Cell cultures were treated with 1 μM flavo for 1 h after which cells were washed and resuspended in prewarmed medium. Colcemid (Life Technologies) at a concentration of 0.1 μg/ml was added 150 min before harvest. Next, cells were swelled in 8 ml hypotonic buffer (20% FBS [v/v], 15 mM KCl) for 15 min. Next, 10 ml fixation buffer (25% acetone, 75% methanol) was added drop by drop. Cells were harvested by centrifugation at 1000 rpm for 5 min, followed by resuspension in 10 ml fixation buffer. Cells were stored at −20°C for 2 days. Finally, the cells were splatted onto the slides and mounted with cover slips using 5 μl of mounting medium containing DAPI (4% n-propyl gallate, 80% glycerol, 1 μg/ml DAPI). Metaphase chromosomes were visualized on a widefield microscope (AxioImager Z1; Carl Zeiss) equipped with a 100× objective lens (Plan Apochromat, NA 1.4; Carl Zeiss), a cooled CCD camera (Orca-ER; Hamamatsu Photonics), differential interference contrast (DIC), and an illumination source (HXP120C; Carl Zeiss);
Hxp 120c
Manufactured by Zeiss
Sourced in Germany
The HXP 120C is a high-performance mercury short-arc lamp designed for a variety of microscopy and imaging applications. It provides a stable, high-intensity light source with a color temperature of approximately 5,600K. The lamp's compact size and efficient thermal management make it suitable for integration into various laboratory equipment and systems.
Automatically generated - may contain errors
Lab products found in correlation
Axio observer z1, by Zeiss (3 mentions)
Orca er, by Hamamatsu Photonics (2 mentions)
Axio imager z1, by Zeiss (2 mentions)
Axiocam mrm rev 3 camera, by Zeiss (2 mentions)
Uncoated glass bottom dishes, by Ibidi (2 mentions)
Plan apochromat 1.4 oil immersion lens, by Zeiss (2 mentions)
Colcemid, by Thermo Fisher Scientific (1 mentions)
Imageem c9100 13, by Hamamatsu Photonics (1 mentions)
Axio scope a1 vario, by Zeiss (1 mentions)
Axiovision, by Zeiss (1 mentions)
L threonine, by Merck Group (1 mentions)
Vivatome, by Zeiss (1 mentions)
Axiocam, by Zeiss (1 mentions)
Rabbit anti gap43 antibody, by Abcam (1 mentions)
Anti gfap cy3, by Merck Group (1 mentions)
Donkey anti rabbit alexafluor 594, by Thermo Fisher Scientific (1 mentions)
Axio examiner z1, by Zeiss (1 mentions)
Rabbit anti iba1, by Fujifilm (1 mentions)
Axio observer z1 inverted microscope, by Zeiss (1 mentions)
Axio scope a1, by Zeiss (1 mentions)
10 protocols using hxp 120c
1
Flavo-induced Metaphase Chromosome Analysis
2
Fluorescence Microscopy of Lipid Monolayers
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Images of fluorescent monolayers at the air/water interface were recorded with an Axio Scope A1 Vario epifluorescence microscope (Carl Zeiss MicroImaging, Jena, Germany) while simultaneously recording the compression of the monolayer. The film balance (see above) below the microscope was mounted on an x-y-z stage (Märzhäuser, Wetzlar, Germany), which was motion controlled by a MAC5000 system (Ludl Electronic Products, Hawthorne, NY, U.S.A.). To ensure a dust-free environment and minimize the evaporation of water, the trough was enclosed by a home-built Plexiglas hood. The microscope was equipped with a compact light source HXP 120 C (mercury short arc reflector lamp), a long working distance objective (LD EC Epiplan-NEOFLUAR 50×), and a filter/beam splitter combination appropriate for the fluorescent dye, all from Carl Zeiss MicroImaging (Jena, Germany). Image data were recorded by an EMCCD camera (ImageEM C9100-13, Hamamatsu, Herrsching, Germany) and acquired by the software AxioVision (Carl Zeiss MicroImaging, Jena, Germany). All presented images show areas of individually contrast-adjusted raw data. Each lipid mixture was doped with 0.05 mol% Rh-DHPE (reducing the amount of brain PE by 0.05 mol%) and once with 1 mol% TopFluor® Cholesterol. The monolayers were spread and the protein was injected as described above.
