20-µm coronal DCX-ir sections DAPI labeled were scanned on a confocal microscope Nikon Eclipse Ti2 (Nikon, Florence, Italy) equipped with an x-y-z motorized stage, a digital camera DS-Qi2 (Nikon, Florence, Italy), and the acquisition and image analysis software NIS-Elements C (Nikon, Florence, Italy). Each section was used to identify (1) the inner molecular layer (IML), which was defined as the first 50 µm from and parallel to the outer border of DAPI-positive cells, and (2) the medial molecular layer (MML), which was defined as the last 100 µm from, and parallel to, the outer border of the IML. DCX-positive dendrites within the IML and MML were imaged and level of branching was evaluated from a maximum-intensity projection of the z-series stack as described by Rosenzweig and Wojtowicz89 (link) and expressed as a percentage of DCX-ir area/µm2 of GCL, IML, or MML including also “orphan” dendrites otherwise excluded by conventional single tracing method of dendrites. The same procedure was adopted for the quantification OxA immunoreactivity in the molecular layers.
Nis elements c
Manufactured by Nikon
Sourced in United Kingdom
NIS-Elements C is a comprehensive software platform designed for microscope imaging and analysis. It provides a unified interface for controlling various Nikon microscope systems and accessories, enabling seamless integration and efficient workflow. The software offers a range of core functionalities for image acquisition, processing, and analysis to support scientific research and investigation.
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Lab products found in correlation
Ds qi2, by Nikon (1 mentions)
Eclipse ti2, by Nikon (1 mentions)
A1r confocal, by Nikon (1 mentions)
Eclipse ti inverted microscope, by Nikon (1 mentions)
Cfi plan apo vc objective, by Nikon (1 mentions)
A1r laser scanning confocal microscope, by Nikon (1 mentions)
Live dead baclight stain, by Thermo Fisher Scientific (1 mentions)
Gene frame, by Thermo Fisher Scientific (1 mentions)
Volocity software, by PerkinElmer (1 mentions)
Polysine slide, by Thermo Fisher Scientific (1 mentions)
Lysotracker green dnd 26, by Thermo Fisher Scientific (1 mentions)
Fitc conjugated goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Eclipse ti e inverted microscope, by Nikon (1 mentions)
A1r system, by Nikon (1 mentions)
Anti human gitrl antibody, by R&D Systems (1 mentions)
Eight well culture slides, by Corning (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Cell Signaling Technology (1 mentions)
8 protocols using nis elements c
1
Quantifying DCX and OxA in Hippocampus
2
Confocal Imaging of Retinal Flat Mounts
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Confocal images spanning entire retinal flat mounts were generated as previously described (Bosco et al., 2012 (link), 2011 (link)) using a confocal imaging system equipped with a 20× lens and a resonant scanner (A1R confocal, Eclipse Ti inverted microscope and NIS-Elements C, Nikon). Multipoint images (625 xy positions) were acquired at high resolution (0.41 µm/px), then stitched and projected as maximal intensity of the inner 30-40 µm of retina (0.8-µm step). To allow image analysis and quantification, the parameters of image acquisition were maintained constant between retinal samples, and, for illustration, images had their brightness and contrast minimally edited. Counts of ONH cells expressing GFP and/or sialoadhesin where manually performed in the central 200×200 µm around the optic disc, visualizing each channel independently in maximum-intensity projections, and verifying colocalization in slice view, through the z plane.
3
Quantifying Presynaptic Protein Colocalization
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L4 larvae were mounted on a 1.5% agarose pad with 20 mM sodium azide in M9 solution for anesthesia. Images were acquired on an inverted confocal microscope (Nikon A1, Nikon) with a 60× objective lens, and were analyzed by NIS-Elements C/NIS-Elements C-ER and ImageJ software, respectively. The Z-stack image was acquired from the whole animals expressing each fluorescent fusion proteins. Co-localization between GLR-1 or GLC-3 GFP fusion proteins and a presynaptic mCherry::RAB-3 fusion protein was quantified by counting the number of GFP pixels overlapping with mCherry signal in a single z-axis flame. The number of pixels in each frame of Z-scan was averaged in each animal and used for quantitative analyses.
4
Biofilm Visualization via CLSM Imaging
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For visualisation of static biofilms with confocal laser scanning microscopy (CLSM), stained coverslips were rinsed with PBS and inverted onto a PBS-filled rubber frame secured on a microscope slide. Imaging was performed using a Nikon A1R confocal laser scanning microscope fitted with CFI PLAN APO VC objective (Nikon 60x/1.40 Oil). Images were captured with NIS-Elements C (v4.4, Nikon) software and processed using Imaris (v8.2, Bitplane) software.
5
Chitin Characterization by UV-Vis and Confocal
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UV-Vis spectra of chitin samples were collected between 250 and 650 nm with a 1 nm resolution, and an average time of 0.1 s.
