Pelco easiglow device

Manufactured by Ted Pella

Sourced in Germany

The PELCO easiGlow is a device used for glow discharging of sample grids. It is designed to prepare sample grids for transmission electron microscopy (TEM) analysis by creating a thin, uniform carbon film on the grid surface.

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Lab products found in correlation

Vitrobot mark 4, by Thermo Fisher Scientific (8 mentions) Vitrobot plunge freezer, by Thermo Fisher Scientific (3 mentions) Ultraufoil r1.2 1 3 300 mesh grid, by Quantifoil (2 mentions) Ultraufoil r1.2 1 3 grids, by Quantifoil (1 mentions) Fei titan krios transmission electron microscope, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

PELCO easiGlow (136 citations) EasiGlow (9 citations)

8 protocols using pelco easiglow device

Cryo-EM of Spike-ACE2 Complex

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Cryo-EM grids were prepared with a Vitrobot Mark IV (Thermofisher Scientific (TFS)). Quantifoil R1.2/1.3 Au 400 holey carbon grids were glow-discharged for 120 s at 15mA using a PELCO easiGlow device (Ted Pella, Inc.). 3.0 μL of a sample containing 14 μM of the corresponding Spike and 25 μM ACE2-Fc was applied to the glow-discharged grids, and blotted for 6 s under blot force 10 at 100% humidity and 4°C in the sample chamber, and the blotted grid was plunge-frozen in liquid nitrogen-cooled liquid ethane.
Grids were screened for particle presence and ice quality on a TFS Glacios TEM (200kV), and the best grids were transferred to TFS Titan Krios G4 TEM. Cryo-EM data was collected using the TFS Titan Krios G4 TEM, equipped with a Cold-FEG, on a Falcon IV detector in electron counting mode. Falcon IV gain references were collected just before data collection. Data was collected using TFS EPU v2.12.1 using aberration-free image shift protocol (AFIS). Movies were recorded at nominal magnification of 96kx, corresponding to the 0.83Å pixel size at the specimen level, with defocus values ranging from -0.7 to -2.5 μm. Exposures were adjusted automatically to 60 e-/Å2 total dose.

Ni D., Turelli P., Beckert B., Nazarov S., Uchikawa E., Myasnikov A., Pojer F., Trono D., Stahlberg H, & Lau K. (2023). Cryo-EM structures and binding of mouse and human ACE2 to SARS-CoV-2 variants of concern indicate that mutations enabling immune escape could expand host range. PLOS Pathogens, 19(4), e1011206.

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Cryogenic Protein Sample Preparation

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4-μL volumes of the respective samples were applied to UltrAuFoil R1.2/1.3 300-mesh grids (Quantifoil Micro Tools GmbH; Germany) that were prior glow discharged in a Pelco easiGlow device (Ted Pella; CA) at 10 mA for 90 s (IM172N22 complex) or at 15 mA for 30 s (IM459 complex). Grids were blotted for 3 s at −3 blot-force setting in a Vitrobot mark IV (ThermoFisher Scientific; operated at 4 °C and 100% humidity) before being plunge frozen in liquid ethane.

Kirk N.S., Chen Q., Wu Y.G., Asante A.L., Hu H., Espinosa J.F., Martínez-Olid F., Margetts M.B., Mohammed F.A., Kiselyov V.V., Barrett D.G, & Lawrence M.C. (2022). Activation of the human insulin receptor by non-insulin-related peptides. Nature Communications, 13, 5695.

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Cryo-EM Structural Determination Protocol

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Cryo-EM grids were prepared by applying 3.5 µl of concentrated sample on to 400-mesh R1.2/1.3 UltrAuFoil grids (Quantifoil Micro Tools GmbH), which had been rendered hydrophilic by glow discharging at 15 mA for 60 s with a PELCO easiGlow device (Ted Pella, Inc.). The sample was adsorbed for 30 s on the grids, followed by blotting and plunge freezing into liquid ethane using a Vitrobot Mark IV plunge freezer (Thermo Fisher Scientific). Cryo-EM data were collected using the automated data acquisition software EPU (Thermo Fisher Scientific) on a Titan Krios G4 transmission electron microscope (Thermo Fisher Scientific), operating at 300 kV and equipped with a cold field emission gun electron source and a Falcon4 direct detection camera. Images were recorded in counting mode at a nominal magnification of ×96,000, corresponding to a physical pixel size of 0.83 Å at the sample level. Datasets were collected at a defocus range of 0.8–2.5 µm with a total electron dose of 60 e⁻/Å⁻². Image data were saved as electron event recordings.

