Spirit transmission electron microscope

Manufactured by Thermo Fisher Scientific

Sourced in Netherlands

The Spirit transmission electron microscope is a high-performance imaging system designed for advanced materials analysis. It utilizes an electron beam to generate detailed images of microscopic structures, enabling users to study the morphology and composition of a wide range of samples.

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Lab products found in correlation

Tryple, by Thermo Fisher Scientific (4 mentions) Em uc6 ultramicrotome, by Leica (3 mentions) Vt 1000m vibratome, by Leica (1 mentions) Em stainer, by Leica (1 mentions) Veleta ccd camera, by Olympus (1 mentions) Morada ccd camera, by Olympus (1 mentions) Tryple express enzyme 1, by Thermo Fisher Scientific (1 mentions) Em fc6 cryo ultramicrotome, by Leica (1 mentions) Tryple express enzyme, by Thermo Fisher Scientific (1 mentions)

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20 protocols using spirit transmission electron microscope

Negative Staining of Purified RS1 Protein

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Negatively stained specimens were prepared from the purified RS1 as previously described [15 (link)]. In brief, 3 μl of the purified protein was adsorbed to glow discharged carbon-coated copper grids, stained with 0.75% (w/v) uranyl formate solution, and the specimens were air dried. These specimens were examined using a Tecnai Spirit transmission electron microscope (FEI) equipped with a LaB6 filament and operated at an accelerating voltage of 120 kV. Micrographs were obtained at a nominal magnification of 49,000X with an FEI Eagle 4K x 4K charge-coupled device (CCD) camera at a defocus value of -1.2 μm using low-dose procedures.

Bush M., Setiaputra D., Yip C.K, & Molday R.S. (2016). Cog-Wheel Octameric Structure of RS1, the Discoidin Domain Containing Retinal Protein Associated with X-Linked Retinoschisis. PLoS ONE, 11(1), e0147653.

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High-Resolution TEM Imaging of Samples

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Samples were prepared as described previously [27 (link)]. Subsequently, the samples were observed using a Spirit transmission electron microscope (FEI Company) operating at 100 kV. Digital images were captured using an AMT imaging system at a high-resolution electron microscopy facility.

Duan J., Huang D., Liu C., Lv Y., Zhang L., Chang F., Zeng X., Li L., Wang W, & Shao G. (2023). USP11-mediated LSH deubiquitination inhibits ferroptosis in colorectal cancer through epigenetic activation of CYP24A1. Cell Death & Disease, 14(7), 402.

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Ultrastructural Imaging of Tissue Samples

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TS tissue samples were collected and then fixed with polyformaldehyde and glutaraldehyde (2.5% in PBS; pH = 7.4) at room temperature for 1 h. After that, tissues were postfixed for 1.5 h with 1% osmium tetroxide insodium cacodylate buffer (0.1 mol L⁻¹), and stained with 1% w/v uranyl acetate for 50 min in double distilled water. After washing and dehydration, samples were embedded in Embed 812 resin and polymerized for 48 h at 60 °C. Polymerized samples were trimmed and sliced to 50 nm sections using a Leica ultramicrotome EM UC6 (Leica, Germany). EM grids coated with carbon film were used to collect on the sections. Spirit transmission electron microscope (FEI Company, the Netherlands) (operating at 100 kV) were used to image for TEM sample grids.

Cui K., Zhu Y., Shi Y., Chen T., Wang H., Guo Y., Deng P., Liu H., Shao X, & Qin J. (2021). Establishment of Trophoblast‐Like Tissue Model from Human Pluripotent Stem Cells in Three‐Dimensional Culture System. Advanced Science, 9(3), 2100031.

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Ultrastructural Analysis of Aged SVZ

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After post-fixation, brains were washed in 0.1 M phosphate buffer (PB) (pH 7,4), cut into 200 μm sections with a VT 1000M vibratome (Leica, Wetzlar, Germany) and treated with 2% osmium tetraoxide in 0.1M PB for 2 h. Then sections were rinsed, dehydrated through a series of ethanol solutions and stained in 2% uranyl acetate at 70% ethanol. Following dehydration, slices were embedded in araldite (Durcupan, Fluka BioChemika, Ronkokoma, NY). To study the SVZ cell organization, serial 1.5 μm semithin sections were cut using a diamond knife and stained with 1% toluidine blue. To identify individual cell types, 60–70 nm ultrathin sections were cut with a diamond knife, stained with lead citrate, and examined under a Spirit transmission electron microscope (FEI Tecnai, Hillsboro, OR). To quantify the SVZ cells in the ventricular wall, we examined the dorsal horn and the entire length of the lateral wall, taking into account the first 20 μm adjacent to the ventricle (0–1 mm anterior to bregma). The quantification was performed in relation to length of the SVZ (cells/mm) and using three different levels per animal (n_young=6, n_aged=6). The analysis was performed with Image Tool software (Evans Technology, Roswell, GA).

