Tecnai biotwin transmission electron microscope

Name: Tecnai biotwin transmission electron microscope
Brand: Thermo Fisher Scientific
SKU: 8yfiCZIBPBHhf-iFJvCt
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The Tecnai Biotwin is a transmission electron microscope designed for biological and materials science applications. It provides high-resolution imaging capabilities for analyzing the structure and composition of samples at the nanoscale level.

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36 protocols using «tecnai biotwin transmission electron microscope»

Electron Microscopy Analysis of Cellular Responses

2025

For TEM images, cells (HFL-1 and H441) were cultured on inserts (50,000 cells/wells) (Costar HTS Transwell-24 System (Individual Transwell inserts with 6.5 mm diameter membranes), Ref 3395, Corning Incorporated, Kennebunk, ME, USA) in 24-well plates. Cells and PCLS were exposed to CuO (1 μg/mL) and soot (10 and 100 μg/mL) for 24 h in corresponding cell culture media. The inserts with cells and PCLS were then fixed in 0.1 M Sorensen’s phosphate buffer, pH 7.4, 1.5% formaldehyde, and 1.5% glutaraldehyde at RT for 1 h. After fixation, the samples were washed three times in 0.1 M Sorensen’s phosphate buffer pH 7.4 and then dehydrated in a graded series of ethanol (50%, 70%, 80%, 90%, and twice in 100%). The samples were embedded in pure Polybed 812, and the polymerized block was sectioned with a Leica UC7 ultramicrotome (Leica Microsystems GmbH, Wetzlar, Germany). The sections were mounted on a pioloform-coated copper Maxtaform H5 grid and stained with 4% Uranyl acetate. Soot and CuO particles in DMEM cell culture media were directly added to the grids as small drops and stained with 4% Uranyl acetate. Images were acquired using a Tecnai BioTWIN transmission electron microscope (Field Electron and Ion (FEI) Company, Hillsboro, OR, USA).

Faruqui N., Orell S., Dondi C., Leni Z., Kalbermatter D.M., Gefors L., Rissler J., Vasilatou K., Mudway I.S., Kåredal M., Shaw M, & Larsson-Callerfelt A.K. (2025). Differential Cytotoxicity and Inflammatory Responses to Particulate Matter Components in Airway Structural Cells. International Journal of Molecular Sciences, 26(2), 830.

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Negative Staining of Extracellular Vesicles

2025

The extracellular vesicles were visualized using negative staining through electron microscopy. The vesicles were purified and placed onto discharged 200-mesh copper EM grids, followed by fixation with 2% paraformaldehyde (PFA). A 1% uranyl acetate solution was used to stain the extracellular vesicles on the EM grids, which were then imaged using an FEI Tecnai BioTwin Transmission Electron Microscope.

Deshpande R.P., Wu K., Wu S.Y., Tyagi A., Smith E.C., Kim J.W, & Watabe K. (2025). MHC-I upregulation by macbecin II in the solid tumors potentiates the effect of active immunotherapy. EMBO Molecular Medicine, 17(4), 797-822.

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Negative Staining and Immunostaining of sEVs

2023

Wu K., Lyu F., Wu S.Y., Sharma S., Deshpande R.P., Tyagi A., Zhao D., Xing F., Singh R, & Watabe K. (2023). Engineering an active immunotherapy for personalized cancer treatment and prevention of recurrence. Science Advances, 9(17), eade0625.

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Visualization and Interaction of Extracellular Vesicles

2023

For negative staining, purified sEVs were placed on the discharged 200 mesh copper EM grids and fixed with 2% paraformaldehyde (PFA). Uranyl acetate solution (1%; Electron Microscopy Sciences) was used to stain the sEVs on the EM grids. The stained sEVs were then imaged by an FEI Tecnai BioTwin Transmission Electron Microscope. For immunostaining of MHC I and MHC II on the surface of sEVs, anti-MHC I (1:100; Santa Cruz Biotechnology) and anti-MHC II (1:100; Santa Cruz Biotechnology) were used to treat the fixed sEVs on the grids. The 10-nm colloidal gold-labeled protein G (1:50; Boster Biological Technology) was then used to label the primary antibodies. Negative staining was performed after immunostaining, and the grid was imaged by transmission electron microscopy.
T cell-p13nsEV interaction assay DC2.4 cells were labeled with PalmGFP (green fluorescent protein) encoded by lentivirus (a gift from X. O. Breakefield). Cells with intense fluorescence were selected by the WOLF cell sorter. The p13nsEV labeled with PalmGFP were isolated by differential ultracentrifuge. To monitor the interaction between p13nsEV and T cells, Vybrant DiD-labeled T cells were seeded in chamber slides, and p13nsEV (20 μg/ml) was added to the media. After 24 hours of incubation, the cells were washed and fixed, followed by sealing the slides with coverslips. The p13nsEV interacting with T cells was examined by observing the GFP signal under the Keyence All-inone Fluorescence Microscope (BZ-X700).

