On each imaging day (timing depending on experiment), GLO-Bot loaded each rhizotron into the imager then closed the door and triggered μManager to capture 5 min exposures on each side of the rhizotron. Inside the imaging system, there are two cameras on top of one another, with one taking an image of the top part of the rhizotron and the other, the bottom part of the rhizotron. After one side was imaged, the rhizotrons were rotated using a Lambda 10-3 Optical Filter Changer (Sutter Instrument, Novato, CA) and the other side of the rhizotron was imaged using the top and bottom camera, yielding four images per rhizotron. If it was the first imaging day or a designated luciferin day (every 6 days), GLO-Bot added 50 mL of 300 μM D-luciferin (Biosynth International Inc, Itasca, IL) to the top of each rhizotron immediately before loading the rhizotron into the imager. Before every replicate, a blank image was taken to account for background noise within the imager.
Lambda 10 3 optical filter changer
Manufactured by Sutter Instruments
The Lambda 10-3 Optical Filter Changer is a device designed to quickly and accurately change optical filters. It is a compact, motor-driven mechanism that can hold up to 10 filters and rapidly switch between them under computer control.
Automatically generated - may contain errors
Lab products found in correlation
Zen acquisition software, by Zeiss (1 mentions)
X cite 120led boost high power led illumination system, by Excelitas (1 mentions)
Orca flash4.0 v3 digital cmos camera, by Hamamatsu Photonics (1 mentions)
Ix81 motorized inverted microscope, by Olympus (1 mentions)
D luciferin, by Biosynth (1 mentions)
3 protocols using lambda 10 3 optical filter changer
1
Rhizotron Imaging Protocol for Root Analysis
2
Multicolor Fluorescence Microscopy Imaging
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Epifluorescent images were acquired with an Olympus ix-81 motorized inverted microscope with CellSens acquisition software v1.0 (Olympus, Center Valley, PA), with a manual Olympus stage and software-based z-step, using an X-Cite 120LED Boost High-Power LED illumination System (Excelitas Technologies, Waltham, MA). Excitation/emission wavelengths were controlled using Lambda 10-3 Optical Filter Changer (Sutter Instrument, Novato, CA) equipped with the following filter excitation wheels located at the light source: AT350/50x, ET402/15x, ET490/20x, ET555/25x, ET645/30x (Chroma, Bellows Falls, VT). The emission wheel located on the camera port contained the following: ET455/50m, ET525/36m, ET605/52m, ET705/72m (Chroma). Between both wheels contained the 89100bs dichroic (Chroma). An Olympus UPlanFL N 60x/1.25 Oil Iris objective lens was used for acquisition. Detector used was an ORCA-Flash4.0 V3 Digital CMOS camera (Hamamatsu, Hamamatsu City, Japan); 2 × 2 binning, 6.5µmx6.5µm pixel size, 2048 × 2048 chip size, with rolling shutter. Confocal images were captured on an LSM 880 with Airyscan with ZEN acquisition software (Zeiss, Jena, Germany). z-stacks were acquired and processed using the Airyscan Processing algorithm included in ZEN.
3
Automated Rhizotron Imaging System
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20 days after sowing, the automated rhizotron handling system (designed by Modular Science, San Francisco) added 50 ml of 300 µM D-luciferin (Biosynth) at the top of each rhizotron and loaded the rhizotron into a fixed stage that was controlled by a Lambda 10-3 optical filter changer (Sutter Instruments, Novato, CA) in the GLO1 imaging system15 . 5-min exposures were taken per rhizotron side.
A shoot image was taken right after the four root images using an ids UI-359xLE-C camera with a Fujinon C-Mount 8–80 mm Varifocal lens that was installed in GLO1. Three LED strips on each side of the camera were switched on before a shoot image was taken.
A shoot image was taken right after the four root images using an ids UI-359xLE-C camera with a Fujinon C-Mount 8–80 mm Varifocal lens that was installed in GLO1. Three LED strips on each side of the camera were switched on before a shoot image was taken.
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