TEM analyses were performed exactly as described recently [44 (link)]. Briefly, 7.5 µL of 1:100 diluted VLP preparations were mixed with 1 µL aqueous contrasting solution, containing 1% methyl cellulose (w/v; Sigma Aldrich) and 2% uranyl acetate (w/v; Polysciences, Warrington, FL, USA). Following incubation for 10 min, 0.5 µL droplets were placed on 200 mesh copper grids (Plano, Wetzlar, Germany) and dried at room temperature, allowing included VLPs to adhere to the films’ surfaces. Images were taken on a JEM 1400 Plus electron microscope (JEOL, Tokyo, Japan), equipped with a 4096 × 4096 pixel CMOS camera (TemCam-F416; TVIPS, Gauting, Germany), and run at an operating voltage of 120 kV. Image acquisition software EMMENU (Version 4.09.83) was used for taking 16-bit images. Image post-processing was performed with the software ImageJ (Version 1.52b; National Institutes of Health, Bethesda, MD, USA).
Jem 1400 plus electron microscope
Manufactured by JEOL
Sourced in Japan, United States, Germany
The JEM-1400 Plus is a transmission electron microscope (TEM) designed and manufactured by JEOL. It is a versatile and reliable instrument capable of high-resolution imaging and analysis of a wide range of samples. The JEM-1400 Plus features a LaB6 electron source, a high-contrast objective lens, and advanced imaging capabilities, making it a powerful tool for various applications in materials science, nanotechnology, and biological research.
Automatically generated - may contain errors
Lab products found in correlation
Veleta 2k 2k ccd camera, by Olympus (8 mentions)
Diamond knife, by Electron Microscopy Sciences (4 mentions)
Epon 812, by TAAB (3 mentions)
Lead stain solution, by Merck Group (3 mentions)
Temcam f416, by TVIPS (2 mentions)
Zetasizer nano z, by Malvern Panalytical (2 mentions)
Dts1070, by Malvern Panalytical (2 mentions)
Uranyl acetate, by Merck Group (2 mentions)
Dmr light microscope, by Leica (2 mentions)
Ultracut r, by Leica (2 mentions)
Em 14830ruby2, by JEOL (2 mentions)
Veleta, by Olympus (2 mentions)
Uranyl acetate, by Polysciences (1 mentions)
Methylcellulose, by Merck Group (1 mentions)
Fcf300 cu, by Electron Microscopy Sciences (1 mentions)
Goldenhance em, by Nanoprobes (1 mentions)
Em uc7, by Leica (1 mentions)
Mvx10, by Olympus (1 mentions)
Em 902a, by Zeiss (1 mentions)
Uranyl formate, by SPI Supplies (1 mentions)
88 protocols using jem 1400 plus electron microscope
1
Transmission Electron Microscopy of VLPs
2
Electron Microscopy Specimen Preparation
3
Visualizing Tau Aggregates by TEM
4
Quantifying Endocytosis-related Gene Expression
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For the analysis of
endocytosis-related gene expression, cells were incubated in 24-well
plates (at a density of 5 × 104 cells and 5 μg
of NPs per well) for 2 h, 8 and 48 h. After incubation, cells were
harvested, RNA was extracted, and gene expression was quantified as
described above. Specific primers for CAV1 and CLTC (involved in caveolar and chlatrin-mediated endocytosis,
respectively) were used for RT-QCR experiments.
For TEM experiments,
cells were seeded in 10 cm plates at a density of 5 × 106 cells per plate and were incubated with 0.5 mg nanoparticles
per plate for 2, 8, and 48 h. After incubation, cells were prefixed
with 0.5% glutaraldehyde for 15 min and centrifuged at 800 g for 10
min. Pellets were fixed with 2% glutaraldehyde overnight at 4 °C.
For embedding the samples and ultramicrotomy, cell pellets were
washed in Sorenson’s buffer (SB) with 6% sucrose and fixed
in 2% glutaraldehyde for 1 h. Then, pellets were washed twice in SB
and osmicated in 1% osmium tetroxide for 1 h. After washing in SB,
pellets were dehydrated in graded acetone (50%, 70%, 96%, 100%) and
embedded in Epon resin 812. Seventy nanometer ultrathin sections were
collected on mesh nickel grids and examined in a JEOL JEM 1400 Plus
electron microscope. Tissue preparations were photographed by using
a digital camera coupled to the electron microscope.
endocytosis-related gene expression, cells were incubated in 24-well
plates (at a density of 5 × 104 cells and 5 μg
of NPs per well) for 2 h, 8 and 48 h. After incubation, cells were
harvested, RNA was extracted, and gene expression was quantified as
described above. Specific primers for CAV1 and CLTC (involved in caveolar and chlatrin-mediated endocytosis,
respectively) were used for RT-QCR experiments.
For TEM experiments,
cells were seeded in 10 cm plates at a density of 5 × 106 cells per plate and were incubated with 0.5 mg nanoparticles
per plate for 2, 8, and 48 h. After incubation, cells were prefixed
with 0.5% glutaraldehyde for 15 min and centrifuged at 800 g for 10
min. Pellets were fixed with 2% glutaraldehyde overnight at 4 °C.
