In accordance to a procedure described previously [56 (link)], vitrified specimens for cryo-TEM were prepared by a blotting procedure using an EM GP grid plunger (Leica, Wetzlar, Germany), and an atmospheric chamber with controlled humidity (80%) and temperature (T = 22 °C). For cryo-fixation, a drop of the sample solution (c = 3 mM) was placed onto an EM grid that is coated with a holey carbon film (C-flat, Protochip Inc., Raleight, NC, USA). After the excess solution was removed with a filter paper, vitrification of the thin film was achieved by rapid plunging of the grid into liquid ethane. The vitrified specimens were kept at temperatures below 108 K during storage, transfer to the microscope, and investigation. Specimens were examined with a Libra 120 Plus TEM (Carl Zeiss Microscopy GmbH, Jena, Germany) operating at 120 kV The microscope was equipped with a Gatan 626 cryotransfer system and with a BM-2k-120 Dual-Speed on axis SSCCD-camera (TRS, Moorenweis, Germany).
Libra 120 plus tem
Manufactured by Zeiss
Sourced in Germany
The Libra 120 Plus TEM is a transmission electron microscope (TEM) manufactured by Zeiss. It is designed to provide high-resolution imaging and analytical capabilities for materials science and life science research. The Libra 120 Plus TEM features advanced electron optics and a user-friendly interface, enabling researchers to obtain detailed information about the structure and composition of their samples.
Automatically generated - may contain errors
Lab products found in correlation
Ultrascan 4 k 4 k ccd, by Ametek (3 mentions)
K2 summit, by Ametek (3 mentions)
Em gp grid plunger, by Leica (2 mentions)
Gatan tomography module, by Ametek (2 mentions)
Tecnai tf20 tem, by Ametek (2 mentions)
Ultrascan 4kx4k ccd, by Ametek (2 mentions)
Apollo 11lb913 microplate reader, by Berthold Technologies (1 mentions)
Wizard2 automatic gamma counter, by PerkinElmer (1 mentions)
Zetasizer nano z, by Malvern Panalytical (1 mentions)
Malvern zetasizer nano s90, by Malvern Panalytical (1 mentions)
Tecnai tf20 tem, by Thermo Fisher Scientific (1 mentions)
Cf200 cu, by Electron Microscopy Sciences (1 mentions)
Em gp plunge freezer, by Leica (1 mentions)
Titan krios g2 tem, by Thermo Fisher Scientific (1 mentions)
18 protocols using libra 120 plus tem
1
Cryo-TEM Specimen Vitrification Procedure
2
Cryo-EM Tilt Series Imaging of IDL
3
Nanoparticle Characterization and Evaluation
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The size and morphology of nanoparticles were examined using a Zeiss Libra 120 Plus TEM operating at 120 kV (Zeiss, Stuttgart, Germany). The DLS method was used to analyze the hydrodynamic size of the synthesized nanoparticles and PEG conjugates. The hydrodynamic diameter and zeta potential measurements were conducted in 1 mM PBS pH 7.4 buffer using a Zetasizer Nano ZS (Malvern Panalytical, Malvern, Worcestershire, UK). The magnetic properties of the obtained nanoparticles were tested using a QD VSM vibrating magnetometer by NanoMagnetics Instruments (Oxford, UK) operating in the range of –2.0 to +2.0 T. Measurements were carried out in the temperature range from 100–300 K with an accuracy of 0.01 K. The SAR values were determined using MaNIaC Controller software supplied with D5 series equipment. The MTS assay absorbance values were evaluated at 490 nm via an Apollo 11LB913 microplate reader (Berthold, Bad Wildbad, Germany). The radioactivity of samples was measured using Wizard® 2 automatic gamma counter (Perkin Elmer, Waltham, MA, USA).
4
Liquid-phase TEM imaging of biological samples
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The liquid phase TEM grids, which contained label-free biological samples, were examined using a Zeiss Libra 120 Plus TEM (Carl Zeiss NTS) at room temperature. The TEM was operated at a high tension of 120 kV with a 20 eV slit for zero-loss in-column energy filtering. Micrographs were acquired using a Gatan UltraScan 4k × 4k charge coupled device (CCD) with a defocus up to 12 µm and magnification from 1000× to 80,000× (corresponding to pixel sizes of 107 Å to 1.48 Å in the specimen). To assess radiation damage, the GroEL sample was subjected to an illumination dose ranging from ~1 to ~36 e−Å−2 under magnification of 40,000× to 80,000× and defocus up to −3 µm. Approximately 300 micrographs were acquired under low-dose conditions using a magnification of 80,000x and a defocus range of ~0.1 to ~1.0 µm. For the HeLa cells, approximately 100 micrographs were acquired under low-dose conditions at a magnification of 1000× to 20,000×.
