Cells were fixed with 3.7% paraformaldehyde in PBS for 10 min at room temperature and then permeabilized in 0.2% Triton X-100 in PBS for 10 min at room temperature. After blocking with 1% BSA in PBS for 1 h at room temperature, the cells were stained with an anti-IRF3 antibody (1:500, #NBP2-67741, Novus Biologicals) in PBS overnight at 4 °C and further stained with an Alexa Fluor 488-conjugated goat anti-rabbit antibody (1:1000, Invitrogen) in PBS for 1 h at room temperature. After staining the cells with DAPI for 5 min at room temperature to visualize nuclei, imaging was performed on an ECLIPSE Ts2R fluorescence microscope (Nikon).
Eclipse ts2r fluorescence microscope
Manufactured by Nikon
Sourced in Japan
The ECLIPSE Ts2R is a fluorescence microscope designed for advanced research applications. It features high-quality optics and illumination systems to enable clear and detailed imaging of fluorescently labeled samples.
Automatically generated - may contain errors
Lab products found in correlation
Cross beam 340 scanning electron microscope, by Zeiss (2 mentions)
Zetasizer nano series, by Malvern Panalytical (2 mentions)
Infinite 200 pro spectrophotometer, by Tecan (2 mentions)
Alexa fluor 488 conjugated goat anti rabbit antibody, by Thermo Fisher Scientific (1 mentions)
Inkredible 3d bioprinter, by Cellink (1 mentions)
1 2 ld plus freeze dryer, by Martin Christ (1 mentions)
Mcr 302 rheometer, by Anton Paar (1 mentions)
Prism 9, by GraphPad (1 mentions)
Nis element, by Nikon (1 mentions)
5 protocols using eclipse ts2r fluorescence microscope
1
Immunofluorescence Staining of IRF3
2
Comprehensive Nanocomposite Hydrogel Characterization
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Scanning electron microscopy (SEM) was performed on a Zeiss cross beam 340 scanning electron microscope, to determine the morphology of the particles. The average size of the particles from SEM images was measured by ImageJ from 35 NPs. An InlensDuo detector (secondary electron (SE) detector) for particles and a SESI detector (SE, and secondary ion detector) for the hydrogels were used. Zeta potential measurements and dynamic light scattering (DLS) were conducted using a Malvern Zetasizer Nano Series. A Nikon ECLIPSE Ts2R fluorescence microscope was used to determine the cell morphology and fluorescence imaging. Cell viability was measured using a Tecan Infinite 200 PRO spectrophotometer. A Christ Alpha 1-2-LD plus freeze-dryer was used to produce porous hydrogel scaffolds. An INKREDIBLE 3D bioprinter (CELLINK, Boston, MA, USA) was used to print all nanocomposite hydrogels into computer-designed 3D structures. Rheological measurements were carried out using an MCR 302 rheometer (Anton Paar, Ashland, VA, USA) with a 25 mm diameter parallel-plate geometry measuring system. A material testing machine (type 066590, ZwickRoell GmbH & Co. KG, Ulm, Germany) was used to determine the compression modulus of the samples.
3
Characterization of Nanoparticle Morphology
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Scanning electron microscopy (SEM) was performed on a Zeiss cross beam 340 scanning electron microscope to determine the morphology of the particles. The average size of NPs from the SEM image was measured by ImageJ software. Zeta potential measurements and dynamic light scattering (DLS) were conducted using a Malvern Zetasizer Nano Series. A Nikon ECLIPSE Ts2R fluorescence microscope was used to determine the cell morphology and fluorescence imaging. Cell viability was measured using a Tecan Infinite 200 PRO spectrophotometer.
4
Quantifying Tumor-Stromal Interactions
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All co-cultures were monitored daily using an ECLIPSE Ts2R fluorescence microscope (Nikon, Tokyo, Japan). Daily ZsG/tdT fluorescence images of the spheroid suspension co-cultures were captured by NIS-Elements (Nikon). The ZsG+ tumor cell area in their co-cultures was measured by ImageJ, normalized to Day 0 and plotted using GraphPad Prism 9.4. ZsG/tdT fluorescence images of the 2D co-culture of HC:tumor and HC+HSC:tumor were captured by an ECLIPSE Ts2R fluorescence microscope on Day 1, 4 and 7. Day 1 and Day 7 images were used to count and determine the percentage of partially ‘eaten’ and intact tdT+ spheroids. Specifically, the tdT fluorescence area of individual normal spheroids on Day 1 to Day 7 was measured using ImageJ, and partially ‘eaten’ and intact spheroids were defined as those with more than 50% and less than 10% reduction in their tdT+ area, respectively. ZsG/tdT fluorescence images of the 2D co-culture of HC:tumor and HC+HSC:tumor were captured every 7 days by a FX Automated Microscope (Lionheart, Winooski, VT, USA) until Day 21. tdT+ HC area within the original seeding region was measured in ImageJ, normalized to Day 1 and plotted using GraphPad Prism 9.4. All measurements were taken from three independent co-cultures.
5
Peptide-Dye Interaction and Salt-Triggered Disruption
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Ethanolic solution of the peptide
(0.02 mM) was incubated with 50 μM of carboxyfluorescein for
24 h. Then, one drop (10 μL) of this solution was drop-casted
on a cover slide and dried at room temperature under vacuum. The fluorescence
microscopic image was taken with a Nikon eclipse Ts2R fluorescence
microscope. For salt-triggered disruption studies with the spheres,
KCl solution (1 mM 5 μL) was added to the dye solution of the
peptide and incubated for 12 h, and the image was taken subsequently.
(0.02 mM) was incubated with 50 μM of carboxyfluorescein for
24 h. Then, one drop (10 μL) of this solution was drop-casted
on a cover slide and dried at room temperature under vacuum. The fluorescence
microscopic image was taken with a Nikon eclipse Ts2R fluorescence
microscope. For salt-triggered disruption studies with the spheres,
KCl solution (1 mM 5 μL) was added to the dye solution of the
peptide and incubated for 12 h, and the image was taken subsequently.
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