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Corresponding organizations : Wake Forest University, Zhengzhou University, Henan Provincial People's Hospital, Henan University, The University of Texas Southwestern Medical Center

Ultrastructural Analysis of Epididymal Sperm

2023

Collected epididymal sperm cells were washed and pelleted by centrifugation and fixed in 2.5% glutaraldehyde and 2% PFA in 0.1 M cacodylate buffer pH 7.4 for one hour at RT. Fixed sperm pellets were rinsed with 0.1 M cacodylate buffer and spun down in 2% agar. The chilled blocks were trimmed, rinsed in the 0.1 M cacodylate buffer, and replaced with 0.1% tannic acid in the buffer for one hour. After rinsing in the buffer, the samples were post-fixed in 1% osmium tetroxide and 1.5% potassium ferrocyanide in 0.1 M cacodylate buffer for one hour. The post-fixed samples were rinsed in the cacodylate buffer and distilled water, followed by en bloc staining in 2% aqueous uranyl acetate for one hour. Prepared samples were rinsed and dehydrated in an ethanol series to 100%. Dehydrated samples were infiltrated with epoxy resin Embed 812 (Electron Microscopy Sciences), placed in silicone molds, and baked for 24 hours at 60°C. The hardened blocks were sectioned in 60 nm thickness using Leica UltraCut UC7. The sections were collected on grids coated with formvar/carbon and contrast stained using 2% uranyl acetate and lead citrate. The grids were imaged using FEI TecnaiBiotwin Transmission Electron Microscope (FEI, Hillsbroro, OR) at 80 kV. Images were taken using MORADA CCD camera and iTEM (Olympus) software.

Relovska S., Wang H., Zhang X., Fernández-Tussy P., Jeong K.J., Choi J., Suárez Y., McDonald J.G., Fernández-Hernando C, & Chung J.J. (2024). DHCR24-mediated sterol homeostasis during spermatogenesis is required for sperm mitochondrial sheath formation and impacts male fertility over time. bioRxiv.

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Corresponding organizations : Yale University, Korea University, The University of Texas Southwestern Medical Center, HumanN (United States)

Top 5 most cited protocols using «tecnai biotwin transmission electron microscope»

Quantifying Melanosome Fibril Formation

Cited 3 times

For conventional Epon embedding of cell samples, Mel220 transfectants were fixed in 2.5% glutaraldehyde/2% sucrose in 0.1 M sodium cacodylate buffer, pH 7.4 (NaCaCo buffer), for 30 min at room temperature, followed by another 30 min in the same fixation solution at 4°C. Subsequently, cells were rinsed with NaCaCo buffer and further processed as described (Carrithers et al., 2009 (link)).
For cryo–immuno electron microscopy, samples were fixed in 2% paraformaldehyde/0.1% glutaraldehyde in PBS for 15 min at room temperature, followed by another 15 min in the same fixation solution at 4°C. Subsequently, cells were rinsed with PBS and further processed as described (Carrithers et al., 2009 (link)). For immunolabeling, cells were stained with the PMEL-specific antibodies HMB45 or HMB50 at 1:25, followed by protein A–gold (University of Utrecht, Utrecht, Netherlands) or gold anti-mouse conjugate (Jackson ImmunoResearch Laboratories), respectively.
Samples were viewed using a Tecnai BioTWIN transmission electron microscope (FEI, Hillsboro, OR) at 80 kV. Images were collected using Morada CCD and iTEM software (Olympus, Tokyo, Japan).
Epon-embedded EM samples were first inspected to qualitatively determine whether the respective Mel220 transfectant formed conventional melanosomes, gave rise to abnormal fibril-containing organelles, or produced no fibrils at all. To quantify fibril formation, we then counted fibril-containing organelles in 15 arbitrarily chosen cells in one view field. We note that the presence of visible fibrils was the only criterion to count a respective compartment as “fibril containing” with no respect to whether the organelle had a conventional melanosomal or abnormal lysosomal morphology. Thus the numbers (mean) indicated in Figures 3, A–E, and 6, A–E, represent the total number of fibril-containing organelles (not the number of conventional melanosomes) per cell. Within those 15 cells, we also determined the percentage of abnormal fibril-containing organelles (abnormal organelles are more spherical than ellipsoid and contain extensive internal membranes, which are often multilamellar (see Figure 3F, 4 and 5, and Supplemental Figures S2B, 9–14, and S6A, panel B).

