SARS-CoV-2 S-6P spike at a concentration of 0.5 mg/mL in PBS was mixed with Fab NA8 or Fab NE12 using a Fab-to-RBD molar ratio of 1:1, and the complex was used for specimen preparation after a brief (5-10 min) incubation at 4°C. To prepare cryo-EM specimens, Quantifoil R 2/2 gold grids were glow-discharged using a PELCO easiGlow device (air pressure: 0.39 mBar, current: 20 mA, duration: 30 s) immediately before vitrification using an FEI Vitrobot Mark IV plunger. The Vitrobot chamber was kept at 4°C and 95% humidity, and the drop volume was 2.7 μL. Datasets were collected at the National CryoEM Facility (NCEF), National Cancer Institute, using a Thermo Scientific Titan Krios G3 electron microscope equipped with a K3 direct electron detector and Gatan Quantum GIF energy filter (slit width: 20 eV) (Table S4 ).
Titan krios g3 electron microscope
Manufactured by Ametek
The Titan Krios G3 is an advanced electron microscope designed for high-resolution structural analysis. It features a stable and powerful electron beam, state-of-the-art optics, and a large, high-quality image sensor. The Titan Krios G3 is capable of producing detailed, high-resolution images of a wide range of samples, making it a valuable tool for researchers and scientists working in the field of structural biology.
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Lab products found in correlation
2 protocols using titan krios g3 electron microscope
1
Cryo-EM Structural Analysis of SARS-CoV-2 Spike Complexes
2
Cryo-EM Structure of SARS-CoV-2 Spike with Fab
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The stabilized SARS CoV-2 spike HexaPro (3) was mixed with Fab A23-58.1 or B1-182.1 at a molar ratio of 1.2 Fab per protomer in PBS. The final spike protein concentration was 0.5 mg/ml. n-Dodecyl β-D-maltoside (DDM) detergent was added shortly before vitrification to a concentration of 0.005%. Quantifoil R 2/2 gold grids were subjected to glow discharging in a PELCO easiGlow device (air pressure, 0.39 mBar; current, 20 mA; duration, 30 s) immediately before specimen preparation. Cryo-EM grids were prepared using an FEI Vitrobot Mark IV plunger with the following settings: chamber temperature of 4°C, chamber humidity of 95%, blotting force of –5, blotting time of 3 s, and drop volume of 2.7 μl. Datasets were collected at the National CryoEM Facility (NCEF), National Cancer Institute, on a Thermo Scientific Titan Krios G3 electron microscope equipped with a Gatan Quantum GIF energy filter (slit width: 20 eV) and a Gatan K3 direct electron detector (table S2). Four movies per hole were recorded in the counting mode using Latitude software. The dose rate was 14.65 e–/s/pixel.
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