5
Structural Analysis of Reconstituted HDL
6
Cryo-EM Imaging of Protein Complexes
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OpNS samples were examined by using a Zeiss Libra 120 Plus TEM (Carl Zeiss NTS) operated at 120 kV high tension with a 10–20 eV energy filter. The OpNS micrographs were acquired under defocus at ~0.6 μm to ~0.9 μm and a dose of ~40–90 e−Å−2 using a Gatan UltraScan 4 K X 4 K CCD under a magnification of 80 kx (each pixel of the micrographs corresponds to 1.48 Å in specimens). The contrast transfer function (CTF) of each micrograph was examined by using ctffind3 software98 (link) and corrected by using the “TF CTS” command in SPIDER99 (link) software or GCTF100 (link) after the X-ray speckles were removed. Particles were then selected from the micrographs with a box size of 192 × 192 by using boxer (EMAN78 (link) software). All particles were masked by using a round mask generated from SPIDER software after a Gaussian high-pass filtering. The reference-free class averages of particles were obtained by using refine2d (EMAN software) based on 13,546 particles of hole-hole homodimer and 4,961 particles of NISTmAb.
7
Physicochemical Characterization of PLGA Nanoparticles
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The particle size, zeta potential, and polydispersity index (PDI) of blank PLGA, S@PLGA, and S@C-PLGA NPs were measured using a Malvern Zetasizer Nano S90 (Malvern Instruments Ltd., UK). The samples were prepared by diluting the nanoparticle dispersion in deionized water and sonicating for 30 s before measurement. The size and zeta potential of the nanoparticles with different concentrations of Stattic loaded were also measured after incubation in cell culture medium for 24 h.
For transmission electron microscope (TEM) sample preparation, the diluted sample was dropped onto a copper grid, stained with 0.25% uranyl acid replacement (UAR) solution for 60 s, and allowed to dry. The morphology of the particles was observed using a Carl Zeiss Libra 120 PLUS TEM (Oberkochen, Germany) at 120 kV.
For transmission electron microscope (TEM) sample preparation, the diluted sample was dropped onto a copper grid, stained with 0.25% uranyl acid replacement (UAR) solution for 60 s, and allowed to dry. The morphology of the particles was observed using a Carl Zeiss Libra 120 PLUS TEM (Oberkochen, Germany) at 120 kV.
8
Tilt Series Cryo-TEM Imaging Protocol
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The TEM holder was tilted at angles ranging from −45° to +45° in 1.5° increments, and imaging was controlled using Gatan tomography software (Zeiss Libra 120 Plus TEM). The TEM was operated at 120 kV high tension with a 20-eV energy filter. The tilt series was acquired under low defocus conditions (<1 μm) using a Gatan UltraScan 4 K X 4 K CCD under a magnification of 80 kx (each pixel of the micrographs corresponds to 1.48 Å in specimens). The total electron doses were 2,800–6,600 e−Å−2. The micrographs were initially aligned using the IMOD103 (link) software package. The CTF was then corrected using TomoCTF104 (link). The tilt series of the particles in square windows of 256 × 256 pixels (~38 nm) were semi-automatically tracked, windowed using IPET software39 (link), and finally binned by 2 to reduce computation time in the subsequent reconstruction.
9
Liquid-Phase TEM Analysis of Biological Samples
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The liquid-phase TEM grids containing label-free biological samples were examined at room temperature using a Zeiss Libra 120 Plus TEM (Carl Zeiss NTS) operated at a high tension of 120 kV with 20 eV in-column energy filtering. Micrographs were acquired on a Gatan UltraScan 4k × 4k CCD using a defocus up to 12 μm and magnification from 1,000× to 80,000× (each pixel of the micrographs corresponded to 107 Å to 1.48 Å, respectively, in the specimen). The radiation damage on the GroEL sample was tested using an illumination dose in the range from ~1 to ~36 e−/Å2 under magnification of 40,000× −80,000× and defocus up to −3 μm. Approximately 300 micrographs were acquired under low-dose conditions, a magnification of 80,000x, and defocus of ~0.1 – ~1.0 μm. Approximately 100 micrographs of HeLa cells were acquired under low-dose conditions at a magnification of 1,000× – 20,000×.
10
Transmission Electron Microscopy of OpNS
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OpNS samples were imaged using a Zeiss Libra 120 Plus TEM (Carl Zeiss NTS) operated at 120 kV high tension with a 20 eV energy filter. The untilt micrographs were acquired at Scherzer defocus and with the dose in the range of ~ 10–25 e− Å−2 at a magnification of 50 kx (each pixel of the micrographs corresponds to 0.24 nm in specimen), or a dose of ~ 60–90 e− Å−2 under a magnification of 125 kx (each pixel of the micrographs corresponds to 0.096 nm in specimens) using a Gatan UltraScan 4 K × 4 K charge-coupled device (CCD). The acquired micrographs were Gaussian high-pass filtered to 150 nm and low-pass filtered to 1 nm after the X-ray speckles were removed.
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