An overnight culture was diluted 100-fold and incubated at 37°C for 2h (in IPTG if appropriate). Then tetracycline was added and 5μl of culture was applied onto a 1.5% low-melting agarose pad (Sigma) containing final concentrations of both IPTG and tetracycline. The agarose pad was then placed onto a coverslip slide (VWR) for imaging. Images were obtained at 30°C with a Nikon Ti inverted microscope with an incubation chamber and a Hamamatsu Orca-ER camera, an automated stage (Prior), and a Lumencor SOLA fluorescent illumination system.
Sola fluorescent illumination system
Manufactured by Lumencor
The SOLA fluorescent illumination system is a high-performance, solid-state light source designed for fluorescence microscopy applications. It provides stable, uniform, and adjustable illumination across a wide range of wavelengths, enabling researchers to optimize their experiments.
Automatically generated - may contain errors
2 protocols using sola fluorescent illumination system
1
Microscopic Imaging of Bacterial Cells
2
Fluorescence Imaging of Bacterial Stress Response
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Imaging was performed with a Nikon Eclipse Ti inverted microscope equipped with an Orca R2 (Hamamatsu) camera, a 60X Plan Apo oil objective (NA 1.4, Nikon), an automated stage (Ludl), a Lumencor SOLA fluorescent illumination system, and a custom-designed temperature-controlled Plexiglas enclosure in which the temperature was maintained at approximately 37°C during imaging. Image acquisition was performed using MATLAB scripts interfacing with μManager, as previously described [29 (link)]. Semrock filter cubes for GFP (GFP1828A) and mKate2 (mCherry-B) were used to image mNeonGreen and mNeptune, respectively. mNeonGreen (used for σB reporters) was imaged with 2x2 pixel binning at approximately 20% illumination power with 200-ms exposures, and mNeptune (used for cell segmentation) was imaged with 1x1 binning at approximately 24% power with 400-ms exposures). Images were captured at 10-minute intervals.
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