Cells were counted using Cellometer Auto 1000 Bright Field Cell Counter (Nexcelom) and 100.000 cells were seeded on 10 μg/ml fibronectin-coated glass bottom 35 mm. Halo-CAAX was labeled using Janelia Fluor 549 HaloTag ligand (200–400 nM; GA1110; Promega) and endogenous SNAP-Profilin-1 were labeled using SNAP-Cell 647-SiR (500 nM– 1 μM; S9102S; NEB), 30 minutes prior to imaging. Cells were imaged in phenol-red free DMEM supplemented with 20 mM HEPES pH 7.4 on a climate-controlled (maintained at 37°C), fully motorized Nikon Ti-Eclipse inverted microscope equipped with Perfect Focus System, an Andor Diskovery illuminator coupled to a Yokogawa CSU-XI confocal spinning disk head with 100 nm pinholes, and a 60x (1.49 NA) APO TIRF objective (Nikon) with an additional 1.8x tube lens, yielding a final magnification of 108x (Andor Technology). Images were recorded at 5 s or 10 s per frame using a scientific CMOS camera with 6.5-μm pixel size (pco.edge).
Csu xi confocal spinning disk head
Manufactured by Nikon
The CSU-XI confocal spinning disk head is a specialized imaging device designed for high-speed, high-resolution confocal microscopy. It features a spinning disk with multiple pinholes that allows for parallel confocal imaging, enabling rapid acquisition of images with improved signal-to-noise ratio.
Automatically generated - may contain errors
Lab products found in correlation
Janelia fluor 549 halotag ligand, by Promega (2 mentions)
Snap cell 647 sir, by New England Biolabs (2 mentions)
Apo tirf 1.49 na objective, by Nikon (2 mentions)
Cellometer auto 1000 bright field cell counter, by Revvity (2 mentions)
Eclipse ti inverted microscope, by Oxford Instruments (1 mentions)
2 protocols using csu xi confocal spinning disk head
1
Imaging Actin Dynamics in Live Cells
2
Live-cell imaging of CAAX and Profilin-1
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Cells were counted using Cellometer Auto 1000 Bright Field Cell Counter (Nexcelom) and 100.000 cells were seeded on 10 µg/ml fibronectin-coated glass bottom 35 mm. Halo-CAAX was labeled using Janelia
Fluor 549 HaloTag ligand (200 -400 nM; GA1110; Promega) and endogenous SNAP-Profilin-1 were labeled using SNAP-Cell 647-SiR (500 nM -1 µM; S9102S; NEB), 30 minutes prior to imaging. Cells were imaged in phenol-red free DMEM supplemented with 20 mM HEPES pH 7.4 on a climate-controlled (maintained at 37 oC), fully motorized Nikon Ti-Eclipse inverted microscope equipped with Perfect Focus System, an Andor Diskovery illuminator coupled to a Yokogawa CSU-XI confocal spinning disk head with 100 nm pinholes, and a 60x (1.49 NA) APO TIRF objective (Nikon) with an additional 1.8x tube lens, yielding a final magnification of 108x (Andor Technology). Images were recorded at 3 Hz frame rate using a scientific CMOS camera with 6.5-µm pixel size (pco.edge).
Fluor 549 HaloTag ligand (200 -400 nM; GA1110; Promega) and endogenous SNAP-Profilin-1 were labeled using SNAP-Cell 647-SiR (500 nM -1 µM; S9102S; NEB), 30 minutes prior to imaging. Cells were imaged in phenol-red free DMEM supplemented with 20 mM HEPES pH 7.4 on a climate-controlled (maintained at 37 oC), fully motorized Nikon Ti-Eclipse inverted microscope equipped with Perfect Focus System, an Andor Diskovery illuminator coupled to a Yokogawa CSU-XI confocal spinning disk head with 100 nm pinholes, and a 60x (1.49 NA) APO TIRF objective (Nikon) with an additional 1.8x tube lens, yielding a final magnification of 108x (Andor Technology). Images were recorded at 3 Hz frame rate using a scientific CMOS camera with 6.5-µm pixel size (pco.edge).
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