Eclipse ti system

Manufactured by Nikon

Sourced in Japan

The Eclipse Ti System is a high-performance microscope platform designed for advanced imaging applications. It features a modular and versatile design, allowing for the integration of various components and accessories to meet the specific needs of researchers and scientists.

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Lab products found in correlation

Nis elements version 3, by Nikon (3 mentions) Lsm 710, by Zeiss (2 mentions) Alexa fluor 488 or 594 conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions) Flag m2 antibody, by Merck Group (1 mentions) Anti fade medium, by Agilent Technologies (1 mentions) Fitc conjugated secondary antibody, by Santa Cruz Biotechnology (1 mentions) Alexa fluor 633 goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Alexa fluor 488 goat anti mouse igg, by Thermo Fisher Scientific (1 mentions) X tremegene 9 transfection reagent, by Merck Group (1 mentions) Alexa fluor 594 goat anti mouse igg, by Thermo Fisher Scientific (1 mentions) Coolsnap camera, by Teledyne (1 mentions) Ab6302, by Abcam (1 mentions) Hydromount, by Electron Microscopy Sciences (1 mentions)

19 protocols using eclipse ti system

Airway Inflammation Cell Analysis

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Cellular recruitment into the airway lumen was assessed in the BALF. The lungs were perfused twice with 1 mL of sterile PBS (pH 7.4) to obtain the BALF. Total cell counts were determined by trypan blue staining 50 μL of BALF and counting viable cells using a hemocytometer. Differential cell counts were performed on cytocentrifuge preparations (Cytospin 3; Thermo Shandon, Pittsburgh, Pa) stained with Wright-Giemsa. A total of 300 cells were counted per sample using light microscopy. Formalin-fixed lungs were embedded in paraffin, sectioned at 4 μm thickness, and stained with hematoxylin and eosin or Masson’s trichrome. Microscopy was performed on a NIKON Eclipse Ti System.^{8 (link),12 (link)}

Liu Z., Chen H., Wang P., Li Y., Wold E.A., Leonard P.G., Joseph S., Brasier A.R., Tian B, & Zhou J. (2020). Discovery of Orally Bioavailable Chromone Derivatives as Potent and Selective BRD4 Inhibitors: Scaffold Hopping, Optimization, and Pharmacological Evaluation. Journal of medicinal chemistry, 63(10), 5242-5256.

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Quantifying Nuclear Localization of Proteins

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Cells were transfected with indicated plasmids on Poly-D-Lysine-coated chambered coverglass (Thermo) and cultured in phenol red-free medium supplemented with 10% charcoal-stripped fetal bovine serum. For IF staining, at 48 hr after transfection, cells were fixed with 4% paraformaldehyde, and incubated with a pan-AR antibody (N-20, Santa Cruz; 1:500) or a FLAG antibody (M2, Sigma; 1:1000) overnight at 4°C and subsequently with Alexa Fluor 488- or 594-conjugated secondary antibody (Invitrogen; 1:1000) for 1 hr at room temperature in the dark. Nuclei were then stained with 4′,6-diamidino-2-phenylindole (DAPI). A Nikon ECLIPSE Ti system with a 40X oil-immersion objective was used for confocal imaging. An average of 6 fields with ~10 cells per field was captured for each group, and image analysis was carried out with the use of the NIH Image J Software. The intensities of the nucleus and the whole cell fluorescence signals were quantitated for calculation of the percentage of nuclear localization.

Zhan Y., Zhang G., Wang X., Qi Y., Bai S., Li D., Ma T., Sartor O., Flemington E.K., Zhang H., Lee P, & Dong Y. (2016). Interplay Between Cytoplasmic and Nuclear Androgen Receptor Splice Variants Mediate Castration Resistance. Molecular cancer research : MCR, 15(1), 59-68.

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Immunofluorescence Analysis of OGG1 and 8-oxodG

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Cells on microscope coverslips were fixed in 4% paraformaldehyde at 4°C and then permeabilized with Triton X100 for 30 min at 37°C. The cells were then incubated for overnight at 4°C with primary Ab to OGG1 (1:1000), 8-oxodezoxyguanosine (1:400). After washing (PBS-Tween 20: PBS-T) cells were incubated for 1 h at room temperature with Alexa 488-conjugated secondary Abs. Nuclei of cells were stained for 15 min with DAPI (4′6-diamidino-2-phenylindole dihydrochloride; 10 ng/mL; blue). Cells were then mounted in anti-fade medium (Dako Inc. Carpinteria, CA) on a microscope slide. Microscopy was performed on a NIKON Eclipse Ti System operated via NIS-Elements Software AR Ver3.22.09 for 64 bit (NIKON Instruments, Tokyo, Japan).