Leonhardt R.M., Vigneron N., Hee J.S., Graham M, & Cresswell P. (2013). Critical residues in the PMEL/Pmel17 N-terminus direct the hierarchical assembly of melanosomal fibrils. Molecular Biology of the Cell, 24(7), 964-981.

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Corresponding organizations : Howard Hughes Medical Institute, Yale University, Ludwig Cancer Research, de Duve Institute

Quantitative Ultrastructural Analysis of Hippocampus and Cortex

Cited 2 times

Sections were viewed and photographed on a Tecnai Biotwin transmission electron microscope (FEI, Hillsboro, OR). For each mouse, one block from the hippocampus and one from the somatosensory cortex was analyzed. Met-immunolabeled profiles were counted and classified in the stratum radiatum, 55 µm distal from the hippocampal CA1 pyramidal cell layer, and in superficial layer V of the somatosensory cortex, approximately 350 (P7) to 400 (P14, P21) µm deep to the pial surface. To insure optimum labeling and to allow quantitative comparisons between groups, only grids near the plastic/tissue interface were selected (Milner et al., 2011 (link)). Within these areas, two to three grid squares, representing 6050 to 9075 µm² of neuropil, were sampled; our previous studies have determined that this amount of sampling is sufficient to represent the population (Barker-Gibb et al., 2001 (link); Milner et al., 2001b (link)). Images of all fields containing immunolabeled profiles were acquired at 16,500× using Advanced Microscopy Techniques software (v3.2, Advanced Microscopy Techniques, Woburn, MA).
Cellular profiles were classified according accepted morphological criteria (Peters et al., 1991 ). Briefly, dendritic shafts contained regular microtubular arrays and were usually postsynaptic to axon terminal profiles. Dendritic spines also were usually postsynaptic to axon terminal profiles and sometimes contained a spine apparatus. Unmyelinated axons were smaller than 0.2 µm, round, contained a few small synaptic vesicles and lacked a synaptic junction in the plane of section (Milner and Bacon, 1989 (link); Milner et al., 2001a (link)). Axon terminals had numerous small synaptic vesicles and had a cross-sectional diameter greater than 0.2 µm. Glial profiles were distinguished by the presence of glial filaments and/or gap junctions (astrocytes), by the absence of microtubules, and/or by their tendency to conform to the boundaries of surrounding profiles (Barker-Gibb et al., 2001 (link); Glass et al., 2001 (link)). Ambiguous profiles, which were more prevalent at P7, were classified as unknown. Asymmetric synapses were characterized by the larger size of the post-synaptic density whereas symmetric synapses had pre- and post-synaptic densities of equal size. Appositions were defined as those contacts between profiles in which no interposing glial processes were seen but that lacked any recognizable synaptic specializations. The number of each class of Met-immunolabeled profiles was tabulated and the data presented as mean ± SEM. Significant differences in the distribution between neocortex and hippocampus, and within a region across the three developmental ages, were determined using Pearson’s χ² test. The cross-sectional area and minimum diameter of terminal profiles in which the entire perimeter could be distinguished were measured using Microcomputer Imaging Device software (MCID, Imaging Research Inc., Ontario, Canada).

Eagleson K.L., Milner T.A., Xie Z, & Levitt P. (2013). Synaptic and extrasynaptic location of the receptor tyrosine kinase Met during postnatal development in the mouse neocortex and hippocampus. The Journal of comparative neurology, 521(14), 3241-3259.

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Corresponding organizations : Keck Hospital of USC, Cornell University, Rockefeller University, MIND Research Institute