3
Fluorescence Imaging of Induced Gene Expression
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Conidia of strain BD1181 were cultured in uncoated glass-bottom dishes (Ibidi® GmbH, Germany; 2.5 mL of medium per well) for 16 h at 25°C (Peñalva, 2005 (link)). After this period, the medium was replaced with fresh medium supplemented with 100 mM L -threonine (Sigma) in place of D -glucose to induce alcA(p)::GFP::fluG expression.
Fluorescence images were acquired using a Zeiss Axio Observer Z1 inverted microscope equipped with a 63x Plan Apochromat 1.4 oil immersion Lens, Axiocam MRm Rev.3 camera, a Zeiss HXP 120C external light source for epifluorescence excitation and fitted with filter set 38HE for green fluorescence (Ex BP 470/40; FT 495; Em BP 525/50) and filter set 43HE for red fluorescence (Ex BP 545/25; FT 570; Em BP 605/70). Numerous samples were observed before taking representative images. Fluorescence levels were measured using Fiji software (Schindelin et al., 2012 (link)).
Fluorescence images were acquired using a Zeiss Axio Observer Z1 inverted microscope equipped with a 63x Plan Apochromat 1.4 oil immersion Lens, Axiocam MRm Rev.3 camera, a Zeiss HXP 120C external light source for epifluorescence excitation and fitted with filter set 38HE for green fluorescence (Ex BP 470/40; FT 495; Em BP 525/50) and filter set 43HE for red fluorescence (Ex BP 545/25; FT 570; Em BP 605/70). Numerous samples were observed before taking representative images. Fluorescence levels were measured using Fiji software (Schindelin et al., 2012 (link)).
4
Structured Illumination Microscopy with FITC and TexasRed
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Images were taken with Axiovert 200 M microscope stage equipped with a mercury short-arc lamp (HXP120c; Carl Zeiss), a structured illumination-aperture correlation unit (VivaTome, Zeiss) with FITC (ex; 494/20–25, em; 536/40–25), and TexasRed (ex; 575/25–25, em; 628/40–25) activations in combination with a triple band dichroic mirror (436/514/604). A 40x (NA 1.3; Carl Zeiss) objective was used for all images. The detector for this system was an Axiocam MRm CCD camera with a pixel size of 6.45×6.45 μm.
5
Multiphoton Immunohistochemistry for Neural Markers
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Heat antigen retrieval in citrate buffer was performed followed by 0.3% TritonX for 15 min and blocking in 5% bovine serum albumin in PBS for 1 h. For GFAP immunohistochemistry, the tissue was probed with the antibody (Anti-GFAP-Cy3, 1:200, Sigma Aldrich) overnight at 4 °C. Rabbit anti- β-III-tubulin (1:500, Covance Inc., Princeton, NJ, USA), rabbit anti-Iba1 (1:200, Wako Chemicals GmbH, Neuss, Germany) and rabbit anti-GAP43 antibody (1:250, abcam, Cambridge, UK) were incubated overnight at 4 °C. After washing with PBS, sections were probed with fluorescent secondary antibodies (donkey anti-rabbit Alexa Fluor 594 (1:500, Invitrogen, Thermo Fisher Scientific, Waltham, MA, USA) at room temperature for 1 h. Stained fluorescent structures were visualized using a microscope Axio Examiner Z.1 equipped with camera AxioCam and metal halide lamp HXP 120c (Carl Zeiss AG, Jena, Germany), or by two-photon fluorescence using multiphoton microscopy.
6
Fluorescent Imaging of Tumor Slices
7
Imaging Fungal Conidiospore Response
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A. nidulans conidiospores were cultured in uncoated glass-bottom dishes (Ibidi GmbH, Germany; 2.5 ml of medium per well) for 16 h at 25°C in adequately supplemented pH 6.8 Käfer's minimal medium containing 0.1% D-glucose, 71 mM sodium nitrate and 25 mM sodium phosphate monobasic, similar to watch minimal medium (WMM; Peñalva, 2005 (link)). After this period, the medium was replaced with fresh medium supplemented with 100 μM CuSO4 to induce CrpA-GFP expression.
