Ix83 epifluorescence microscope

Manufactured by Olympus

The IX83 epifluorescence microscope is an advanced imaging system designed for high-performance fluorescence microscopy. It features a modular and versatile design that allows for various configurations to meet diverse research needs. The IX83 provides efficient fluorescence excitation and high-quality image capture, enabling researchers to study a wide range of biological samples with precision and clarity.

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Lab products found in correlation

Orca fusion scmos camera, by Olympus (3 mentions) Cellsense software, by Olympus (2 mentions) Proteinase k, by Thermo Fisher Scientific (2 mentions) Ab91605, by Abcam (1 mentions) Hydroxyurea, by Thermo Fisher Scientific (1 mentions) Rnase a, by Thermo Fisher Scientific (1 mentions) Trizol, by Thermo Fisher Scientific (1 mentions) High capacity cdna reverse transcription kit, by Thermo Fisher Scientific (1 mentions) Gotaq green master mix, by Promega (1 mentions) Propidium iodide, by Thermo Fisher Scientific (1 mentions) Calcein am, by Thermo Fisher Scientific (1 mentions)

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IX83 microscope (347 citations) Epifluorescence microscope (122 citations) IX83 inverted epifluorescence microscope (10 citations)

7 protocols using ix83 epifluorescence microscope

Studying NFAT5 Protein Condensation

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Recombinant NFAT5 CTD and PLD fused to sfGFP fusion proteins at varying concentrations were mixed with 5% Dextran (a molecular crowding agent) in droplet buffer (20 mM Na-HEPES pH7.4, 1 mM TCEP) and NaCl (or other salts) were added from a concentrated stock to achieve the indicated osmolarity. The protein solution was incubated for 10 min at room temperature and loaded onto a glass slide with a coverslip. Slides were then imaged with a ×10 dry objective (NA 0.3) on an Olympus IX83 epifluorescence microscope equipped with an Orca Fusion scMOS camera. Droplets containing sfGFP and mCherry were imaged using the FITC and TRITC channels, respectively. Droplets without GFP were visualized using the brightfield channel. Unless indicated, images presented are of droplets settled on the glass coverslip. All assays were performed at room temperature.
To pellet condensates formed by NFAT5 GFP-CTD (Figure 5G) droplet reactions (containing 5% Dextran, 70 μM GFP-CTD, 1000 mOsm/L NaCl in droplet buffet) were centrifuged at 20,000×g for 10 min. Supernatant was removed and condensate pellets were resuspended droplet buffer (20 mM Na-HEPES pH7.4, 1 mM TCEP) with 5% dextran (but no added NaCl) to determine if the condensates were reversible.

Khandwala C.B., Sarkar P., Schmidt H.B., Ma M., Kinnebrew M., Pusapati G.V., Patel B.B., Tillo D., Lebensohn A.M, & Rohatgi R. (2023). Direct ionic stress sensing and mitigation by the transcription factor NFAT5. bioRxiv.

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Cryosectioning Fibrin-Encapsulated Cardiac Cells

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At the end of the experiment the fibrin encapsulated cells are removed from the BCTM and fixed using fresh 4% paraformaldehyde at room temperature for 1 hour before moving to 4 °C. Samples are then placed in OCT solution and frozen for cryosectioning of 10 micron thick slices. Sections were immunolabeled with rabbit anti-cardiac troponin T (Abcam, ab91605) and mouse monoclonal anti-actin (alpha-sarcomeric) antibody (Sigma, A2172). Nuclei are counter stained with DAPI. Images were taken using an Olympus IX83 epifluorescence microscope.

Rogers A.J., Kannappan R., Abukhalifeh H., Ghazal M., Miller J.M., El-Baz A., Fast V.G, & Sethu P. (2019). Hemodynamic Stimulation Using the Biomimetic Cardiac Tissue Model (BCTM) Enhances Maturation of Human Induced Pluripotent Stem Cell Derived Cardiomyocytes. Cells, tissues, organs, 206(1-2), 82-94.

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High-Resolution Microscopy of Knock-In Cells

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Unless otherwise specified, HCT116 and U2OS knock-in cells were seeded at 15,000 cells/well in 18-well ibidi glass-bottom μ-slides, allowed to adhere o/n, treated as indicated and imaged with an Olympus IX83 epifluorescence microscope equipped with an Orca Fusion scMOS camera and a 100× oil objective (NA 1.45). At least 15 z planes in 0.26 μm steps were taken per position and channel. Images were deconvoluted with the Olympus cellSense software using the proprietary constrained iterative deblurring algorithm.