Aguilera-Aguirre L., Bacsi A., Radak Z., Hazra T.K., Mitra S., Sur S., Brasier A.R., Ba X, & Boldogh I. (2014). Innate Inflammation Induced by the 8-Oxoguanine DNA Glycosylase-1-KRAS-NF-κB Pathway. Journal of immunology (Baltimore, Md. : 1950), 193(9), 4643-4653.

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Mitochondrial Morphology and Membrane Potential

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Mitochondrial morphology was assessed under four conditions: control, Mcl-1S3, control plus Drp1, and Mcl-1S3 plus Drp1. Under each condition, cells were transfected with 1 μg of mtDSred (excitation/emission: 556/586 nm) and stained with 1 μM calcein (excitation/emission: 495/515 cm) for 10 min at 37°C to mark the mitochondria and to normalize for cell volume.
The Ψ_m was assessed under two conditions: control and Mcl-1S3. Under each condition, cells were incubated with 10 nM TMRM (excitation/emission: 548/573 nm) for 20 min at 37°C. Steady-state and poststimulation (FCCP 10 μM) dye intensities were quantified.
All of the experiments were performed on a confocal Nikon Eclipse T_i system. Fluorescent images were captured and analyzed using NisElements 3.2 for membrane potential and Imaris 4.0 software for morphology. To improve image resolution, Z-series acquisitions were deconvolved using Huygens Essential 3.3 software.

Morciano G., Giorgi C., Balestra D., Marchi S., Perrone D., Pinotti M, & Pinton P. (2016). Mcl-1 involvement in mitochondrial dynamics is associated with apoptotic cell death. Molecular Biology of the Cell, 27(1), 20-34.

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Immunofluorescence Analysis of Cell Cycle Markers

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Cell on coverslips were fixed in 4% paraformaldehyde at 4°C for 15 min. The air-dried cells were permeabilized with acetone-methanol (ratio: 1:1). The cells were then incubated for 60 min at 37°C with primary antibodies to TP53-serine¹⁵, y-pH2A.X, p16^INK4a or p21^WAF/CIP1. After washing cells (3 times for 15 min) with PBS-Tween 20 (PBS-T), they were incubated with affinity-purified FITC-conjugated secondary antibodies (Santa Cruz Biotechnology) for 1 h at room temperature. Nuclei of cells were stained with 10 ng/ml DAPI (4′6-diamidino-2-phenylindole dihydrochloride) for 15 min and then mounted in antifade medium (Dako Inc. Carpinteria, CA) on microscope slides. Microscopy was performed on a NIKON Eclipse Ti System using Nikon NIS Elements Version 3.5 (NIKON Instruments, Tokyo, Japan). Magnification: ×125.

German P., Saenz D., Szaniszlo P., Aguilera-Aguirre L., Pan L., Hegde M.L., Bacsi A., Hajas G., Radak Z., Ba X., Mitra S., Papaconstantinou J, & Boldogh I. (2016). 8-oxoguanine DNA glycosylase1–driven DNA repair --a paradoxical role in lung aging. Mechanisms of ageing and development, 161(Pt A), 51-65.

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Quantifying Cellular Senescence and Lipofuscin

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Senescent associated (SA)–galactosidase (SA-β-gal) expression was undertaken as described previously (Dimri et al., 1995 (link)). Briefly, cells were fixed in 2% formaldehyde/0.2% glutaradehyde then incubated at 37 °C with β-gal staining solution (1 mg of X-Gal per ml in dimethylformamide) in buffer containing 40 mM citric acid/sodium phosphate; 5 mM potassium ferrocyanide; 5 mM potassium ferricyanide; 150 mM NaCl and 2 mM MgCl2). Cultures were evaluated after 16 h incubation. Lipofuscin: cells on microscope cover-slips were placed in a thermo-controlled microscopic chamber and cellular autofluorescence were visualized at 580–630 nm. Microscopy and co-localization was performed on a NIKON Eclipse Ti System using Nikon NIS Elements Version 3.5 (NIKON Instruments, Tokyo, Japan). Magnification: ×125.

