Human brain sections (50 μm) were pre-treated with 10 Mm Tris, pH 6.0, at 85°C for 5 min. The slices were treated with formic acid (20%) for 20 min to detect the β-amyloid protein. The mouse (50 μm coronal sections) and human brain sections were “treated for 20 min in 0.1 M PB:methanol (1:1) with 1% hydrogen peroxide and then incubated for 1 h in 0.1 M PB with 1% BSA and 0.3% Triton X-100. Slices were then incubated with primary antibodies, mouse anti-human amyloid beta protein (1:250, SIGNET) and rabbit anti-BACE1 C-terminal (485–501; 1:100, Calbiochem), overnight at 4°C in 0.1 M PB with 0.3% BSA and 0.3% Triton X-100. The slices were incubated with a biotinylated mouse secondary antibody and then incubated with ABC-HRP complex (Pierce Biotechnology) for 2 h. Diaminobenzidine (DAB) was used to develop the staining. The tissues were dehydrated, covered with mounting solution and observed on an Eclipse E200 optical microscope (Nikon)”.
Eclipse e200 optical microscope
Manufactured by Nikon
Sourced in Japan
The Eclipse E200 is an optical microscope designed for high-quality observation and imaging. It features a sturdy, ergonomic construction and offers a range of magnification options to accommodate various sample types. The Eclipse E200 is a reliable and versatile tool for laboratory applications.
Automatically generated - may contain errors
Lab products found in correlation
Abc hrp complex, by Thermo Fisher Scientific (2 mentions)
Protein quantification kit, by Bio-Rad (1 mentions)
Vetflurano, by Virbac (1 mentions)
Paraplast, by Merck Group (1 mentions)
Ab81289, by Abcam (1 mentions)
Sc 56, by Santa Cruz Biotechnology (1 mentions)
Dab hrp brown detection system, by Bio SB (1 mentions)
9 protocols using eclipse e200 optical microscope
1
Immunohistochemical Analysis of Amyloid-Beta and BACE1
2
Immunohistochemical Evaluation of NOTCH1 Expression
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The immunohistochemical evaluation was performed by a senior pathologist and an experienced examiner of the Instituto Nacional de Cancerología-México in a double-blind fashion. A Nikon ECLIPSE E200 optical microscope with a 10x eyepiece and 10x and 40x objective lens was used. The staining intensity was defined as: weak (including + and ++), when the immunoreaction was visible only with a 40x objective; and intense (+++), when staining was visible even using a 10x objective. Localization was categorized as membranal, cytoplasmic, nuclear, or combinations among them. Percentage of positivity was assessed by quartiles. Technique validation was performed using an alternative NOTCH1 antibody in a representative set of samples.
3
Quantifying Inflammatory Markers in BALF
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The remaining rats in each group were sacrificed 24 h following CLP and a tracheal intubation was performed upon separating the trachea and the main bronchus. The left lung was washed three times with 0.5 ml pre-cooled PBS. The BALF was centrifuged for 10 min at 1,500 × g at 4°C. The protein concentrations in the BALF were detected using a protein quantification kit (Bio-Rad Laboratories, Inc., Hercules, CA, USA), according to the manufacturer's protocol. The levels of TNF-α (cat. no. ab208348), IL-1β (cat. no. ab100704) and IL-6 (cat. no. ab100712) in the BALF were determined using ELISA kits (Abcam, Cambridge, UK). The TNF-α, IL-1β and IL-6 content in the samples was calculated using a standard calibration curve. The detection ranges of the TNF-α, IL-1β and IL-6 ELISA assays were 12.5–800, 31.25–2,000 and 62.5–4,000 pg/ml, respectively. Concentrations of samples that were initially over the limit of the standard curve were measured following dilution.
The BALF cells were resuspended with PBS, and a small number of the suspended cells were dropped onto a slide. Following Wright-Giemsa staining for 10 min at room temperature, polymorphonuclear neutrophils (PMN) were counted using an Eclipse E200 optical microscope (Nikon Corporation, Tokyo, Japan) under a high-power field (magnification, ×400). A total of five randomly selected non-overlapping fields on each slide were observed.
The BALF cells were resuspended with PBS, and a small number of the suspended cells were dropped onto a slide. Following Wright-Giemsa staining for 10 min at room temperature, polymorphonuclear neutrophils (PMN) were counted using an Eclipse E200 optical microscope (Nikon Corporation, Tokyo, Japan) under a high-power field (magnification, ×400). A total of five randomly selected non-overlapping fields on each slide were observed.
4
Yeast Budding Lifespan Analysis
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The budding lifespan of individual mother yeast cells was defined as the number of mitotic cycles (buddings) during cell’s life. After overnight growth, the cells were arrayed on a YPD plate using a micromanipulator. The budding lifespan was determined microscopically by a routine procedure with the use of a micromanipulator as described previously [39 (link)]. The number of buds formed by each mother cell reflects its reproductive potential (budding lifespan). In each experiment, 45 single cells were analyzed. The results represent measurements for at least 90 cells analyzed in two independent experiments. The analysis was performed by micromanipulation using the Nikon Eclipse E200 optical microscope with an attached micromanipulator.
