Cells were seeded to ultra-low attachment 6-well plates (Corning 3,471) with cell density of 2,500 and 3,000 cells per well for BT549 and HS578t, respectively. Cells were cultured in mammosphere medium, containing DMEM/F12 supplemented with B27 (Gibco 12587010) and 20 ng/ml EGF (R&D 236-EG), for 7–8 days. Images of mammospheres were captured by Nikon NIS-Elements D software. Mammosphere with diameter >50 µm were counted using the Nikon NIS-Elements D software.
Nis elements d software
Manufactured by Nikon
Sourced in Japan, United Kingdom, Canada
NIS-Elements D is a comprehensive software suite for microscope imaging and analysis. It provides a range of tools and functionalities to capture, process, and analyze images from a variety of Nikon microscopes. The software offers core functions for image acquisition, visualization, and measurement, enabling users to perform various microscopy tasks efficiently.
Automatically generated - may contain errors
Lab products found in correlation
Bx43 microscope, by Olympus (4 mentions)
Ds fi3 camera, by Nikon (4 mentions)
Las x software, by Leica (2 mentions)
Ni e compound microscope, by Nikon (2 mentions)
Rm2135 rotary microtome, by Leica (2 mentions)
Ds fi3, by Nikon (2 mentions)
Hoechst 33342, by Thermo Fisher Scientific (2 mentions)
Eclipse ti u, by Nikon (2 mentions)
Eclipse 80i microscope, by Nikon (2 mentions)
Epidermal growth factor (egf), by R&D Systems (1 mentions)
Eclipse ti e, by Nikon (1 mentions)
Cfi plan fluor, by Nikon (1 mentions)
Ds fi1, by Nikon (1 mentions)
Ccd camera, by Nikon (1 mentions)
Ds qi2 camera, by Nikon (1 mentions)
Eclipse ni u microscope, by Nikon (1 mentions)
Vectashield antifade mounting medium, by Vector Laboratories (1 mentions)
Alexa fluor 594 conjugated donkey anti rabbit, by Thermo Fisher Scientific (1 mentions)
Fdx 35, by Nikon (1 mentions)
Eg1150h machine, by Leica (1 mentions)
52 protocols using nis elements d software
1
Mammosphere Formation Assay Protocol
2
Muscle Fiber and Nuclei Analysis
3
Muscle Fiber and Nuclei Analysis
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For myofiber cross-sectional area measurements, images of the entire muscle cross-section were captured and then the cross-sectional area of at least 70 randomly selected myofibers of each myofiber type (Type I, IIa, IIx, and IIb) were measured by tracing the laminin stained periphery of individual myofibers with Nikon NIS-Elements D software (Nikon). For myonuclei and interstitial nuclei identification, 3–5 randomly selected 8.4 mm2 regions of each muscle cross-section were captured, and then all TRIM28 and P-TRIM28(S473) positive nuclei, as well as total nuclei were identified using both the NIS-Elements D software (Nikon) and the Leica LASX software (Leica). The distinction between myonuclei versus interstitial nuclei was achieved by defining nuclei that resided within the dystrophin layer of individual myofibers as myonuclei, while those that resided outside of the dystrophin layer were designated as interstitial nuclei. All image analyses were performed by investigators that were blinded to the sample identification.
4
Visualization of Gold Nanorod Uptake in A549 Cells
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A549 cells were plated onto 22 × 22 mm glass cover-slips in a 6-well plate at a density of 1 × 105 per well and allowed to grow for two days. The DMEM medium was then replaced with 2 ml of the same medium containing each AuNR type at a concentration of 1 × 1011 NP ml−1. After 4 h incubation, the AuNR-medium was removed and the cell monolayer on the cover-slip was twice-rinsed with DPBS (14190-094, Life Technologies, UK), fixed in 4% paraformaldehyde/DPBS for 10 min at room temperature and rinsed with DPBS twice. The fixed coverslips were mounted and sealed onto glass slides. Bright and dark-field microscopy imaging was performed with an inverted microscope (Nikon Eclipse Ti-E, Nikon UK Ltd, UK) and an oil coupled 100× objective (CFI Plan Fluor, Nikon UK Ltd, UK). Images were recorded with a 5 Megapixel colour camera (DS-Fi1, Nikon UK Ltd, UK) and saved using the NIS-Elements D software (Nikon UK Ltd, UK). Open-source software package ImageJ69 (link) was used to crop and enhance the contrast of saved images.
