To analyze the ability of cells to enter stationary phase, overnight cultures were diluted to a final concentration of 1 × 105 cells/ml in YPD and incubated at 30°C. At 1, 2, 3, 4, and 5 days after starting the culture, 1 ml of cell suspension was harvested. Cell concentration was enumerated using a hemocytometer, and cell morphology was determined by analysis of at least 100 cells on a Leica DM750 microscope (Leica Biosystems, Wetzlar, Germany). For ploidy analysis, cells were fixed with 70% ethanol, stained with PI, and analyzed by flow cytometry as described above. To assess cells after stationary-phase release, cells were grown for 96 h under nutrient starvation. Cells were then centrifuged to pellet the cells, washed with PBS twice, and resuspended to a final concentration of 2.5 × 105 cells/ml in YPD broth and incubated at 30°C with shaking at 250 rpm. The cell morphology was determined by analysis of at least 100 cells per time point on a Leica DM750 microscope (Leica Biosystems, Wetzlar, Germany).
Dm750 microscope
Manufactured by Leica
Sourced in Germany, Switzerland, United States, China
The Leica DM750 is a high-quality microscope designed for laboratory use. It features a sturdy construction, a range of magnification levels, and various illumination options to support a variety of scientific applications. The core function of the DM750 is to provide clear, detailed imaging of samples under observation.
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Lab products found in correlation
Icc50 hd camera, by Leica (9 mentions)
Icc50w camera, by Leica (5 mentions)
Icc50, by Leica (5 mentions)
Las ez software, by Leica (5 mentions)
Diaminobenzidine tetrahydrochloride, by Vector Laboratories (3 mentions)
Axio imager z1m microscope, by Zeiss (3 mentions)
Application suite 4, by Leica (3 mentions)
Laz ez software, by Leica (3 mentions)
Icc50 hd digital camera, by Leica (3 mentions)
Avidin biotin hrp complex reagent solution, by Vector Laboratories (2 mentions)
M205a stereomicroscope, by Leica (2 mentions)
Szx9 stereomicroscope, by Olympus (2 mentions)
Phalloidin tritc, by Merck Group (2 mentions)
Alexa fluor 488 594 conjugated secondary antibodies, by Thermo Fisher Scientific (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Biotium (2 mentions)
Nis elements software, by Nikon (2 mentions)
Mc120 hd, by Leica (2 mentions)
Cm1860, by Leica (2 mentions)
Image pro software, by Media Cybernetics (2 mentions)
Ventana benchmark system, by Roche (2 mentions)
149 protocols using dm750 microscope
1
Stationary Phase Characterization in Yeast
2
Histological Analysis of Liver and eWAT
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Tissues from the liver and eWAT were necropsy-dissected and fixed in a 4% paraformaldehyde solution for at least 24 h. After dehydrating with ethanol and embedding the tissue in paraffin, the staining of tissues with a thickness of 4 to 5 μm was performed by hematoxylin and eosin (H&E). Observation of histopathological changes was performed under a Leica DM750 microscope (Nussloch, Germany). The size of the adipocytes was calculated by Image J (version 1.53).
3
Histopathological Evaluation of Ovarian Tissue
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The ovarian tissues fixed in 4% paraformaldehyde buffer were embedded in paraffin after serial dehydration steps, then sectioned at a thickness of 4 μm and stained with hematoxylin for 8 min and eosin for 5 min (HE staining). The histopathological changes of the ovaries in the sections were viewed using a Leica DM750 microscope (Germany).
4
Assay for Senescence-Associated Beta-Galactosidase Activity in T Cells
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SA-β-gal activity in senescent T cells was detected as we described previously.10 12 15 (link) CD4+ or CD8+ T cells were cocultured with iTreg cells or nTreg cells at a ratio of 4:1 or 2:1 in anti-CD3/CD28-coated 24-well plates for different time points, and cocultured responder T cells were then stained for SA-β-gal expression. For tumor-induced T cell senescence analysis, anti-CD3/CD28 activated CD4+ T cells or CD8+ T cells were cocultured with or without tumor cells or control NIH/3T3 cells at ratio of 1:1 for 1 day and then separated and cultured for an additional 3 days. For SA-β-gal staining, T cells were washed in PBS (pH7.2), fixed in 4% formaldehyde, and incubated overnight at 37°C with freshly prepared SA-β-gal staining solution (1 mg/mL X-gal, 5 mM K3Fe (CN)6, 5 mM K4Fe (CN)6, 2 mM MgCl2 in PBS at pH6.0). The stained T cells were examined for SA-β-gal expression with a computerized image system composed of a Leica ICC50 camera system equipped on a Leica DM750 microscope.
For some experiments, the cocultured T cells were determined for SA-β-gal expression in the presence of various MAPK or ATM inhibitors. Concentrations of inhibitors used in this study were as following: U0126 (10 µM), SB203580 (10 µM), SP600125 (10 µM), and KU55933 (10 µM).
For some experiments, the cocultured T cells were determined for SA-β-gal expression in the presence of various MAPK or ATM inhibitors. Concentrations of inhibitors used in this study were as following: U0126 (10 µM), SB203580 (10 µM), SP600125 (10 µM), and KU55933 (10 µM).
