L6 cells transfected with FoxO1 biosensor were seeded onto a 96-well plate for treatment. When cells reached 50–60% confluency, they were treated with conditioned media collected from THP-1 cells that were stimulated with LPS in the presence of control or copper leachates for 24 hours. After incubation time, all cells were starved in 0% FBS AMEM for 90 min, followed by treatment with 100 nM insulin. Images were taken using EVOS FL Auto 2 over a span of 30 min to observe fluorescent translocation. Nuclear fluorescence signal was traced from the nucleus to the cytosol in all treatments with number of cells being 10 for each treatment. Data quantitation was performed using Celleste software from Thermo Scientific.
Celleste software
Manufactured by Thermo Fisher Scientific
Sourced in United States
Celleste is a software solution developed by Thermo Fisher Scientific for managing and analyzing data from cell culture experiments. The software provides tools for organizing experimental data, monitoring cell growth, and generating visualizations of experimental results.
Automatically generated - may contain errors
4 protocols using celleste software
1
FoxO1 Biosensor Translocation Assay
2
Live-cell Imaging of Mitotic Dynamics
3
Imaging HEK293 and Caco2 Cell Dynamics
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HEK293 cells were transfected with plasmids encoding FILIP1L-eGFP and mCherry-PFDN1, and incubated for 24 h. Cells were then placed in the EVOS Onstage Incubator at 5% CO2, 20% O2 and 80% humidity. Fluorescent images were acquired at 20-minute intervals. Caco2 clones were incubated with SPY-595 DNA and SPY-650-tubulin fluorescent dyes (Cytoskeleton) for 1 h and placed in the EVOS Onstage Incubator. Forty random fields were selected, and fluorescent and phase contrast images were acquired at 5-minute intervals. Images acquired over the initial 4 h were used to quantify data. Durations longer than 4 h demonstrated substantial fluorescent signal bleaching. Images were acquired by an EVOS FL Auto 2 microscope (Thermo) at 20X objective magnification (z-stack of 1.7 μmol/L thickness). Mitotic length was quantified as nuclear envelope breakdown (NEBD) to Anaphase. Time to cytokinesis completion was quantified as NEBD to membrane fission by phase contrast. Acquired images were analyzed and quantified using Celleste software (Thermo, Version 4.1.1). Detailed quantification procedures were written in Supplementary Information.
4
Quantifying EGFP-c-Fos Cell Death
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For measuring cell death, EGFP‐c‐Fos transfected cells were fixed by 4% paraformaldehyde after drug treatment, and permeabilized by 0.1% Tx‐100 in PBS. After incubated with TUNEL reaction mixture (TMR Red; Roche, Germany) for 1 hr at 37°C in dark, cells were imaged by fluorescent microscopy (EVOS FL Auto2; Thermo Fisher Scientific, USA). Images were processed and analyzed by Celleste software (Thermo Fisher Scientific, USA).
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