5
Visualizing cAMP Wave Propagation
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To visualize cAMP wave propagation, 5x105 cells/cm2 were plated on 1% non-nutrient agar plates and developed in dark moist conditions at 22°C. On a real-time basis, the aggregates were filmed at an interval of 30 s/frame, using a Nikon CCD camera and documented with NIS-Elements D software (Nikon, Japan). For visualizing cAMP optical density waves, image pairs were subtracted [92 (link)] using Image J (NIH, Bethesda, MD).
6
Morphological Analysis of Biological Samples
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Morphological characteristics were evaluated by optical microscopy (Nikon Eclipse NI optical microscope, Nikon Instruments, Amsterdam, Netherlands) after staining with May-Grünwald Giemsa medium (Sigma 32856). The photographs were obtained from a Nikon OS-Fi2 camera (Nikon Metrology Inc, Irvine, CA, USA) and the images were subsequently analyzed using the NIS-Elements D software (version 4.00). Photographs with magnifications of 100×, 200× and 500× were obtained.
7
Mammosphere Formation and Propagation
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For first generation of mammosphere formation, cells were seeded to ultra-low attachment 6-well plate (Corning 3471) with cell density of 1000 and 2500 cells per well for HCC1806 and MDA-MB-468 cells, respectively. Cells were cultured in mammosphere medium, containing DMEM/F12 supplemented with B27 (Gibco 12587010) and 20 ng/ml EGF (R&D 236-EG) for 5–6 days. Images of mammospheres were captured by the Nikon Eclipse Tis2 microscope at ×4 magnification objective. Number of mammospheres with diameter ≥70 µm were counted using the Nikon NIS-Elements D software. Second-generation mammosphere-formation assay was performed as previously described [28 (link)]. Briefly, mammospheres from first generation were collected, dissociated by trypsin and washed by mammosphere medium once. All cells were then seeded to a new ultra-low attachment 6-well plate with similar densities and cultured for 5–6 days. Ratio of second-generation mammosphere number to first-generation mammosphere number was calculated.
8
Assessing Cryopreservation Outcomes through Larval Abnormalities
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Percentage of abnormal D-larvae was calculated as an indicator of the toxicity of the CPAs. On the contrary, the percentage of normal D-larvae developed after thawing was used to assess cryopreservation outcome. Both parameters were obtained after examining n = 100 cells for each replicate and treatment, under the microscope Nikon ECLIPSE 2000-5 and using Nikon NIS Elements-D software, version 4.13. The discrimination between normal and abnormal D-larvae was determined under a microscope attending to previous work focused on shell larval morphology and guidelines from other experts in the shell abnormalities and abnormally developing larvae of related mollusks in ecotoxicological larval bioassays25 (link),53 (link),54 (link),62 (link),63 (link). Typical larval abnormalities found included delayed development, deviations from the D-larvae shell shape like indented margins or hinge deformations (concave or convex hinges) or presence of clear protruding mantle.
9
Quantifying DNA Damage via γH2AX
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Images of immunofluorescence-stained samples (prepared as described in the previous section) were taken using an ECLIPSE Ni-U with DS-Qi2 camera (Nikon, Tokyo, Japan) with a 40× lens, and analyzed using NIS-Elements D software (Nikon). For each experimental setting, at least 200 cells within four to five fields were recorded. The γH2AX and DAPI signals were identified by ImageJ (version 1.48, National Institutes of Health, Bethesda, MD, USA), from which intranuclear occupancy by γH2AX signals was calculated. Unhit cell was defined as cell in which intranuclear occupancy by the γH2AX signals was less than 1%.
10
In situ Hybridization for HIV and KSHV
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For in situ hybridization (ISH), we used a RNAscope 2.5 duplex detection kit and chromogenic probes (Advanced Cell Diagnostics, Hayward, CA) to detect HIV and KSHV signals in the KS samples. The detection process was conducted in accordance with the manufacturer’s instructions. HIV and KSHV reactive particles were marked in red and green. Images were captured using an ECLIPSE Ni-U microscope and processed using NIS-Elements D software (Nikon, Tokyo, Japan).
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