Liver samples were fixed 10% NBF for 24 h and processed for paraffin embedding. After cutting, the 5 μm sections were stained with hematoxylin and eosin (H&E) and examined using a Nikon microscope (Eclipse E200-LED, Tokyo, Japan). Other sections were stained with anti-Bax as previously described [40 (link),41 ]. Briefly, the tissue sections were blocked in 3% hydrogen peroxide (H2O2) and then probed with a rabbit polyclonal Bax antibody overnight at 4 °C. The sections were washed and probed with a biotinylated secondary antibody, followed by streptavidin/peroxidase conjugate for 30 min and then diaminobenzidine. The sections were counterstained with hematoxylin and examined using a Nikon microscope (Eclipse E200-LED, Tokyo, Japan).
Eclipse e200 led
Manufactured by Nikon
Sourced in Japan
The Eclipse E200-LED is a research-grade microscope from Nikon. It features LED illumination and is designed for a variety of microscopy applications.
Automatically generated - may contain errors
Lab products found in correlation
Rmflt3l, by Thermo Fisher Scientific (1 mentions)
Rmscf, by Thermo Fisher Scientific (1 mentions)
Diaphot 200, by Nikon (1 mentions)
Methocult gf m3434, by STEMCELL (1 mentions)
Anti pcna antibody, by Santa Cruz Biotechnology (1 mentions)
Anti bax antibody, by Santa Cruz Biotechnology (1 mentions)
Anti bcl 2 antibody, by Santa Cruz Biotechnology (1 mentions)
Szx illb200, by Olympus (1 mentions)
Orange g, by Merck Group (1 mentions)
Alcian blue, by Merck Group (1 mentions)
Ds fi1 camera, by Nikon (1 mentions)
Histostain plus bulk kit, by Thermo Fisher Scientific (1 mentions)
Axioskop microscope, by Zeiss (1 mentions)
Microtome, by Leica camera (1 mentions)
Nis br, by Nikon (1 mentions)
50 protocols using eclipse e200 led
1
Histological Analysis of Liver Tissue
2
Quantification of Fish Sperm Characteristics
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Milt was collected using a micropipette with a sterile micro tip that was adjusted every 0.01 mL (Cejko et al., 2011) , and the total volume of expressible milt was noted. The volume of milt was determined by dividing the expressible milt by the total body weight of the fish.
Milt was diluted 1000-fold with an immobilizing solution containing 153 mM NaCl in order to prevent sperm aggregation and achieve the proper concentration for counting. Sperm concentrations were measured by a hemocytometer (Zadmajid et al., 2018) (link). A compound Eclipse E200-LED light microscope connected to a video display (× 100 and × 400 magnification) was used for cell counting.
The sperm motility was analyzed by depositing 1 mL of sperm on a slide glass. The assessments were conducted at a room temperature of 26 • C. Sperm motility was measured using a video recorder connected to a threenocular microscope (Eclipse E200-LED, Nikon, Japan) (100 × and 400 × magnification). Based on Rurangwa et al. (2004) (link), a semi-quantitative technique was used to quantify the sperm motility.
For staining procedure, a mixture of 1 mL milt and 1 mL eosin 2 % was utilized to assess sperm viability using a light microscope (Eclipse E200-LED, Nikon, Japan) connected to a video monitor (× 1000 magnification). The process was examined for 15 s.
Milt was diluted 1000-fold with an immobilizing solution containing 153 mM NaCl in order to prevent sperm aggregation and achieve the proper concentration for counting. Sperm concentrations were measured by a hemocytometer (Zadmajid et al., 2018) (link). A compound Eclipse E200-LED light microscope connected to a video display (× 100 and × 400 magnification) was used for cell counting.
The sperm motility was analyzed by depositing 1 mL of sperm on a slide glass. The assessments were conducted at a room temperature of 26 • C. Sperm motility was measured using a video recorder connected to a threenocular microscope (Eclipse E200-LED, Nikon, Japan) (100 × and 400 × magnification). Based on Rurangwa et al. (2004) (link), a semi-quantitative technique was used to quantify the sperm motility.
For staining procedure, a mixture of 1 mL milt and 1 mL eosin 2 % was utilized to assess sperm viability using a light microscope (Eclipse E200-LED, Nikon, Japan) connected to a video monitor (× 1000 magnification). The process was examined for 15 s.
3
Histological Tissue Processing and Staining
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The liver, kidney, and pancreas were washed in saline and fixed in 10% neutral buffered formalin for 24 hours. Ascending grades of ethyl alcohol were then used to dehydrate the samples, and xylene was used as the clearing agent. The samples were then mounted in molten paraplast at 58°C–62°C; 4–5 μm slices were cut from the prepared blocks and stained with hematoxylin and eosin. The preparations were visualized using a Nikon microscope (Eclipse E200-LED, Tokyo, Japan).
