The leg samples were decalcified in 10% Ethylene Diamine Tetraacetic Acid for 4 weeks at RT and then processed for embedding in paraffin blocks. Tissues were sectioned into 5‐μm thick slices for TRAcP staining, hematoxylin and eosin (H&E) staining, and IHC. Tibia femurs sections underwent H&E staining. Stained sections were examined using an Eclipse TS100 inverted light microscope (Nikon Instruments Inc). The total number of TRAcP‐positive cells and the osteoclast surface per bone surface (Oc.S/BS) were calculated using ImageJ software. IHC analysis was performed with heat‐induced antigen retrieval in sodium‐citrate buffer (Dako). Primary antibodies used was anti‐phospho‐PI3K at 1:100. Biotinylated secondary antibody used was the EnVision + system HRP kit (Dako), and then nuclei were stained with hematoxylin.
Eclipse ts100 inverted light microscope
Manufactured by Nikon
Sourced in Japan
The Eclipse TS100 is an inverted light microscope manufactured by Nikon. It is designed for basic observation and documentation of samples.
Automatically generated - may contain errors
Lab products found in correlation
Envision system hrp kit, by Agilent Technologies (1 mentions)
Sodium citrate buffer, by Agilent Technologies (1 mentions)
Coolpix digital camera, by Nikon (1 mentions)
Mayer haematoxylin solution, by Merck Group (1 mentions)
Oil red o solution, by Merck Group (1 mentions)
Mda mb 231, by European Collection of Authenticated Cell Cultures (1 mentions)
U 87 mg, by European Collection of Authenticated Cell Cultures (1 mentions)
A2780cis, by Merck Group (1 mentions)
Cisplatin, by Merck Group (1 mentions)
Mcf 7, by European Collection of Authenticated Cell Cultures (1 mentions)
A2780, by European Collection of Authenticated Cell Cultures (1 mentions)
4 protocols using eclipse ts100 inverted light microscope
1
Osteoclast Histomorphometric Analysis
2
BPA-Induced Lipid Accumulation in THLE-2 Cells
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Oil Red O staining was performed to examine cytoplasmic lipid accumulation in BPA-treated THLE-2 cells. THLE-2 cells were trypsinised and sub-cultured at 37̊C for 24 h with fresh growth medium in 24-well plates at a density of 2x105 cells/well. Subsequently, THLE-2 cells were treated with 35 µg/ml BPA at 37̊C for 24, 48 and 72 h. The BPA-treated cells were fixed with cold 4% paraformaldehyde in PBS (pH 7.4) for 10 min at room temperature before being stained with Oil Red O solution (Sigma Aldrich; Merck KGaA) at 37˚C for 2 h. The stained cells were counterstained with Mayer haematoxylin solution at room temperature for 5 min (Sigma Aldrich; Merck KGaA) and mounted with an aqueous mounting medium. The morphology of the stained cells was examined using an Eclipse TS100 Inverted Light Microscope (Nikon Corporation) at 200x magnification and images were captured with a Nikon COOLPIX digital camera (Nikon Corporation).
3
Clonogenic Assay for H1299 Cells
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For standard clonogenic assay, molten-agarose (45°C) was added to complete media (DMEM/F12 with 10% FBS and 1% PSA) at 0.6% final-concentration. This DMEM/F12-agarose mix was quickly pipetted into a 12-well plate and allowed to reach the room temperature. Next, H1299 cells (2.0 x 105 cells/well) were similarly suspended in 0.3%-agarose and were quickly pipetted onto the base agarose layer. This layer was again allowed to completely solidify at room temperature. Wells were then treated with either vehicle-control, DBeQ (50μM, positive-control), DDN or DDNDBeQ (50μM). After the treatment, the plate was kept at 37°C in a CO2-incubator till colonies were visible (~4 days). Images were captured using the Nikon Eclipse TS100 inverted light microscope (10x phase objective). The average area of clearly visible colonies were counted and quantified using the Infinity Analyze software.
4
Cultivation and Characterization of Cancer Cell Lines
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Human ovarian cancer cell lines A2780 (cisplatin sensitive human epithelial ovarian cancer cell line) and A2780cis (the resistant variant), human breast cancer cell lines MCF7 and MDA-MB-231, and human glioblastoma cancer cell lines U-87 MG and T98G were purchased from European Collection of Authenticated Cell Cultures (England). A2780 and A2780cis cell lines were grown in complete RPMI-1640 medium containing 10 % FBS, 2 mM glutamine, 0.1 mg/ml each of penicillin and streptomycin. To maintain A2780cis resistance to cisplatin, 1 μM of cisplatin (Sigma-Aldrich, St. Louis, USA) was added to the media every 3 passages. MCF7 and MDA-MB-231 cell lines were grown in EMEM containing 10 % FBS, 2 mM glutamine, 1 % non-essential amino acids (NEAA), 0.1 mg/ml each of penicillin and streptomycin. U-87 MG and T98G cell lines were cultured in EMEM, 2 mM Glutamine, 10 % FBS and 1 % Sodium Pyruvate (NaP). All cell cultures were kept in 5 % (v/v) CO 2 humidi ed atmosphere at 37 °C (Binder, Germany).
Morphological phenotypes of cell lines were assessed when the cell density was up to 70 % con uence using Eclipse TS100 inverted light microscope (Nikon, Japan).
Morphological phenotypes of cell lines were assessed when the cell density was up to 70 % con uence using Eclipse TS100 inverted light microscope (Nikon, Japan).
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