Cells were seeded into 8-well μ-slides (ibidi GmbH, Germany) with 2 × 104 cells/well and left to recover for 24 h. For organelle tracking, the medium was replaced with serum- and phenol red-free medium with 50 nM MitoTracker Red CMXRos, MitoTracker Green FM or LysoTracker Red (Life technologies, Vienna, Austria). For calcium imaging, cells were incubated with 0.5 µM Rhod-2 AM (Abcam, Cambridge, UK) in serum- and phenol red-free medium for 30 min at 4 °C. After 1 h, cells were washed and imaged with the Nikon Eclipse Ti-e fluorescence microscope with differential interference contrast and RFP or GFP filter settings and a sCMOS pco.edge camera. Life-cell imaging was performed in an environmental chamber pre-heated to 37 °C with 5% CO2. For non-fluorescence imaging, phase-contrast pictures were taken with the Nikon Eclipse Ti inverted microscope with a Nikon DS-Fi1c camera. Contrast and brightness were adjusted with ImageJ. Cell area was calculated as mean occupied area per cell from at least two different sections in one well at the end of life-cell imaging (48 h) using ImageJ and then normalized to control.
Ds fi1c camera
Manufactured by Nikon
Sourced in Japan
The DS-Fi1c is a digital microscope camera from Nikon. It features a 5-megapixel CMOS sensor and can capture images at resolutions up to 2560 x 1920 pixels. The camera is designed for integration with Nikon's microscope systems and software.
Automatically generated - may contain errors
Lab products found in correlation
Nis elements software, by Nikon (4 mentions)
Sc 6026, by Santa Cruz Biotechnology (3 mentions)
Eclipse te300, by Nikon (3 mentions)
A 16001, by Thermo Fisher Scientific (2 mentions)
Evans blue dye, by Merck Group (2 mentions)
Nis element d software, by Nikon (2 mentions)
Eclipse lv100nd, by Nikon (2 mentions)
Te200 microscope, by Nikon (2 mentions)
Ds u2 pc control unit, by Nikon (2 mentions)
Eclipse ni e, by Nikon (2 mentions)
Isolectin b4, by Vector Laboratories (2 mentions)
Lysotracker red, by Thermo Fisher Scientific (1 mentions)
μ slide 8 well, by Ibidi (1 mentions)
Mitotracker red cmxros, by Thermo Fisher Scientific (1 mentions)
Mitotracker green fm, by Thermo Fisher Scientific (1 mentions)
Eclipse ti e fluorescence microscope, by Nikon (1 mentions)
Eclipse ti inverted microscope, by Nikon (1 mentions)
Rhod 2 am, by Abcam (1 mentions)
Ni u microscope, by Nikon (1 mentions)
Euparal, by Carl Roth (1 mentions)
30 protocols using ds fi1c camera
1
Live-cell Imaging of Organelles and Calcium
2
Quantifying Cell Morphometry in N. benthamiana
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Stem segments of N. benthamiana were fixed in 4% (v/v) paraformaldehyde in PEM buffer (100 mM PIPES, 10 mM MgSO4, 10 mM EGTA, pH 6.9) at room temperature for 2 h. After washing in demineralized water, stem segments were glued to the vibratome stage using superglue (Roticoll, Carl Roth). Sections of 60 µm thickness, prepared with a vibrating microtome (HM 650V, ThermoScientific, Germany), were stained with 0.5% (w/v) astra blue, 0.5% (w/v) chrysoidine, and 0.5% (w/v) acridine red, and mounted in Euparal (Roth) after dehydration in isopropyl alcohol. Slides were imaged with a Nikon Ni-U microscope equipped with a Nikon DS-Fi1c camera. Images were subsequently processed to quantify cell number, shape, and size, using the method described previously (Andriankaja et al., 2012 (link)). Cellular analysis was performed on three regions from two independent sections obtained from three representative plants per treatment.
3
Automated Microscopic Analysis Protocol
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Microscopic analysis was performed using an automated Nikon Eclipse Ni-E microscope (Nikon, Tokyo, Japan) equipped with a Nikon DSFi1c camera (Nikon, Tokyo, Japan) in NIS-Elements Advanced Research 4.00.00 software (Nikon, Tokyo, Japan). Microphotographs were taken at 400× magnification.
4
Histopathological Analysis of Ovarian Tissues
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For histopathological analysis, ovarian tissues were fixed in 10% neutral buffered formaldehyde and embedded in paraffin. Cross-sections (5 μm) were obtained and stained with haematoxylin&eosin (H&E). Ovarian damage was evaluated by considering stromal congestion, oedema and infiltration, follicular degeneration and oedema and necrosis in the corpus luteum. The severity of these parameters was calculated as 0: absent, 1: minimal, 2: moderate and 3: severe (17 (link)). Sections were examined under light microscopy (Nikon Eclipse i5 light microscope with a Nikon DS-Fi1c camera and the Nikon NIS Elements version 4.0 image analysis systems. Nikon Instruments Inc., Tokyo, Japan). All tissue samples were evaluated by a single histologist who was blinded to the groups.