For embedding the samples and ultramicrotomy, cell pellets were
washed in Sorenson’s buffer (SB) with 6% sucrose and fixed
in 2% glutaraldehyde for 1 h. Then, pellets were washed twice in SB
and osmicated in 1% osmium tetroxide for 1 h. After washing in SB,
pellets were dehydrated in graded acetone (50%, 70%, 96%, 100%) and
embedded in Epon resin 812. Seventy nanometer ultrathin sections were
collected on mesh nickel grids and examined in a JEOL JEM 1400 Plus
electron microscope. Tissue preparations were photographed by using
a digital camera coupled to the electron microscope.
5
Immunogold Labeling of Lzts1 in Brain
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For immunogold EM labeling, cerebral hemispheres were fixed with 4% paraformaldehyde for 3 h at 4 °C, immersed in a graded series of sucrose solutions (10%, 15%, and 20%), and embedded in OCT compound. Frozen sections (10 µm thick) were cut on a cryostat and mounted on silane-coated slides. After blocking, sections were incubated with the primary antibody (20878-1-AP, 1:1000, rabbit anti-Lzts1 pAb, Proteintech Group) in PBS overnight at 4 °C. Sections were incubated with 1.4-nm gold-coupled goat anti-rabbit Fab fragment secondary antibodies (7204, 1:100, Nanoprobes Inc., NY, US) at room temperature for 2 h and postfixed with 2.5% glutaraldehyde, followed by gold enhancement of the immunogold particles using GoldEnhance EM (Nanoprobes). After treatment with 0.5% OsO4, the sections were stained with 1% uranyl acetate, dehydrated and embedded in Epon 812. Ultrathin sections (70-nm thick) were cut using a diamond knife (DiATOME, Hatfield, US) on an ultramicrotome (Leica EM UC7), collected on formvar-coated single-slot copper grids, stained with lead citrate, and examined under a JEM-1400Plus electron microscope (JEOL, Tokyo, Japan).
6
Exosome Ultrastructure Imaging
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Whole exosome extracts were fixed in 2% glutaraldehyde and then absorbed onto formovar-coated grids for 20 min in a dry environment. The grids were examined in a Jeol JEM 1400 Plus electron microscope (JEOL USA, Inc., MA, USA) at 80 kV.
7
Ultrastructural Analysis of Pollen Tube Basal Region
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In vitro-germinated pollen tubes were observed 8 h after germination under the Olympus MVX10 fluorescence stereomicroscope (Olympus) using the RFP filter set. Pollen tubes whose vegetative nucleus and sperm nuclei remained in the basal area were dragged out one-by-one and aligned in one place on the medium with a 27 gauge needle. Those pollen tubes were collected with small pieces of growth medium and fixed with 2% glutaraldehyde and 2% paraformaldehyde in 50 mM cacodylate buffer pH 7.4 at 4 °C. The tissue segments were washed in buffer and post-fixed for 3 h in 2% aqueous osmium tetroxide at 4 °C. The tissue was then dehydrated in a graded ethanol series, transferred into propylene oxide, infiltrated, and embedded in Quetol 651. Cross-sections around the basal area of the pollen tubes (90 nm thickness) were stained with 2% aqueous uranyl acetate and lead citrate and examined using a JEOL JEM 1400Plus electron microscope at 100 kV.
8
Electron Microscopy Sample Preparation
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Cultured cells were fixed with 1% glutaraldehyde in PBS at pH 7.3 for 1 h at 4°C and then postfixed with 1% osmium tetroxide in 0.1 M phosphate buffer at pH 7.3 for 1 h at 4°C, followed by dehydration in a graded series of ethanol. Following dehydration, they were embedded in Epon 812 (TAAB Laboratories Equipment, Berkshire, England) and then observed under a JEM-1400 Plus electron microscope (JEOL, Japan).
9
Electron Microscopy Imaging of Extracellular Vesicles
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Drops of 4 µL of selected EV samples were loaded on formvar or carbon coated 300 mesh copper grids (Plano, Wetzlar, Germany) for 2 min. The grids were washed in double distilled water and contrasted with either 1% uranyl acetate (SPI Supplies, West Chester, USA) or 0.75% uranyl formate (SPI Supplies) for 1 min. Grids were air dried and examined either on a JEM1400 Plus electron microscope (JEOL, Tokyo, Japan) equipped with a LaB6 cathode at 120 kV using a TemCam-XF416® FastScan CCD camera system (TVIPS, Gauting, Germany) for image acquisition or an EM 902A (Zeiss, Oberkochen, Germany) electron microscope at 80 kV with a Morada slow scan CCD camera connected to a PC running ITEM 5.2 capturing software (Olympus SIS, Münster, Germany).
10
Cauda Epididymis Ultrastructure Imaging
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Cauda epididymis samples were prepared for TEM analysis as described previously.25 Sections were examined using a JEM‐1400 plus electron microscope (JEOL, Tokyo, Japan) at 80 kV with a CCD Veleta 2K × 2X camera (Olympus).
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