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Quantifying Pulmonary Fibrosis in Tissue Sections

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Formalin-fixed lungs were embedded in paraffin, sectioned at a 4 μm thickness, and stained with hematoxylin and eosin or Masson's trichrome. Microscopy was performed on a NIKON Eclipse Ti System ^{16 , 17 (link)}. Pulmonary fibrosis was graded using a modified Ashcroft scoring method ^{16 , 17 (link)}. To determine aggregate histopathology score, the entire left and right lungs were longitudinally sectioned. Each lung was scored separately (score range, 0 to 9) and scores combined (total score range, 0 to 18) ^{16 , 17 (link)}.

Tian B., Hosoki K., Liu Z., Yang J., Zhao Y., Sun H., Zhou J., Rytting E., Kaphalia L., Calhoun W.J., Sur S, & Brasier A.R. (2018). Mucosal Bromodomain-Containing Protein 4 (BRD4) Mediates Aeroallergen-induced Inflammation and Remodeling. The Journal of allergy and clinical immunology, 143(4), 1380-1394.e9.

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Histological Lung Tissue Analysis

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The formalin-fixed lungs were embedded in paraffin, sectioned at a 4-μm thickness, and stained with hematoxylin and eosin or Masson’s trichrome. Microscopy was performed on a Nikon Eclipse Ti system, similar as described in (22 (link)).

Ren J., Wu W., Zhang K., Choi E.J., Wang P., Ivanciuc T., Peniche A., Qian Y., Garofalo R.P., Zhou J, & Bao X. (2021). Exchange Protein Directly Activated by cAMP 2 Enhances Respiratory Syncytial Virus-Induced Pulmonary Disease in Mice. Frontiers in Immunology, 12, 757758.

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Airway Immune Cell Analysis

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Cellular recruitment into the airway lumen was assessed in the BALF. Lungs were perfused twice with 1 mL of sterile PBS (pH 7.4) to obtain the BALF. Total cell counts were determined by trypan blue staining of 50 μL of BALF and counting viable cells using a hemocytometer. Differential cell counts were performed on cytocentrifuge preparations (Cytospin 3; Thermo Shandon, Pittsburgh, Pa) stained with Wright-Giemsa. A total of 300 cells were counted per sample using light microscopy. Formalin-fixed lungs were embedded in paraffin, sectioned at a 4 μm thickness, and stained with hematoxylin and eosin or Masson’s trichrome. Microscopy was performed on a NIKON Eclipse Ti System.

Liu Z., Li Y., Chen H., Lai H.T., Wang P., Wu S.Y., Wold E.A., Leonard P.G., Joseph S., Hu H., Chiang C.M., Brasier A.R., Tian B, & Zhou J. (2022). Discovery, X‑ray Crystallography, and Anti-inflammatory Activity of Bromodomain-containing Protein 4 (BRD4) BD1 Inhibitors Targeting a Distinct New Binding Site. Journal of medicinal chemistry, 65(3), 2388-2408.

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Lung Lavage and Histological Analysis

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Lungs were perfused twice with 0.75 mL of sterile PBS (pH 7.4) to obtain the BALF. Total cell counts were determined by trypan blue staining 50 µL of BALF and counting viable cells using a hemocytometer. Differential cell counts were performed on cytocentrifuge preparations (Cytospin 3; Thermo Shandon, Pittsburgh, Pa) stained with Wright-Giemsa. A total of 300 cells were counted per sample using light microscopy. Formalin-fixed lungs were embedded in paraffin, sectioned at a 4 µm thickness, and stained with hematoxylin and eosin or Masson's trichrome. Microscopy was performed on a NIKON Eclipse Ti System (Tian et al., 2016 (link)).

Tian B., Liu Z., Yang J., Sun H., Zhao Y., Wakamiya M., Chen H., Rytting E., Zhou J, & Brasier A.R. (2018). Selective Molecular Antagonists of the Bronchiolar Epithelial NFκB-Bromodomain-Containing Protein 4 (BRD4) Pathway in Viral-induced Airway Inflammation. Cell reports, 23(4), 1138-1151.

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