Broder Schmidt H., Jaafar Z.A., Erik Wulff B., Rodencal J.J., Hong K., Aziz-Zanjani M.O., Jackson P.K., Leonetti M.D., Dixon S.J., Rohatgi R, & Brandman O. (2022). Oxaliplatin disrupts nucleolar function through biophysical disintegration. Cell reports, 41(6), 111629.

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Cell Cycle Synchronization and Imaging Protocol

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Yeast cultures were grown to an OD600 of 0.4–0.6 at 30°C and then pretreated with 100 mM hydroxyurea (Alfa Aesar) for 2 h at 30°C. After 2 h of pre-treatment with hydroxyurea (HU), a saturating concentration of α-factor (10 μM) was added, and cultures were then fixed after 240 min using an overnight ethanol fixation at −20°C. After ethanol fixation, yeast were resuspended and washed twice in 50 mM sodium citrate buffer (pH 7.2). Next, the cultures were incubated with 20 mg/ml RNase A (Thermo Fisher Scientific) for a minimum of 1 h at 37°C. After RNase incubation, proteinase K (Thermo Fisher Scientific) was added to the cultures at a final concentration of 0.4 mg/ml and incubated at 55°C for a minimum of 1 h then placed at 4°C overnight. For imaging, cells were pelleted, then washed and mounted in 1× PBS (pH 7.4). Cells were then imaged on the IX83 epifluorescence microscope (Olympus).

Simke W.C., Johnson C.P., Hart A.J., Mayhue S., Craig P.L., Sojka S, & Kelley J.B. (2022). Phosphorylation of RGS regulates MAP kinase localization and promotes completion of cytokinesis. Life Science Alliance, 5(10), e202101245.

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High-Resolution Microscopy of Knock-In Cells

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Immunofluorescence and RT-PCR Analysis of iPSCs

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iPSCs at p6 were fixed with 4% PFA for 10 min and washed with DPBS. Fixed cells were incubated in primary antibodies overnight at 4 °C and in secondary antibodies for 30 min at room temperature. Nuclear staining was done with DAPI. Images were acquired using an Olympus IX83 epifluorescence microscope. Random areas on the DAPI images were selected for cell counting. Cells positive for pluripotency markers (Nanog or Oct3/4) were manually counted using ImageJ Software and reported as the percentage of total cells (DAPI).
RNA was isolated from cells at p10 using TRIzol (Invitrogen) following the manufacturer’s protocol. RT-PCR for pluripotency markers was performed by generating cDNA according to the manufacturer’s protocol (High-Capacity cDNA Reverse Transcription Kit, Applied Biosystems). PCR was performed using GoTaq Green Master Mix (Promega) and samples were run on 1% agarose gel. The PCR cycle was: 5 min 95 °C, 38 × (15 s 95 °C, 30 s 55 °C, 45 s 72 °C), 5 min 72 °C, ∞ 4 °C.

Madencioglu D.A., Kruth K., Shin M., Andreasen N., Wassink T, & Williams A. (2021). Generation of a human induced pluripotent stem cell line from a patient diagnosed with schizophrenia carrying a 16p11.2 deletion. Stem cell research, 59, 102636.

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Cytocompatibility Evaluation of Functionalised AZ31 Substrates

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The murine osteoblast cell line (MC3T3-E1) (ATCC® CRL-2593™, LGC standard) was used for cytocompatibility studies. Cells were cultured under 37 °C, 5% CO 2 and 95% relative humidity in DMEM/F-12 containing 10% FBS and 1% penicillin and streptomycin. Cells were seeded (10 4 cells.cm -2 ) on HA functionalised AZ31-MT-A coated and uncoated AZ31 substrates and maintained in complete DMEM/F12 (1 cm 2 = 1.25 ml) in accordance to ISO 10993-12 [17] (link) for 1 and 3 days before the live-dead staining was performed. Calcein AM (eBioscience 65-0853) and propidium iodide (eBioscience 00-6990) were dissolved in DMSO (1 mg/ml) and used at 1:500 ratio in complete DMEM/F12. Cells cultured on the experimental substrates (10 × 10 mm) were washed with PBS and incubated with fluorescent dyes for 20 min at 37 °C. After incubation cells were rinsed with PBS and imaged using an Olympus IX83 epifluorescence microscope fitted with a 10X objective.

Agarwal S., Labour M.N., Hoey D., Duffy B., Curtin J, & Jaiswal S. (2018). Enhanced corrosion resistance and cytocompatibility of biomimetic hyaluronic acid functionalised silane coating on AZ31 Mg alloy for orthopaedic applications. Journal of materials science. Materials in medicine, 29(9).

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