Capilla-Gonzalez V., Cebrian-Silla A., Guerrero-Cazares H., Garcia-Verdugo J.M, & Quiñones-Hinojosa A. (2014). Age-Related Changes in Astrocytic and Ependymal Cells of the Subventricular Zone. Glia, 62(5), 790-803.

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TEM Sample Preparation for Peptides

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The samples for TEM were prepared by following a literature protocol.^{15 (link)} For each sample, a 5 μL aliquot of peptide solution (20 μM in 10 mM sodium phosphate pH 7.4 or 10 mM Tris-maleate pH 6.4) was placed onto the carbon-coated surface of a copper grid (400 square mesh, Electron Microscopy Sciences). After 1 min, the grid was washed with a 5 μL aliquot of Milli-Q water, stained with a 5 μL aliquot of 2% uranyl acetate (UA, Electron Microscopy Sciences) in Milli-Q water three times, and air-dried for at least 15 min before imaging. A FEI Technai Spirit Transmission Electron Microscope was employed for all TEM imaging (W.M. Keck Microscopy Facility, Whitehead Institute, Cambridge, MA). TEM images were obtained with at least two independently prepared peptides and representative images are presented.

Chairatana P., Chu H., Castillo P.A., Shen B., Bevins C.L, & Nolan E.M. (2015). Proteolysis triggers self-assembly and unmasks innate immune function of a human α-defensin peptide †Electronic supplementary information (ESI) available: Tables of amino acid sequences and characterization of peptides employed in this work, calculated sedimentation coefficients, sedimentation velocity results, and sedimentation equilibrium results for proHD6. Figures of characterization of HD6 in ileal fluid, working model of HD6 maturation, SDS-PAGE, HPLC traces, antibacterial activity assays and in vitro trypsin-catalyzed degradation of recombinant proHD6, characterization of products from trypsinized proHD6, TEM of trypsinized proHD6 in Tris-maleate buffer and at different time points, SEM of bacterial agglutination by trypsinized proHD6, cytotoxicity studies of proHD6 and HD6 against T84 cells, sedimentation velocity analysis, and sedimentation equilibrium analysis of proHD6. See DOI: 10.1039/c5sc04194e Click here for additional data file.. Chemical Science, 7(3), 1738-1752.

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Electron Microscopy Sample Preparation

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Cells were fixed with a solution containing 2.5% glutaraldehyde overnight at 4 °C. After fixation, the samples were washed with 0.1 M sodium phosphate buffer for three times, and were post-fixed in 1.0% osmium tetroxide for 1 h. Then the samples were blocked with 1% uranyl acetate, dyed for 1 h, and dehydrated through a graded series of acetone to 100% and placed into liquid Spurr epoxy resin for saturating overnight at 37 °C. Then, the samples were polymerized in a 65 °C oven for 2 days and the embedded samples were cut using Leica ultramicrotome EM UC6 (Leica). Later, the samples were stained with uranyl acetate and lead citrate in a Leica EM Stainer and imaged under a Spirit Transmission Electron Microscope (FEI Company) operating at 120 kV.

Wang S., Hu B., Ding Z., Dang Y., Wu J., Li D., Liu X., Xiao B., Zhang W., Ren R., Lei J., Hu H., Chen C., Chan P., Li D., Qu J., Tang F, & Liu G.H. (2018). ATF6 safeguards organelle homeostasis and cellular aging in human mesenchymal stem cells. Cell Discovery, 4, 2.

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Quantitative Ultrastructural Analysis of Remyelination

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At 14 days post-lysolecithin injection, mice were transcardially perfused with 2% PFA/2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer with 2 mM CaCl₂. Spinal cords were processed and EM performed with a FEI Spirit transmission electron microscope, according to standard procedures (Davalos et al., 2012 (link)). We examined ∼300 axons (∼75 axons per mouse, N = 4 mice per group) within the rim of active remyelination at the lesion edge, and calculated g-ratios as the diameter of the axon divided by the diameter of the axon plus the surrounding myelin sheath. Unmyelinated axons were excluded from G-ratio graph and analysis.