5
BrdU Immunohistochemistry in Mouse Brain
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The mice were subjected to transcardial perfusion, their brains were removed and cryotome sections were collected. Every fifth section from the region between bregma -1.82 mm and -2.18 mm (according to Paxinos and WatsonMouse Brain Atlas) was analyzed. For DNA hydrolysis, tissue sections were washed twice in phosphate-buffered saline, incubated in 2 N HCl for 30 min at 37°C, following neutralization in borate buffer (0.1 M, pH 8.5) for 15 min. Immunohistochemistry (IHC) was performed with specific anti-BrdU primary antibodies (Oxford Biotechnology, Oxford, UK). After incubation with anti-BrdU, sections were incubated with the biotinylated secondary antibody for 1 h at 25 °C and then with avidin-biotin-HRP complex reagent solution (Vector Laboratories, Burlingame, CA, USA). The peroxidase reaction was finally performed using diaminobenzidine tetrahydrochloride (Vector Laboratories). Digital images of IHC staining were captured using a Leica DM750 microscope. Every fifth section was taken from the region between 1.46 to 2.80 mm posterior to bregma. BrdU-positive cells were quantified.
6
Phyllanthus microcarpus Fruit Herbivores
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The field study was conducted from 2011 to 2013 in Pingxiang, Guangxi Zhuang Autonomous Region, and from 2009 to 2014 in several nature reserves in Hainan Province, China. Specimens examined in this study were collected or reared from fruits of Phyllanthus microcarpus
The type specimens are deposited in the Insect Collection, College of Life Sciences, Nankai University, Tianjin, China and some paratypes are deposited in the
The type specimens are deposited in the Insect Collection, College of Life Sciences, Nankai University, Tianjin, China and some paratypes are deposited in the
Department of Entomology, Natural History Museum, London, UK
7
Insect Collection Preservation and Examination
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The present study is based on the examinations of the specimens collected by light traps. Adults were examined using an Olympus SZX9 stereomicroscope. Permanent mounting methods of genitalia and venation follow the techniques introduced by Li (2002) . Images of adults and genitalia were taken by using a Leica M205A stereo microscope and a Leica DM750 microscope.
All the studied specimens, including the types of the new species, are deposited in the
All the studied specimens, including the types of the new species, are deposited in the
Insect Collection of College of Life Sciences, Nankai University
8
Histological Assessment of Liver Samples
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Three gross sections of the liver samples were taken from the left lateral lobe, right medial lobe, and caudate lobe, kept in a tissue cassette with the respective identity, and fixed in 10% buffered neutral formalin solution until processing. Sections of 4 μm thickness were taken from each tissue block, stained with hematoxylin for 10 minutes, and counterstained in 2% eosin solution for 20 seconds. The stained slides were coverslipped using DPX mounting medium. Histomorphologic abnormalities were scored based on the type and severity of morphologic changes according to Yahya et al. [33 (link)]. Selected images of the sections were captured under a magnification of 10× by Leica DM750 microscope using ICC50 HD camera.
9
Immunofluorescence and Histological Staining
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Cells from the in vitro experiments were washed in PBS and fixed in 10% (v/v) formalin for 15 min (RT). Cell permeabilization was performed with 0.2% (v/v) Triton X-100 for 15 min (RT, intracellular staining) or 30 min (4 °C, nuclear staining), and blocking with a 3% (w/v) BSA solution for 30 min (RT). Samples were then incubated with primary antibodies (Supplemental Table S2 ) diluted in 1% BSA for 1 h (RT). After washing with PBS, Alexa Fluor (594/488)-conjugated secondary antibodies (1:500; Molecular Probes) were applied. Cellular morphology was observed after staining F-actin with Phalloidin-TRITC (1:100, Sigma-Aldrich) for 1 h (RT) and nuclear counterstain performed using DAPI (0.02 mg/ml, Biotium) for 15 min (RT). Images were acquired with an Axio Imager Z1m microscope (Zeiss).
For histological analysis, 4-μm paraffin-embedded sections were stained with hematoxylin and eosin (H&E), according to standard procedures, and analyzed with a DM750 microscope (Leica). For immunofluorescence studies, heat-mediated antigen retrieval was performed in the deparaffinized sections using the citrate buffer (pH 6.0), and the remaining procedure was performed as above described for the in vitro ones.
For histological analysis, 4-μm paraffin-embedded sections were stained with hematoxylin and eosin (H&E), according to standard procedures, and analyzed with a DM750 microscope (Leica). For immunofluorescence studies, heat-mediated antigen retrieval was performed in the deparaffinized sections using the citrate buffer (pH 6.0), and the remaining procedure was performed as above described for the in vitro ones.
10
Histopathological Liver Assessment Protocol
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Histopathological changes were investigated in formalin-fixed liver tissues (4-μm sections) using hematoxylin and eosin (H&E) staining, and necrotic areas were scored as described previously [23 (link)]. Modified Elastic Van Gieson (EVG) staining was performed using Sirius red to evaluate fibrosis areas. Specimens were examined under a DM750 microscope (Leica, Wetzlar, Germany). Fibrosis areas were evaluated using NIS-Elements software (Nikon instruments, Tokyo, Japan) as described previously [18 (link)].
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