Fluorescence images were acquired using a Zeiss Axio Observer Z1 inverted microscope equipped with a 63x Plan Apochromat 1.4 oil immersion Lens, Axiocam MRm Rev.3 camera, an Zeiss HXP 120C external light source for epifluorescence excitation and fitted with filter set 38HE for green fluorescence (Ex BP 470/40; FT 495; Em BP 525/50) and filter set 43HE for red fluorescence (Ex BP 545/25; FT 570; Em BP 605/70). Same exposure time and microscope settings were applied for all image acquirement. Numerous cells were observed for each time before taking representative images. Fluorescence levels were measured using ImageJ software (http://imagej.nih.gov/ij ; U.S National Institutes of Health, Bethesda, Maryland, USA).
Fluorescence images were acquired using a Zeiss Axio Observer Z1 inverted microscope equipped with a 63x Plan Apochromat 1.4 oil immersion Lens, Axiocam MRm Rev.3 camera, an Zeiss HXP 120C external light source for epifluorescence excitation and fitted with filter set 38HE for green fluorescence (Ex BP 470/40; FT 495; Em BP 525/50) and filter set 43HE for red fluorescence (Ex BP 545/25; FT 570; Em BP 605/70). Same exposure time and microscope settings were applied for all image acquirement. Numerous cells were observed for each time before taking representative images. Fluorescence levels were measured using ImageJ software (
8
Monitoring Downy Mildew Pathogenesis
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Aniline blue staining was used to monitor P. viticola mycelium development according to Hood and Shew [75 (link)]. Leaf discs were inoculated with zoospore suspensions as described above. Samples were collected at 24, 48 and 72 hpi and documented with an epifluorescence microscope (ZEISS Axio Scope.A1; Kübler HXP-120C lighting device; Filter set: Zeiss 05; software AxioVision Rel. 4.8). Programmed cell death was studied by trypan blue staining at 24, 28, 32 and 48 hpi as described in Feechan et al. [76 (link)]. For a photographic record of leaf disc tissues a ZEISS Axio Lab.A1 microscope with a Zeiss Axiocam MRc camera and Zen blue software were used.
9
Fluorescent Tagging and Microscopy of Yeast Proteins
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Cells were grown shaking in liquid synthetic complete medium supplemented with 100 µg/ml adenine at 25°, to OD600 = 0.2–0.3, and processed for fluorescence microscopy as described previously (Silva et al. 2012 (link)). Rfa1 was tagged with cyan fluorescent protein (clone W7) (Heim and Tsien 1996 (link)) and Cdc13 with yellow fluorescent protein (clone 10C) (Ormö et al. 1996 (link); Khadaroo et al. 2009 (link)). Fluorophores were visualized with oil immersion on a widefield microscope (AxioImager Z1; Carl Zeiss, Thornwood, NY) equipped with a 100× objective lens (Plan Apochromat, numerical aperture 1.4; Carl Zeiss), a cooled charge-coupled device camera (Orca-ER; Hamamatsu Photonics), DIC, and an illumination source (HXP120C; Carl Zeiss). Eleven optical sections with 0.4 µm spacing through the cell were imaged. Images were acquired and analyzed using Volocity software (PerkinElmer). Images were pseudocolored according to the approximate emission wavelength of the fluorophores.
10
Cell Morphology Assessment via Fluorescence
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To assess cell morphology,
the media were removed from plates containing H9c2 cells, and 100
μL of calcein AM and propidium iodide solution (0.1% v/v each dissolved in phosphate-buffered saline (PBS), with
all reagents from Sigma-Aldrich) was added. After a 10-min incubation
in the dark, cells were imaged using a Zeiss Axiovert 40 fluorescence
microscope (magnification 100×) with an HXP 120 C metal halide
illuminator (Carl Zeiss, Germany).
the media were removed from plates containing H9c2 cells, and 100
μL of calcein AM and propidium iodide solution (0.1% v/v each dissolved in phosphate-buffered saline (PBS), with
all reagents from Sigma-Aldrich) was added. After a 10-min incubation
in the dark, cells were imaged using a Zeiss Axiovert 40 fluorescence
microscope (magnification 100×) with an HXP 120 C metal halide
illuminator (Carl Zeiss, Germany).
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