Petersen M.A., Ryu J.K., Chang K.J., Etxeberria A., Bardehle S., Mendiola A.S., Kamau-Devers W., Fancy S.P., Thor A., Bushong E.A., Baeza-Raja B., Syme C.A., Wu M.D., Rios Coronado P.E., Meyer-Franke A., Yahn S., Pous L., Lee J.K., Schachtrup C., Lassmann H., Huang E.J., Han M.H., Absinta M., Reich D.S., Ellisman M.H., Rowitch D.H., Chan J.R, & Akassoglou K. (2017). Fibrinogen activates BMP signaling in oligodendrocyte progenitor cells and inhibits remyelination after vascular damage. Neuron, 96(5), 1003-1012.e7.

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Correlative Light-Electron Microscopy (CLEM)

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CLEM is described in full in Supplemental Experimental Procedures. In brief, endogenous CPAP was depleted by RNAi for 72 hr, simultaneous with induction of the transgene. For dual marker experiments, cells were transfected with a tagRFP-Centrin 1 expression vector 16 hr prior to fixation. Cells were fixed, then washed thoroughly with 0.1 M cacodylate buffer (pH 7.4), and imaged by light microscopy. Immediately afterward, samples were post-fixed for 40 min in 1.0% osmium tetroxide, then 30 min in 1.0% uranyl acetate in water, before being dehydrated through increasing concentrations of alcohol and then embedded in Durcupan ACM resin (Fluka). The coverslips were then placed face down on a glass slide coated with mold-releasing agent (Glorex), with approximately 1 mm of resin separating the two. These regions were mounted on blank resin blocks with acrylic glue and trimmed with glass knives to form a block ready for serial sectioning. Series of between 150 and 300 thin sections (50 nm thickness) were cut with a diamond knife mounted on an ultramicrotome (Leica UC7). These sections were contrasted with lead citrate and uranyl acetate and images taken using an FEI Spirit transmission electron microscope equipped with an Eagle CCD camera.

Sharma A., Aher A., Dynes N.J., Frey D., Katrukha E.A., Jaussi R., Grigoriev I., Croisier M., Kammerer R.A., Akhmanova A., Gönczy P, & Steinmetz M.O. (2016). Centriolar CPAP/SAS-4 Imparts Slow Processive Microtubule Growth. Developmental Cell, 37(4), 362-376.

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Ultrastructural Analysis of hMSCs

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WT and SIRT7-deficient hMSCs at middle passage (P6) were harvested by TrypLE™ (Thermo Fisher Scientific) and centrifuged at 500 ×g for 5 min (RT). The collected pellets were fixed with 4% PFA in PBS, pH 7.4, on ice overnight. Using graded series of ethanol, cells were dehydrated and then infiltrated with Lowicryl resin HM20. Images were collected with a Spirit transmission electron microscope (FEI Company) operating at 100 kV.

Bi S., Liu Z., Wu Z., Wang Z., Liu X., Wang S., Ren J., Yao Y., Zhang W., Song M., Liu G.H, & Qu J. (2020). SIRT7 antagonizes human stem cell aging as a heterochromatin stabilizer. Protein & Cell, 11(7), 483-504.

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Ultrastructural Analysis of ZKSCAN3 hMSCs

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ZKSCAN3^+/+ and ZKSCAN3^-/^- hMSCs were collected enzymatically by TrypLE (Gibco) and centrifuged at 1500 g for 5 min at RT. The pellets were fixed with 4% glutaraldehyde in PBS at 4°C overnight. Samples were dehydrated in a graded series of ethanol, infiltrated and embedded in Lowicryl resin HM20. Two hundred nanometre sections were obtained and imaged by a Spirit transmission electron microscope (FEI Company) operating at 100 kV.

Hu H., Ji Q., Song M., Ren J., Liu Z., Wang Z., Liu X., Yan K., Hu J., Jing Y., Wang S., Zhang W., Liu G.H, & Qu J. (2020). ZKSCAN3 counteracts cellular senescence by stabilizing heterochromatin. Nucleic Acids Research, 48(11), 6001-6018.

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