5
Quantifying Extracellular Amyloid Plaques in Aged Transgenic Mouse Brains
6
Yeast Lifespan Modulation by Curcumin
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The total lifespan was defined as the length of life of a single mother cell expressed in units of time. The total lifespan was calculated as the sum of reproductive and post-reproductive lifespans. The reproductive lifespan was defined as the length of time between the first and the last budding, and the post-reproductive lifespan as the length of time from the last budding until cell death. The lifespan of the Saccharomyces cerevisiae yeast was determined as previously described by (Minois et al. 2005 (link)) with small modification (Molon et al. 2018 (link)). Ten microliter aliquots of an overnight grown culture of yeast were collected and transferred on YPD plates with solid medium containing Phloxine B (10 μg/mL) and without (control) or with 200 μM and 300 μM curcumin. Phloxine B was used to stain Saccharomyces cerevisiae dead cells. Dead yeast cells lose membrane integrity and Phloxine B entered cell space giving pink/red coloration of cytosol. In each experiment, forty-five single cells were analysed. During manipulation, the plates were kept at 28 °C for 15 h and at 4 °C during the night. The results represent measurements for at least 90 cells analysed in two independent experiments. The analysis was performed by micromanipulation using the Nikon Eclipse E200 optical microscope with an attached micromanipulator.
7
Quantitative Analysis of Liver Fibrosis
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Six weeks after Bdl, the rats from each experimental group were euthanized with an excess of isoflurane (Vetflurano®; Virbac, São Paulo, Brazil), and liver samples were collected from the medial lobe and right lobe. The specimens were fixed in 10% buffered formalin for 48 h and embedded in Paraplast (SigmaAldrich, Germany). Four-µm-thick sections were obtained with a semiautomatic microtome, and stained with masson's trichrome for fibrosis and collagen analysis, Bras Cienc (2019) 91(3) e20190220 4 | 14 according to the manufacturer's instructions or placed on polysinine-l coated slides for immunohistochemical assessment.
QUAlITATIVe ANAlYSIS OF THe FIBROSIS deGRee
The degree of liver fibrosis was qualitatively assessed using a Nikon eclipse e200 optical microscope at 100x magnification by the Knodell scoring system, which classified the histopathological alterations, as follows (Brunt 2000) : score 0: no fi brosis; score 1: fi brous portal expansion; score 3: bridging fi brosis (portal-portal or portal-central); score 4: cirrhosis. The average score taken from 10 non-overlapping random fi elds per section was used to generate a single score for each specimen, in each experimental group (Sant'anna et al. 2011a ).
QUAlITATIVe ANAlYSIS OF THe FIBROSIS deGRee
The degree of liver fibrosis was qualitatively assessed using a Nikon eclipse e200 optical microscope at 100x magnification by the Knodell scoring system, which classified the histopathological alterations, as follows (Brunt 2000) : score 0: no fi brosis; score 1: fi brous portal expansion; score 3: bridging fi brosis (portal-portal or portal-central); score 4: cirrhosis. The average score taken from 10 non-overlapping random fi elds per section was used to generate a single score for each specimen, in each experimental group (Sant'anna et al. 2011a ).
8
Optical Microscopy of Fungal Propagules
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For optical microscopy, samples of either MS or P propagules were taken at 24, 48, 72, and 96 h post-inoculation of liquid media, centrifuged to recover propagules, washed twice with sterile water, and observed with a Nikon eclipse e200 optical microscope (Nikon, Japan) at 100× magnification.
9
Immunohistochemical Analysis of Tumor Angiogenesis
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Tumors were excised, processed, and embedded in paraffin. Consecutive tumor sections of 4 µm thickness were cut and mounted on positively charged slides. Immunohistochemistry was performed to evaluate PN; PI was evaluated with an anti-PCNA primary antibody (sc-56, dilution 1:500; Santa Cruz Biotechnology, Inc., Dallas, TX, USA); and MVD was evaluated with an anti-CD34 primary antibody (ab81289, dilution 1:500; Abcam, Cambridge, UK). Positive staining was detected with the DAB HRP Brown detection system (Bio SB, Santa Barbara, CA, USA). Negative controls corresponded to incubation without primary antibodies; human tonsil and vascular tumor were used as positive controls for anti-PCNA and anti-CD34, respectively. PN was evaluated as the percentage of necrotic tissue contained in the tumor, PI was estimated as the percentage of positive cells to anti-PCNA in the viable tumor, and MVD was quantified as the mean value of the vessel count in 10 high-power fields of hot spots [49 (link)]. A Nikon ECLIPSE E200 optical microscope (Nikon Instruments Inc., Melville, NY, USA) with a 10× eyepiece and 10× and 40× objective lens was used.
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