Immunogold Labeling for Ultrastructural Analysis

Cited 1 time

Tissue samples were cut into 2 mm³ cubes and fixed overnight with paraformaldehyde and glutaraldehyde. Fixed samples were dehydrated in increasing concentrations of alcohol and infiltrated with LR Gold resin™ (EMS, Hatfield, USA). Samples were then embedded in fresh resin with benzyl (EMS) and polymerized with ultraviolet light. Ultrathin sections were cut on an Ultracut UCT Leica™ with diamond knives (Diatome; Biel, Switzerland), picked up with pioloform-coated nickel grids, and processed for immuno-labeling. To block unspecific staining, sections were floated on glycine followed by blocking solution (EMS). Sections were then incubated with the primary antibody (anti-53BP1, anti-γH2AX (pSer139), Bethyl Laboratories; anti-pKu70 (pSer6), anti-pDNA-PKcs, Novus Biologicals, Littleton, USA; anti-H3K9me3, Abcam Inc., Cambridge, USA; anti-CD34, BD Biosciences). After rinsing, secondary antibody conjugated with 6-nm or 10-nm gold-particles (EMS) was applied to the grids. Sections were rinsed and post-fixed with glutaraldehyde. All sections were stained with uranylacetate and examined using a Tecnai Biotwin™ transmission electron microscope (FEI Company, Eindhoven, The Netherlands).

Schuler N, & Rübe C.E. (2013). Accumulation of DNA Damage-Induced Chromatin Alterations in Tissue-Specific Stem Cells: The Driving Force of Aging?. PLoS ONE, 8(5), e63932.

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Corresponding organizations : Saarland University

Ultrastructural Examination of Neuronal Cell Death

Cited 1 time

Primary neurons were grown on poly-l-lysine-coated aclar plastic coverslips. We fixed the cells overnight at 4°C in 0.05M phosphate buffer (PB; pH 7.4) containing 2 % glutaraldehyde and 0.1M sucrose. The coverslips were processed for electron microscopy as previously described with modifications.^{24 (link)} We incubated the coverslips in 2% osmium tetroxide in PB for 1 hour followed by embedding in Epon-812. Ultrathin sections (70nm) using a Leica UC6 ultratome were collected on 400-mesh thin-bar copper grids (Electron Microscopy Sciences, Fort Washington, PA, USA) and counterstained with 5% uranyl acetate and Reynold’s lead citrate. Micrographs were taken on a Tecnai Biotwin transmission electron microscope (FEI, Hillsboro, Oregon, USA). We quantified the percentage of cells displaying necrotic or apoptotic morphology (n=33–43/condition and replicate). Using ImageJ v.1.49 (http://imagej.nih.gov/ij/), we measured mitochondrial size as percentage area of total area of the cytoplasm^{25 (link)} (comprising >1800 mitochondria in total). Analysis was performed by an investigator blinded to treatment group assignment.

Zille M., Karuppagounder S.S., Chen Y., Gough P.J., Phil D., Bertin J., Finger J., Milner T.A., Jonas E.A, & Ratan R.R. (2017). Neuronal Death After Hemorrhagic Stroke In Vitro and In Vivo Shares Features of Ferroptosis and Necroptosis. Stroke, 48(4), 1033-1043.

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Corresponding organizations : Burke Medical Research Institute, Cornell University, MIND Research Institute, Rockefeller University, Yale University

Electron Microscopy of Extracellular Vesicles

EVs were isolated using Vn96 from supernatant from isotype and anti-CD24 treated cells as described above. Two 750-µL aliquots of vesicle-containing media were pooled for each Vn96 pull-down. Pellets were resuspended in 20 µL of PBS and MV were dispersed by digestion overnight with 25 µg proteinase K enzyme^{17 (link)} at 37 °C. The digested samples were centrifuged at 17,000 × g for 15 min to remove undigested Vn96-EV material. Dispersed EV (10 μl) were placed on formvar-carbon electron microscope grids (Canemco; Montreal, Canada) and allowed to dry for 30 min. Grids were floated sample-side down pyrogen-free water. Grids were then fixed with 3.7% paraformaldehyde for 15 min, followed by two washes with water by flotation. Grids were contrasted with 2% uranyl acetate (w/v), followed by one additional water wash. All solutions were filtered using 0.1-µm syringe filters (4611; Pall Corp; Port Washington, NY). Dried grids were then viewed using a Tecnai Biotwin Transmission Electron Microscope (TEM) (FEI; Hillsboro OR). Images were captured using an XR-41 camera with an AMT capture engine V602 (Advanced Microscopy Techniques; Woburn, MA).

Ayre D.C., Chute I.C., Joy A.P., Barnett D.A., Hogan A.M., Grüll M.P., Peña-Castillo L., Lang A.S., Lewis S.M, & Christian S.L. (2017). CD24 induces changes to the surface receptors of B cell microvesicles with variable effects on their RNA and protein cargo. Scientific Reports, 7, 8642.

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Corresponding organizations : Memorial University of Newfoundland, Atlantic Cancer Research Institute, Dalhousie University

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