated with holey carbon (C-flat; Protochips, Raleigh, NC). For low pH treatment of VLPs, the grid was floated for three minutes on top of a 2-ml droplet of SPG buffer at the desired pH. The grids were vitrified using a plunger device (CP3; Gatan, Pleasanton, CA) by blotting the sample for 3 s followed by plunge-freezing into a mixture of liquid ethane (37%) and propane (63%) [41 (link)].

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Li S., Sun Z., Pryce R., Parsy M.L., Fehling S.K., Schlie K., Siebert C.A., Garten W., Bowden T.A., Strecker T, & Huiskonen J.T. (2016). Acidic pH-Induced Conformations and LAMP1 Binding of the Lassa Virus Glycoprotein Spike. PLoS Pathogens, 12(2), e1005418.

Publication 2016

bovine serum albumin C-flat

Bovine serum albumin Buffer Carbon Colloidal gold Device Ethane Plasma Propane

Corresponding Organization : Philipps University of Marburg

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Variable analysis

independent variables

PH of SPG buffer
Mixing of purified LASV or VLP sample with colloidal 6-nm gold particles coupled to bovine serum albumin

dependent variables

Morphology of LASV or VLP samples

control variables

Volume of purified LASV or VLP sample (3 μl)
Volume of colloidal 6-nm gold particles coupled to bovine serum albumin (3 μl)
Blotting time (3 s)
Plunge-freezing method (liquid ethane (37%) and propane (63%))
Plasma cleaned EM grids coated with holey carbon

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

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