Ekundayo B., Torre D., Beckert B., Nazarov S., Myasnikov A., Stahlberg H, & Ni D. (2022). Structural insights into the regulation of Cas7-11 by TPR-CHAT. Nature Structural & Molecular Biology, 30(2), 135-139.

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Cryo-EM Grid Preparation Protocol

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A 4 μL volume of the above stock solution was then applied to UltrAuFoil R1.2/1.3 300-mesh grids (Quantifoil Micro Tools GmbH; Großlöbichau, Germany) which were glow discharged prior to sample application in a Pelco easiGlow device (Ted Pella; Redding, California) at 15 mA for 30 s. Grids were blotted for 3 s at -3 blot-force setting in a Vitrobot mark IV (ThermoFisher Scientific; operated at 4°C and 100 % humidity) before being plunge frozen in liquid ethane.

Xu Y., Kirk N.S., Venugopal H., Margetts M.B., Croll T.I., Sandow J.J., Webb A.I., Delaine C.A., Forbes B.E, & Lawrence M.C. (2020). How IGF-II Binds to the Human Type 1 Insulin-like Growth Factor Receptor. Structure(London, England:1993), 28(7), 786-798.e6.

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Vitrified Exosome Imaging Protocol

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Purified exosomes in PBS buffer were vitrified on Quantifoil Cu R200 2/2 (Electron Microscopy Sciences, Ref. No.: Q2100CR2) grids. Prior to sample application, the grids were glow discharged using a Pelco easiGlow device (Ted Pella Inc.) at 15 mA for 30 s. Samples were deposited by transferring 3 μl of sample onto the glow-discharged side of the grid, blotted, then plunge frozen in liquid ethane, using a Vitrobot plunge freezer (Thermo Fisher Scientific), with the following settings: 22°C, 80% humidity, Blot-force = –5, 60 s wait time and a blotting time of 3 s. All data were collected on an FEI Titan Krios transmission electron microscope (Thermo Fisher Scientific) operated at 300 keV and equipped with a Gatan BioQuantum energy filter and a K2 direct electron detector. A condenser aperture of 70 μm and no objective aperture were chosen for data collection. Data were acquired in parallel illumination mode using EPU (Thermo Fisher Scientific) software at a nominal magnification of 33 kx (4.3 Å pixel size).

Deiana M., Andrés Castán J.M., Josse P., Kahsay A., Sánchez D.P., Morice K., Gillet N., Ravindranath R., Patel A.K., Sengupta P., Obi I., Rodriguez-Marquez E., Khrouz L., Dumont E., Abad Galán L., Allain M., Walker B., Ahn H.S., Maury O., Blanchard P., Le Bahers T., Öhlund D., von Hofsten J., Monnereau C., Cabanetos C, & Sabouri N. (2023). A new G-quadruplex-specific photosensitizer inducing genome instability in cancer cells by triggering oxidative DNA damage and impeding replication fork progression. Nucleic Acids Research, 51(12), 6264-6285.

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Cryo-EM sample preparation of HAdV-F41

