Images were acquired with a Nikon Eclipse Ni-U epifluorescence microscope running NIS-elements software (version 5.11.03; Nikon). Immunofluorescence was visualized with an LED illumination system (X-Cite 120 LED Boost, Excelitas Technologies) and captured with a Nikon DS-Qi2 digital camera. Immunofluorescence images were acquired using Plan Fluor 4×, Plan-Apochromat 10× differential interference contrast (DIC) N1 or Plan Fluor 20× DIC N2 objectives. Bright-field images were acquired with a 10× objective on an Olympus BX61 microscope. Figures were made using Photoshop version 22.5 (Adobe). When brightness and/or contrast adjustments were made in a figure, these changes were made equally to all photomicrographs.
Eclipse ni u epifluorescence microscope
Manufactured by Nikon
The Eclipse Ni-U epifluorescence microscope is a laboratory equipment designed for fluorescence microscopy. It features a high-performance optical system and a comprehensive range of fluorescence filter sets to enable detailed observation and analysis of fluorescently labeled samples.
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4 protocols using eclipse ni u epifluorescence microscope
1
Nikon Epifluorescence Microscopy Protocol
2
Fluorescence imaging of microbial communities
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Cells were fixed in growth medium with the addition of 2.5% formaldehyde for 2 h at room temperature. After this period, they were washed three times in 1× phosphate-buffered saline and then stored at −20 °C in a mixture of absolute ethanol and 1× phosphate-buffered saline (1:1). The oligonucleotide probes used in this study were the general bacteria-targeting EUB338 (ref. 59 (link)), the Methanomicrobiales-targeting MG1200 (ref. 60 (link)) and the Lokiarchaea-specific DSAG-Gr2-1142 (ref. 7 (link)) (Supplementary Table 5 ). The DNA-hybridization chain reaction procedure was performed as described previously61 (link) and the samples were imaged on the Eclipse Ni-U epifluorescence microscope (Nikon) using Gryphax (v.1.1.8.153). Images were processed using ImageJ62 (link).
3
Quantifying Neuronal Marker Expression
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H9 TrkB‐, H9 TrkC‐, H9 Tuj1‐, A10 TrkB‐, A10 TrkC‐ and A10 Tuj1‐positive cells were counted in 20 randomly selected fields, two coverslips, each using a Nikon Eclipse Ni‐U epifluorescence microscope at 40X magnification. The number of TrkB‐ or TrkC‐positive cells was found not to be significantly different from the number of Tuj1‐positive cells, neither for H9 (TrkB, TrkC) nor A10 (TrkC) neurons.
4
Telomere Length Analysis via FISH
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Microscope observations were carried out with the Nikon Eclipse Ni-U epifluorescence microscope under 100 × and 60 × planachromatic immersion objectives. Extended-depth-of-focus (EDF) images were obtained by capturing 10–15 different focal planes of the same object by a cooled monochrome DSQi1 camera (Nikon), stacking them and combining using EDF function of the NIS Elements software (Laboratory Imaging, Ltd.). The intrachromosomal (TTTAGGG)n signals can be generated by all FISH-techniques and all telomeric probes, but they quickly fade, which requires finishing acquisition within 1–2 days after mounting the preparations. The superimposing and uniform image processing of the FISH-images and chromosome measurements were performed using Adobe Photoshop (Adobe Systems) and ImageJ software (Fiji package, https://imagej.net/Fiji/Downloads ). Six best mitotic metaphases from each plant were added to the pool for statistical calculations (in total 90 metaphase plates), which were done with Microsoft Excel 2016.
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