Qualitative images were acquired with a confocal laser‐scanning microscope (Zeiss LSM 710). For quantitative image analysis, the Operetta CLS High‐Content Analysis System (Perkin Elmer) was used to automatically acquire 25 planes per organoid section, with a spacing between planes of 1 μm. Images were modified with the ZEN blue Software. For live imaging (supporting videos), the Cell Observer SD and the CSU‐X1 Spinning Disc Unit (ZEISS) were used. One hundred frames were acquired in each video using the ZEN blue software, during 3054.06 s (163 line), 3053.27 s (EPI line), and 3053.66 s (K7 line). The videos were processed and modified with Adobe Premiere and Screenpresso softwares in order to obtain a representative time‐line of the phagocytosis process. The shown videos represent 29.97 frames per second, resulting in a total number of 659.34 frames (163 line), 689.31 frames (EPI line), and 449.55 frames (K7 line).
Cell observer sd
Manufactured by Zeiss
Sourced in Germany, Poland
The Cell Observer SD is a high-performance microscope system designed for live-cell imaging and time-lapse experiments. It features a fast, sensitive camera and advanced illumination technology to capture high-quality images of dynamic cellular processes. The system is capable of acquiring images at high speed and resolution, making it suitable for a wide range of applications in cell biology research.
Automatically generated - may contain errors
Lab products found in correlation
Lsm 710, by Zeiss (7 mentions)
Axio observer microscope platform, by Zeiss (3 mentions)
Syto9, by Thermo Fisher Scientific (2 mentions)
Coverslips, by Zeiss (2 mentions)
Goat anti mouse secondary antibodies conjugated with atto 488, by Abcam (2 mentions)
Prolong gold antifade containing dapi, by Thermo Fisher Scientific (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions)
Ab16667, by Abcam (2 mentions)
Hoechst 33258, by Thermo Fisher Scientific (2 mentions)
Lsm 780, by Zeiss (2 mentions)
Oil objective lens, by Zeiss (2 mentions)
Axiovert 200m, by Zeiss (2 mentions)
Operetta cls high content analysis system, by PerkinElmer (1 mentions)
Zen software, by Zeiss (1 mentions)
Auriga 60, by Zeiss (1 mentions)
Propidium iodide, by Merck Group (1 mentions)
Advia, by Siemens (1 mentions)
Cal 520 am, by AAT Bioquest (1 mentions)
Brainphystm neuronal medium, by STEMCELL (1 mentions)
Plan apochromat 20x 0, by Zeiss (1 mentions)
69 protocols using cell observer sd
1
Quantitative Confocal Imaging of Phagocytosis
2
Cortex Intravital Microscopy Imaging
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Cortex intravital microscopy (IVM) was performed as previously described by Jenne et al. [20 (link)]. Briefly, mice were anaesthetised by intraperitoneal injection with ketamine (100 µg/g bw + xylazine (10 µg/g bw)), placed on a heating pad to control the body temperature and fixed in a custom-made stereotactic frame. The skin was sterilized by 70% ethanol, opened and the periosteum was removed using a scalpel. The skull was opened to create a cranial window using a drill and removed using regular fine forceps (LST). Next a rubber ring (6 mm diameter; conventional hardware store) was carefully installed and fixed with fibrin glue (3M Vetbond) on the skull surrounding the cranial window to stabilize a droplet of prewarmed PBS (PAN Biotech), which was used to moisturize the tissue and to keep the conditions as physiologic as possible. Spinning disc microscopy was performed on an upright spinning disc confocal microscope (CellObserver SD, Zeiss, Germany) equipped with a HXP120c + LSM T-PMT lighting unit and FLUAR water immersion objectives (10x, 63x were used). The body temperature and depth of anaesthesia of the mice were constantly controlled. Images and movies were acquired using the ZEN software (Zeiss, Germany). Quantification of cell velocities was performed using ImageJ 1.53a (Wayne Rasband, National Institute of Health, USA).
3
Evaluating hASCs Attachment and Morphology on Biomaterials
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The attachment of hASCs onto biomaterials surface, as well as the morphology and growth pattern of cultures were determined after 120 h of cells propagation. The microscopic evaluation of hASCs on biomaterials surface was performed using scanning electron microscope (SEM, Auriga 60, Zeiss, Oberkochen, Germany) and confocal microscope Cell Observer SD (Zeiss, Oberkochen, Germany) as described previously [36 (link),37 (link),38 (link)]. To prepare samples for observations the cultures were fixed with 4% paraformaldehyde. The specimens were either dehydrated in graded ethanol series (concentrations ranges from 50% to 100%, every 10%) for SEM analysis or stained with fluorescent dyes for confocal microscope observations. To visualize the nuclei and cytoskeleton the cultures were stained with diamidino-2-phenylindole (DAPI; 1:800) and atto-488-labeled phalloidin (1:800). The staining was performed using well established protocols [8 (link),26 (link),30 (link)]. All reagents used during this protocol derived from Sigma Aldrich, Poznan, Poland.
