Fluorescence microscopy was performed using a Nikon Eclipse Ti-E microscope with MicroManager V 1.4 and NIS-Elements Advanced Research V 5.02 software. For time-lapse experiments, cells were loaded in microfluidic plates (CellASIC ONIX2, Merck Millipore) at 30 °C [4 (link)]. Heat shock was induced with the microfluidic plate temperature controller by increasing the temperature to 42 °C for 30 min, before returning it to 30 °C, or by shifting cultures for 30 min in a shaking incubator (Kühner, LT W Lab-Therm) pre-heated at 42 °C. Images were recorded every 10 min, and quantified as described in the figure legends.
Eclipse ti e microscope
Manufactured by National Instruments
The Eclipse Ti-E microscope is a high-performance inverted research microscope designed for a wide range of applications in life science research. It features a motorized, fully automated operation and is compatible with a variety of imaging techniques, including brightfield, phase contrast, DIC, and fluorescence microscopy.
Automatically generated - may contain errors
Lab products found in correlation
Micromanager v 1, by National Instruments (2 mentions)
Cellasic onix2, by Merck Group (2 mentions)
Rat anti cldu, by Novus Biologicals (2 mentions)
Mouse anti idu, by BD (2 mentions)
Anti single strand dna, by Merck Group (2 mentions)
Bodipy 493 503, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 488 conjugated anti rabbit igg secondary antibody, by Thermo Fisher Scientific (1 mentions)
Permafluor, by Thermo Fisher Scientific (1 mentions)
5 protocols using eclipse ti e microscope
1
Time-lapse Fluorescence Microscopy of Heat Shock
2
Replicating DNA Analysis Protocol
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Analysis of replicating DNA were performed according to the methods described previously32 (link). In brief, cells were pulse-labeled with 50 μM CldU for 15 min, followed by second labeling with 50 μM IdU for 15 min before analysis. Cells were harvested and DNA fibers were stretched onto glass slides in a DNA lysis buffer (200 mM Tris-HCl at pH 7.4, 0.5% SDS, 50 mM EDTA). Single replicating DNA was detected with rat anti-CldU (Novus), mouse anti-IdU (Becton Dickinson) and anti-single-strand DNA (EMD Millipore) antibodies. Images were captured using the Nikon Eclipse TiE microscope and processed using NIS Elements AR imaging software.
3
Replicating DNA Analysis Protocol
4
Fibroblast Response to Glyoxal and AICAR
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Fibroblasts (3.8×104) were seeded onto 12 mm coverslips. After 24 hours, 0.6 mM glyoxal, 1 mM AICAR (5-aminoimidazole-4-carboxamide ribonucleotide) (Sigma #A9978), or both, were added for a further 24 hours and the cells were then fixed with 2% formaldehyde for 20 min. BODIPY 493/503 (Thermo Fisher Scientific, #D3922) and anti-perilipin-2 staining (Santa Cruz Biotechnology, #sc-377429) were performed according to the protocol of Listenberger et al.23 (link) The coverslips were then incubated for 1 hour at room temperature with Alexa Fluor 488-conjugated anti-rabbit IgG secondary antibodies (1:1000) (Thermo Fisher Scientific, #A11034). The nuclei were labeled with 2 µg/mL 4',6-diaminido-2-phenylindole (DAPI) and the coverslips were then mounted with PermaFluor (Thermo Fisher Scientific, #TA-030-FM). Images were acquired with a Nikon ECLIPSE Ti-E microscope and NIS-Element software.
5
Time-lapse Fluorescence Microscopy of Heat Shock
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Fluorescence microscopy was performed using a Nikon Eclipse Ti-E microscope with MicroManager V 1.4 and NIS-Elements Advanced Research V 5.02 software. For time-lapse experiments, cells were loaded in microfluidic plates (CellASIC ONIX2, Merck Millipore) at 30 °C [4 (link)]. Heat shock was induced with the microfluidic plate temperature controller by increasing the temperature to 42 °C for 30 min, before returning it to 30 °C, or by shifting cultures for 30 min in a shaking incubator (Kühner, LT W Lab-Therm) pre-heated at 42 °C. Images were recorded every 10 min, and quantified as described in the figure legends.
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