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Purified HAdV-F41 was used at 4.2 mg/ml (pH 7.4) and 1.6 mg/ml (pH 4.0). The recombinant HAdV-F41 PB (fPB) was purified as described before (29 (link)) and used at 1 mg/ml in PBS buffer, supplemented with 5% glycerol. A HAdV-F41 sample at pH 4.0 was prepared by adding 2 μl of a 0.5 M citric acid/1 M Na₂HPO₄ (pH 3.4) solution to 25 μl of HAdV-F41 (pH 7.4), followed by incubating on ice for 15 min. Samples were vitrified on QUANTIFOIL Cu R200 2/2 (Electron Microscopy Sciences, catalog no. Q2100CR2) and QUANTIFOIL Cu R200 1.2/1.3 (Electron Microscopy Sciences, catalog no. Q3100CR1.3) grids for the virus particles and the recombinant protein, respectively. Before sample application, the grids were glow discharged using a PELCO easiGlow device (Ted Pella Inc.) at 15 mA for 30 s. Sample was applied by transferring 3 μl of sample onto the glow-discharged side of the grid, blotted, and plunge-frozen in liquid ethane, using a Vitrobot plunge freezer (Thermo Fisher Scientific), with the following settings: 22°C, 80% humidity, a blot force of −20, and a blotting time of 3 s. For HAdV-F41 at pH 7.4, sample was applied twice with a blotting step, using the same settings as above, between applications (51 (link)).

Rafie K., Lenman A., Fuchs J., Rajan A., Arnberg N, & Carlson L.A. (2021). The structure of enteric human adenovirus 41—A leading cause of diarrhea in children. Science Advances, 7(2), eabe0974.

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Cryo-EM Structure of Omicron BA.1 and mACE2-Fc

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Cryo-EM grids were prepared with a Vitrobot Mark IV (Thermofisher Scientific (TFS)). Quantifoil R1.2/1.3 Au 400 holey carbon grids were glow-discharged for 120 s at 15mA using a PELCO easiGlow device (Ted Pella, Inc.). 3.0 μL of a sample containing 9 μM Omicron BA.1 and 16 μM mACE2-Fc was applied to the glow-discharged grids, and blotted for 6 s under blot force 10 at 100% humidity and 4°C in the sample chamber, and the blotted grids were plunge-frozen in liquid nitrogen-cooled liquid ethane.
Grids were screened for particle presence and ice quality on a TFS Glacios TEM (200kV), and the best grids were transferred to TFS Titan Krios G4 TEM. Cryo-EM data was collected using TFS Titan Krios G4, equipped with a Cold-FEG and Selectris X energy filter, on a Falcon IV detector in electron counting mode. Falcon IV gain references were collected just before data collection. Data was collected using TFS EPU v2.12.1 using aberration-free image shift protocol (AFIS). Movies were recorded at the nominal magnification of 165kx, corresponding to the 0.726Å pixel size at the specimen level, with defocus values ranging from -0.7 to -2.0 μm. Exposures were adjusted automatically to 60 e-/Å² total dose.

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Cryo-EM Sample Preparation of Adenovirus

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Purified HAdV-F41 was used at 4.2 mg/mL (pH=7.4) and 1.6 mg/mL (pH=4.0). The recombinant HAdV41 penton base (PB41) was purified as described before (27) and used at 1 mg/mL in PBS buffer, supplemented with 5% glycerol. A HAdV41 sample at pH=4.0 was prepared by adding 2 µL of a 0.5 M citric acid / 1 M Na2HPO4 pH=3.4 solution to 25 µL of HAdVF-41 pH=7.4 followed by incubating on ice for 15 minutes. Samples were vitrified on Quantifoil Cu R200 2/2 (Electron Microscopy Sciences, Cat#: Q2100CR2) and Quantifoil Cu R200 1.2/1.3 (Electron Microscopy Sciences, Cat#: Q3100CR1.3) grids for the virus particles and the recombinant protein, respectively. Prior to sample application the grids were glow discharged using a Pelco easiGlow device (Ted Pella Inc.) at 15 mAmp for 30 s. Sample was applied by transferring 3 µL sample onto the glow-discharged side of the grid, blotted and plunge frozen in liquid ethane, using a Vitrobot plunge freezer (Thermo Fisher Scientific), with the following settings: 22 °C, 80% humidity, Blotforce = -20 and a blotting time of 3 s. For HAdVF-41 pH=7.4 sample was applied twice with a blotting step, using the same settings as above, between applications (49) .

Rafie K., Lenman A., Fuchs J., Rajan A., Arnberg N., & Carlson L.A. (2020). The structure of enteric human adenovirus 41 - a leading cause of diarrhea in children.

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