4
Visualizing Fungal Endosymbiotic Bacteria
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One-week-old fungal cultures (symbiotic R. microsporus ATCC 62417 and aposymbiotic R. microsporus ATCC 62417/S) were used to visualize the presence or absence of endosymbiotic M. rhizoxinica. The fungal hyphae were stained with 5 μM Syto 9 (Invitrogen) for 5 to 10 min. Fluorescent microscopy was carried out using a spinning disc microscope (Axio Observer microscope-platform equipped with Cell Observer SD; Zeiss), and images were captured using Zeiss-Zen software.
5
In Vivo Microscopy of Hepatic Injury
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For IVM of the liver, mice were prepared as previously described (Volmering et al., 2016 (link)). In brief, mice were anesthetized and placed on a heating pad to maintain body temperature. To induce focal injury, a thin platinic wire was heated and shortly pressed on the exposed liver lobe. The liver lobe was exposed, but no focal injury was induced in sham-operated mice. The necrotic tissue was visualized by 40 ng/ml propidium iodide (Sigma-Aldrich), and neutrophils were labeled by i.v. injection of neutrophil-specific Alexa Fluor 488–conjugated Ly-6G antibody (clone 1A8; BioLegend). Immediately after preparation, the exposed liver was visualized with an upright spinning disk confocal microscope (CellObserver SD; ZEISS) equipped with a 5×/0.25 FLUAR objective, and time-lapse Z stacks were recorded for 4 h. The number of adherent neutrophils was determined for different time points per field of view by using FIJI. The serum levels of the transaminases GOT and GPT before and 4 h after focal hepatic necrosis of WT and Skap2−/− mice were determined with a hematology analyzer (ADVIA; Siemens).
6
Imaging Mitotic Aberrations in Glioblastoma Cells
7
Calcium Imaging of Neuronal Differentiation
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For calcium imaging, NPCs were incubated for 56 days in 3 N medium20 (link) for cortical-like differentiation. For further 14 days of differentiation, media were exchanged for BrainPhysTM Neuronal Medium (STEMCELL Technologies Germany GmbH, Köln, Germany) plus supplements (20× NeuroCult SM1 Supplement, 10× N2 Supplement, 100 µg/ml BDNF, 100 µg/ml GDNF, 100 mg/ml cAMP, and 50 mg/ml L-ascorbic acid). For visualization of calcium signals, neurons were stained with 1 µM Cal520® AM (AAT Bioquest®, CA, USA) for 30 min and imaged for 10 min at a rate of 20 frames per second using spinning disc confocal microscopy (Cell Observer SD, Carl Zeiss Microscopy GmbH, Oberkochen, Deutschland equipped with an an iXon DV885 EMCCD camera, Andor Technology, Belfast, UK).
8
Confocal Imaging of GCGR Localization
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Confocal microscopy imaging was performed to monitor the location of GCGR in live cells. The HEK293 suspension cells (1× 105) were seeded in a 35 mm glass bottom dish and transfected with a modified pcDNA6 expression vector, pcDNA6-sfGFP-GCGR and pcDNA6-sfGFP-GCG(1-29)-FLAG-5GSA-GCGR, including Wild-type GCGR and p.Phe320del. We confirmed the expression and location of GCGR by confocal microscopy imaging. (Cell Observer SD, Zeiss, Germany).
9
Visualizing Arabidopsis Plant Structures
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Hypocotyls, leaves with pavement cells, stomata, and trichomes of 5–8 DAG Arabidopsis plants with PLDα1-YFP expression were documented with spinning disk microscope (Cell Observer SD, Carl Zeiss, Germany) equipped with Plan-Apochromat 20x/0.8 (Carl Zeiss, Germany) and Plan-Apochromat 63x/1.40 Oil (Carl Zeiss, Germany) objectives. Cells were imaged with excitation laser 514 nm and with emission filter BP535/30 for YFP. Cotyledons, petioles and guard cells were documented with confocal laser scanning microscope LSM 710 (Carl Zeiss, Germany) equipped with Plan-Apochromat 20x/0.8 (Carl Zeiss, Germany) and alpha Plan-Apochromat 63x/1.46 Oil (Carl Zeiss, Germany) objectives. Plants of 6 DAG were stained with 4 μM FM4-64 (Invitrogen, USA) diluted in half-strength liquid MS medium for 90 min before imaging. Samples were imaged with excitation lasers 514 nm for YFP and 561 nm for mRFP and FM4-64, beam splitters MBS 458/514 for YFP, MBS 458/561 for mRFP and MBS 488/561for FM4-64. Emission spectrum used were 519–550 nm for YFP, 590–610 nm for mRFP and 651–759 nm for FM4-64.
10
Vesicular Motility Analysis in Neurons
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For vesicular motility analysis, images for time‐lapse movies were taken on a Zeiss Cell observer SD spinning‐disk microscope with an air‐cooled Evolve 512 EMCCD camera at 5 Hz for 60 s (RAB11), 2 Hz for 150 s (RAB4, RAB5, mitochondria), or 1 Hz for 300 s (RAB7) with an 63× oil immersion objective (NA = 1.4). During image acquisition, neurons were kept in a climate chamber (37°C, 5% CO2). Unless noted otherwise, kymographs of vesicular movement from at least four dendrite segments per cell and at least six neurons per condition and experiment were generated and manually analyzed using ImageJ software (Multiple Kymograph plugin by J. Rietdorf and A. Seitz).
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