4
Testes Histological Analysis Protocol
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The testes were fixed in 10% neutral buffered formalin for 24 h at room temperature. The tissues were dehydrated using an ascending series of alcohol concentrations, cleared in xylene, and embedded in paraffin wax. Paraffin blocks were sectioned at 5 μm thickness then hydrated and stained with hematoxylin and eosin according to the Drury and Wallington [28 ] method. Sections were examined using a Nikon microscope (Eclipse E200-LED, Tokyo, Japan). Images represent the 400x magnification.
5
Immunohistochemical Analysis of NF-κB, iNOS, and αSMA
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To investigate NF-κB, iNOS and αSMA, the paraffinized sections were blocked with hydrogen peroxide (0.1%) containing methanol for 15 min to damage the endogenous peroxidase. After blocking, the samples were incubated with the primary antibodies at 4 °C overnight. Thereafter, the samples were washed twice with phosphate-buffered saline and incubated with biotinylated secondary antibody labelled with horseradish peroxidase (HRP). The reactions were developed via an HRP-catalysed reaction with diaminobenzidine (DAB), followed by counterstaining with haematoxylin. Images were recorded at 400× magnification (Nikon Eclipse E200-LED, Tokyo, Japan). Afterward, the color intensity of each examined protein was semi-quantitatively scored by a blinded pathologist. The intensity was expressed as + (weak immunoreaction), ++ (moderate immunoreaction), +++ (strong immunoreaction), or ++++ (very strong immunoreaction).
6
Histological Analysis of Burned Tissues
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Burned tissues were dehydrated in escalating grades of ethanol, cleaned with xylene, and embedded in molten paraplast before being fixed in 10% neutral buffered formalin for 24 h. Hematoxylin and eosin were used to stain the finished blocks, which were divided into pieces that were 4–5 m thick. A Nikon microscope (Eclipse E200-LED, Tokyo, Japan) was used to examine tissue slices at the microscopic level. For the purpose of observing the deposition of collagen fibres, additional sections were dyed with Sirius Red.
7
Histological Analysis of Kidney Tissue
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The right kidney was fixed in 10% neutral buffered formaldehyde for 24 hours, dehydrated in alcohol, cleared in xylene, and mounted in molten paraffin wax. The paraffin-embedded tissues were sectioned at 4–5 µm and stained with hematoxylin and eosin. Sections were examined using a light microscope (Eclipse E200-LED; Nikon Corporation, Tokyo, Japan).
8
Histopathological Evaluation of Hippocampal Injury
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The hippocampal tissue was kept for 24 h at room temperature in 10% neutral-buffered formalin as a fixative, then dehydrated, paraffinized, and sectioned (4–5μm). Sections were stained with hematoxylin and eosin for light microscopy analysis. Light microscope photomicrographs were captured using a Nikon microscope (Eclipse E200-LED, Tokyo, Japan) at an original magnification of × 400. The severity of the brain injury was graded using the numerical method that described by Al-Quraishy et al41 with five grades of severity of injury specified as follows: 1 = minimal (<1%); 2 = slight (1%–25%); 3 = moderate (26%–50%); 4 = moderate/severe (51%–75%); and 5 = severe (76%–100%).
9
Myeloid and Erythroid Differentiation Assay
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De novo isolated or post–48-h liquid culture cells were plated in MethoCult GF M3434, supporting myeloid and erythroid differentiation, or MethoCult GF M3630, supporting preB cell differentiation (STEMCELL Technologies), supplemented with 0–20 ng/ml rmFlt-3L and 0–100 ng/ml rmSCF (PeproTech), and cultured at 37°C and 5% CO2. Colonies were scored between days 7 and 10 after plating, and images were captured using an inverted microscope (Diaphot 200; Nikon) at 4× with SPOT imaging software (v.5.0.15). Single representative colonies were isolated and used to confirm cell lineage by FACS analysis for the surface markers CD11b (M1/70), Gr-1 (RB6-8C5), and B220 (RA3-6B2) and Wright-Giemsa staining of fixed cytospins. Cytospin images were captured using an inverted microscope (Eclipse E200-LED; Nikon) at 100× with SPOT imaging software.
10
Immunohistochemical Analysis of Apoptosis Markers
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For immunohistochemistry investigations, the paraffin sections were mounted on glass slides and dewaxed. The antigen sites were revealed by washing the sections with boiled water after treatment for 10 min with 0.03% H2O2 in absolute methanol to stop endogenous peroxidase activity. Sections were incubated at 4 °C overnight with (1:50) polyclonal rabbit anti- Bcl-2 antibody, anti-Bax antibody, anti-caspases-3 antibody and anti-PCNA antibody (Santa Cruz, CA, USA). To get rid of the unbound primary antibodies, sections were washed with phosphate buffer saline (PBS). Afterward, sections were incubated for 30 min with goat-derived secondary anti-rabbit antibody conjugated to horseradish peroxidase at 37 °C. Antigen-antibody interactions were finally detected by incubating the sections for 10 min at room temperature with the chromogen 3,3′-diaminobenzidine tetrachloride (DAB-H2O2) as substrate. Testicular sections were visualized using 400× magnification lens (Nikon Eclipse E200-LED, Tokyo, Japan).
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