5
Qualitative Assessment of Blood-Retinal Barrier Leakage
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Blood-retinal barrier leakage was evaluated qualitatively using Evans blue dye. The use of this dye in the evaluation of BRB leakage has been observed to pose several problems, 39 although these problems are particularly relevant only for quantitative evaluation of BRB leakage. 40 After the mice were deeply anesthetized with avertin, Evans blue dye, dissolved in normal saline (30 mgÁmL À1 ), was injected into the left ventricle and allowed to circulate through the body for 5 minutes. After the enucleated eyes were fixed in 4% paraformaldehyde, the retinas were flat mounted and examined with a fluorescence microscope (Eclipse E800; Nikon Corp.), and images were acquired (DS-Fi1c camera; Nikon Corp.). For each experimental condition, six retinas from six different mice were used.
6
Imaging Polar Capsule Discharge Dynamics
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The polar capsule discharge process was recorded using an Andor Neo sCMOS black and white camera attached to a Nikon TI inverted epi-fluorescent microscope at an average rate of 200 frames per second, and resolution of 0.9 μm/pixel. Movies were then analyzed using Nikon’s image analysis program NIS-Elements version 3.22.15. The size of the polar capsule and the length of the elongating tubule were measured, and relative dye load was obtained from image pixel intensity throughout the discharge process. Color video clips were recorded using a Nikon DS-Fi1c camera (average 12 frames per second, and resolution 0.4 μm/pixel), mounted on a Nikon Eclipse 90i confocal microscope.
7
Retinal Vessel Visualization and Quantification
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To visualize retinal vessels, 6 retinas for each age group were incubated with FITC-conjugated isolectin B4 (1:200; Vector Laboratories, Burlingame, CA, USA) diluted in PBS containing 5% BSA and 2% Triton X-100. After 48 h incubation at 4 °C, retinas were rinsed in PBS and flat-mounted on polarized glass slides with the RGC layer facing up. Images of retinal vessels were obtained following scanning acquisition with a 10x Plan-Apochromat objective using an epifluorescence microscope (Ni-E; Nikon-Europe, Amstelveen, The Netherlands) equipped with a digital camera (DS-Fi1c camera, Nikon-Europe) and subsequent automatic digital reconstruction with NIS-Elements software (Nikon-Europe, Amstelveen, The Netherlands). Angio Tool software was used to quantify vessel density as percent of the area occupied by vessels and lacunarity as the size distribution of gaps or lacunae between vessels [28 (link)]. Values of vessel density and lacunarity were normalized to those measured at 2 months.
8
Retinal Ganglion Cell Labeling
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Mice under avertin anesthesia (n = 5 for each group) were euthanized before removing their eyes. After isolation, retinas were fixed for 90 min at 4 °C in 4% paraformaldehyde in 0.1 M phosphate buffer (PB). Fixed retinas were then stored at 4 °C in 25% sucrose in 0.1 M PB. Retinal ganglion cells were labeled using an antibody against Brn3a (1:100 in PB containing 5% BSA and 2% TritonX-100; sc-6026; Santa Cruz Biotechnology, Santa Cruz, CA, USA) incubating the retinas for 24 h at 4 °C. Retinas were then washed with PB and incubated overnight at 4 °C in an AlexaFluor488-conjugated secondary antibody (1:100; A-16001; Molecular Probes, Eugene, OR, USA). Finally, retinas were washed with PB, mounted on glass slides with the photoreceptor side facing down and viewed with a fluorescence microscope (Ni-E; Nikon-Europe). Images were acquired with DS-Fi1c camera (Nikon-Europe) and Brn3a-positive cells were counted using NIS-Elements software (Nikon-Europe).
9
Retinal Vasculature Examination in Mice
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For inner BRB assessment, 3 mice for each age group were anesthetized by intraperitoneal injection of avertin (1.2% tribromoethanol and 2.4% amylene hydrate in distilled water, 0.02 mL/g body weight; Sigma-Aldrich, St. Louis, MO, USA). Mice underwent systemic perfusion with 0.5% Evans blue dye (Sigma-Aldrich, St. Louis, MO, USA) in PBS intracardially injected into the left ventricle for 10 min. Then, mice were euthanized and explanted retinas were flat-mounted onto glass slides. Retinas were examined using an epifluorescence microscope (Ni-E; Nikon-Europe, Amstelveen, The Netherlands) equipped with a 10x Plan-Apochromat objective and a digital camera (DS-Fi1c camera, Nikon-Europe, Amstelveen, The Netherlands).
10
Quantifying Whitefly Neurodegeneration
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Whiteflies were collected and placed in 4% paraformaldehyde fixative overnight at 4°C, washed in 70% ethanol and processed into the Tissue-Tek O.T.C. Compound (Sakura). After freezing at −20°C, the embedded whiteflies were sectioned at 10 μm using a Leica CM1950 freezing microtome and stained with H & E according to the standard protocol. Images were taken using a Nikon light microscope, equipped with a DS-Fi1c camera (Nikon), and the images were generated using the NIS-Element D software (Nikon). The appearance of vacuolar lesions in the brain neuropil was the typical symptom of neurodegeneration, and six levels of neurodegeneration (0, 1, 2, 3, 4, and 5) were defined for quantification in previous research (Cao et al., 2013 (link)). The same standard was applied to quantify the neurodegeneration of whiteflies.
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