Ultraufoil r1.2 1 3 au300 mesh grids

Manufactured by Quantifoil

The UltrAuFoil R1.2/1.3 Au300 mesh grids are a type of laboratory equipment produced by Quantifoil. They are designed to serve as sample supports for various imaging and analysis techniques. The grids are composed of a gold (Au) material and have a specified mesh size of 300.

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Lab products found in correlation

Vitrobot mark 4, by Thermo Fisher Scientific (4 mentions) Mk 2 vitrobot, by Thermo Fisher Scientific (3 mentions) K3 detector, by Ametek (2 mentions) K2 summit direct detector, by Ametek (2 mentions) K3 direct detector, by Ametek (2 mentions) Titan krios g3, by Ametek (2 mentions) Titan krios g3, by Thermo Fisher Scientific (2 mentions) Pelco easiglow, by Ted Pella (2 mentions) Pelco easiglow unit, by Ted Pella (1 mentions) Titan krios, by Thermo Fisher Scientific (1 mentions) 595 filter paper, by Ted Pella (1 mentions) Post gif k3 direct detector, by Ametek (1 mentions) R1.2 1 3 cu rh 200 mesh grids, by Quantifoil (1 mentions)

5 protocols using ultraufoil r1.2 1 3 au300 mesh grids

Cryo-EM Imaging of Purified Samples

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UltrAuFoil R1.2/1.3 Au300 mesh grids (Quantifoil) were glow discharged for 1 min on each side at 25 mA using a Pelco easiGlow unit (Ted Pella, Inc.). 3.5 μl of purified sample were applied to the grid and blotted with filter paper (595 Filter Paper, Ted Pella, Inc.) for 2.5 s using a Mk. II Vitrobot (Thermo Fisher Scientific) with a −1 mm offset and then plunge frozen into liquid ethane.
Cryo-EM movies were recorded using SerialEM v3.8⁴⁰ in super-resolution mode on a 300 kV Titan Krios (Thermo Fisher Scientific) equipped with a post-GIF K3 direct detector (Gatan, Inc.). A total of 9732 movies were recorded at a nominal magnification of 81,000×, corresponding to a super-resolution pixel size of 0.54 Å, with a total dose of 46 electrons/Å² and 40 frames per movie.

Xu Y., Han H., Cooney I., Guo Y., Moran N.G., Zuniga N.R., Price J.C., Hill C.P, & Shen P.S. (2022). Active conformation of the p97-p47 unfoldase complex. Nature Communications, 13, 2640.

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Cryo-EM Sample Preparation and Data Acquisition

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3 µL of sample was applied to glow-discharged UltrAuFoil R1.2/1.3 Au 300 mesh grids (Quantifoil) and vitrified on a Vitrobot Mark IV (Thermo Fisher Scientific) set to 4 °C and 100% humidity and 10 s blot time. Data were collected on a Titan Krios G3i 300 kV electron microscope (Themo Fisher Scientific) equipped with GIF Quantum energy filter and K3 detector (Gatan). Data acquisition was performed in EFTEM NanoProbe mode with a 50 µM C2 aperture at an indicated magnification of ×105,000 with zero-loss slit width of 25 eV. The data was collected automatically with homemade scripts for SerialEM^{57 (link)} performing a 9-hole beam-image shift acquisition scheme with one exposure in the centre of each hole. Experimental parameters are listed in Supplementary Table 1.

Burger W.A., Pham V., Vuckovic Z., Powers A.S., Mobbs J.I., Laloudakis Y., Glukhova A., Wootten D., Tobin A.B., Sexton P.M., Paul S.M., Felder C.C., Danev R., Dror R.O., Christopoulos A., Valant C, & Thal D.M. (2023). Xanomeline displays concomitant orthosteric and allosteric binding modes at the M4 mAChR. Nature Communications, 14, 5440.

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Cryo-EM Imaging of RACK1 Proteins

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UltrAuFoil R1.2/1.3 Au300 mesh grids (Quantifoil) were glow discharged using a Pelco easiGlow (Ted Pella, Inc.) for 25 s at 25 mA. 3.5 μL of sample were applied to the glow discharged grid, and grids were vitrified using a Mk. II Vitrobot (Thermo Fisher Scientific) operating at ~85% relative humidity, 4°C, and 2.5 s blot time, and then plunge frozen into liquid ethane.
A total of 3,842 cryo-EM movies of S²⁷⁸E RACK1 samples were recorded using a 300kV Titan Krios G3 (Thermo Fisher Scientific) equipped with a K2 Summit direct detector (Gatan, Inc.) at a nominal magnification of 105K x, corresponding to 1.348 Å pixel size. Movies were recorded using SerialEM with a defocus range of −1.0 to −3.0 μm and at a dose rate of 1.0 e⁻/Å²/frame with a total exposure of 40 frames and each movie recording time was 8 s (Mastronarde, 2005 (link)).
For WT RACK1 samples, a total of 2,141 movies were recorded on a 300kV Titan Krios G3 equipped with a K3 direct detector (Gatan) at a nominal magnification of 81,000× corresponding to 1.058 Å pixel size. Data were collected in super-resolution mode using SerialEM with a defocus range of from −0.8 to −1.8 μm and at a dose rate of 1.1 e⁻/Å²/frame with a total exposure of 40 frames and each movie recorded for 2.53 s.

Rollins M.G., Shasmal M., Meade N., Astar H., Shen P.S, & Walsh D. (2021). Negative charge in the RACK1 loop broadens the translational capacity of the human ribosome. Cell reports, 36(10), 109663.

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Cryo-EM Imaging of RACK1 Proteins

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Rollins M.G., Shasmal M., Meade N., Astar H., Shen P.S, & Walsh D. (2021). Negative charge in the RACK1 loop broadens the translational capacity of the human ribosome. Cell reports, 36(10), 109663.

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Cryo-EM Structural Analysis Protocol

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Samples (3 µl) were applied to glow-discharged Quantifoil R1.2/1.3 Cu/Rh 200 mesh grids (Quantifoil) (M4R-G_i1-Ipx and M4R-G_i1-Ipx-LY298) or UltrAuFoil R1.2/1.3 Au 300 mesh grids (Quantifoil) (M4R-G_i1-Ipx-VU154 and M4R-G_i1-Ach) and were vitrified on a Vitrobot Mark IV (Thermo Fisher Scientific) set to 4°C and 100% humidity and 10 s blot time. Data were collected on a Titan Krios G3i 300 kV electron microscope (Thermo Fisher Scientific) equipped with GIF Quantum energy filter and K3 detector (Gatan). Data acquisition was performed in EFTEM NanoProbe mode with a 50 µM C2 aperture at an indicated magnification of ×105,000 with zero-loss slit width of 25 eV. The data were collected automatically with homemade scripts for SerialEM performing a nine-hole beam-image shift acquisition scheme with one exposure in the center of each hole. Experimental parameters specific to each collected data set is listed in Table 2.

Vuckovic Z., Wang J., Pham V., Mobbs J.I., Belousoff M.J., Bhattarai A., Burger W.A., Thompson G., Yeasmin M., Nawaratne V., Leach K., van der Westhuizen E.T., Khajehali E., Liang Y.L., Glukhova A., Wootten D., Lindsley C.W., Tobin A., Sexton P., Danev R., Valant C., Miao Y., Christopoulos A, & Thal D.M. (2023). Pharmacological hallmarks of allostery at the M4 muscarinic receptor elucidated through structure and dynamics. eLife, 12, e83477.

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