Confocal microscopy imaging was performed using an NIS-Elements C, Nikon, and exciting the sample using a 489.3 nm laser. The 512 × 512 images were acquired at a pixel size of 0.62 μm (optical resolution of 0.25 μm) and a Z step of 0.35 μm.
Confocal microscopy imaging was performed using an NIS-Elements C, Nikon, and exciting the sample using a 489.3 nm laser. The 512 × 512 images were acquired at a pixel size of 0.62 μm (optical resolution of 0.25 μm) and a Z step of 0.35 μm.
6
Quantifying Biofilm Biovolume by Confocal Microscopy
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Biofilms for visualization experiments were cultured on sterile glass coverslips incubated in wells of six-well tissue culture dishes. Following growth, biofilms were rinsed with PBS and incubated with Live/Dead BacLight stain (Molecular Probes, Eugene, OR) for 15 min at 20°C. For confocal laser scanning microscopy, each stained coverslip was rinsed with PBS and inverted onto a PBS-filled Gene frame (25 μL, 1.0 × 1.0 cm, Thermo Fisher Scientific, Waltham, MA) secured on a microscope slide.
Imaging was performed using a Nikon A1R confocal laser scanning microscope fitted with CFI PLAN APO VC objective (Nikon 60×/1.40 Oil).
Images were captured with NIS-ElEmEnts C (v4.4, Nikon, Kingston upon Thames, UK) software and processed using ImarIs (v8.2, Bitplane, Zurich, Switzerland) software. Biovolume quantification of Z-stacks was conducted using VolocIty software (v6.3, PerkinElmer, Waltham, MA), set to identify objects ≥1 μm 2 as S. gordonii cells. At least three Z-stacks (image size 1024 × 1024) from three different fields of view were analyzed for each strain. The data were analyzed from three independent experiments. Statistical significance of differences between biofilm biovolume was assessed using analysis of variance with Tukey's post-hoc test.
Imaging was performed using a Nikon A1R confocal laser scanning microscope fitted with CFI PLAN APO VC objective (Nikon 60×/1.40 Oil).
Images were captured with NIS-ElEmEnts C (v4.4, Nikon, Kingston upon Thames, UK) software and processed using ImarIs (v8.2, Bitplane, Zurich, Switzerland) software. Biovolume quantification of Z-stacks was conducted using VolocIty software (v6.3, PerkinElmer, Waltham, MA), set to identify objects ≥1 μm 2 as S. gordonii cells. At least three Z-stacks (image size 1024 × 1024) from three different fields of view were analyzed for each strain. The data were analyzed from three independent experiments. Statistical significance of differences between biofilm biovolume was assessed using analysis of variance with Tukey's post-hoc test.
7
Visualizing RNA-DNA Hybrids in Cells
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Day 8 FLDCs from C57BL/6 mice were transfected with 1 μg/ml fluorescently labelled R:D45 or the fluorescently labelled non-complementary oligonucleotides ssRNA45 and ssDNA60 using Lipofectamine LTX. After 1 h, cells were adhered to polysine slides (Thermo Scientific), fixed with 4% PFA and nuclei stained with DAPI. To stain endolysosomal compartments, 2 μM LysoTracker Green DND-26 (Life Technologies) was added to the cells for 1 h before fixation. To visualise RNA:DNA hybrids using the S9.6 antibody, fixed cells were permeabilised using 0.2% (v/v) Triton X-100/PBS for 2 min, blocked for 1 h with 2% (w/v) BSA/PBS and incubated with 1 μg/ml S9.6 antibody, which was detected using 1:500 dilution FITC-conjugated goat anti-mouse IgG (Molecular Probes). Images were taken on a Nikon A1R confocal microscope comprising of a Nikon Eclipse TiE inverted microscope with Perfect Focus System. Image capture was performed using Nikon Nis-Elements C (Nikon Instruments Europe). 3 channel images were acquired by consecutive scanning with only one laser line active per scan.
8
Quantification of GITRL and GITR Expression
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CRL5946 cells were seeded on eight-well culture slides (Corning Life Sciences) with each well 104 cells and treated with cisplatin 2 ug/ml or γ-ray radiation 7.5 Gy. After incubating for 96 h, the medium was removed and the slide was washed twice with cold PBS. The viable cells were fixed with 4% paraformaldehyde for 10 min. The slides were stained with anti-human GITRL antibody (Clone:109101, R&D SYSTEMS), anti-mouse IgG Antibody-AF488 conjugated (Invitrogen), anti-human GITR antibody-PE conjugated (Clone#621, Biolegend Inc.), and DAPI (Cell Signaling) following the commercial instructions. The fluorescence images of whole sides were captured by Nikon A1R+ system, which is built onto a Nikon Eclipse TI-E inverted microscope with Nikon NIS Elements C for acquisition and analysis of images.
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