Images were obtained using an Olympus Fluoview-Fv1000, Leica SP8X, Zeiss 700, Zeiss LSM 880 with Airyscan and Nikon Ti2-E confocal microscopes. All images were processed with Image J software (NIH) and the montage of figures were performed using Adobe Photoshop CC. Schemes were designed using Adobe Illustrator CS3.
Lsm 880
Manufactured by Nikon
Sourced in Germany
The Nikon LSM 880 is a laser scanning microscope designed for high-resolution imaging and analysis of biological samples. It offers advanced features for confocal microscopy, including a multi-photon excitation capability, enabling researchers to capture detailed images of living cells and tissues.
Automatically generated - may contain errors
Lab products found in correlation
Airyscan, by Nikon (3 mentions)
Ti2 e, by Nikon (1 mentions)
Fluoview fv1000, by Leica camera (1 mentions)
Epifluorescence microscopy, by Nikon (1 mentions)
Anti gfap, by Cell Signaling Technology (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Vector Laboratories (1 mentions)
Propidium iodide dye, by Thermo Fisher Scientific (1 mentions)
Axioplan 2, by Zeiss (1 mentions)
Axiocam hrc, by Zeiss (1 mentions)
Prolong gold antifade, by Thermo Fisher Scientific (1 mentions)
Z fix, by Thermo Fisher Scientific (1 mentions)
N sim, by Nikon (1 mentions)
Mitotracker, by Thermo Fisher Scientific (1 mentions)
Formaldehyde, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (1 mentions)
Plan apo, by Nikon (1 mentions)
Imagej, by Nikon (1 mentions)
12 protocols using lsm 880
1
Multimodal Imaging and Processing Protocol
2
Immunofluorescence Protocol for Astrocyte Markers
3
Fluorescence Microscopy of YFP-Expressing Roots
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Detection of YFP was achieved using a YFP filter (excitation, 488 nm; emission, 505–550 nm) and a Karl Zeiss Lsm880 or a Nikon Eclipse TE-2000 Model-E2 confocal microscope. Roots were mounted in propidium iodide dye (Invitrogen) and the imaging settings were 488 nm excitation and >585 nm emission.
4
Embryo Imaging and Cell Counting
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Images of embryos were taken using an AxioCam HRC camera on a Zeiss Axioplan 2 microscope in DIC mode. High-resolution fluorescence images were taken using a Leica TCS-SP5-MP, a Zeiss LSM880 with AiryScan or a Nikon A1R confocal microscope. EMC number was counted from images taken on compound or confocal microscopes using ImageJ software as follows: (i) the image was loaded into ImageJ; (ii) ‘Cell counter’ was selected from ‘Analyze’ items on the Plugins menu; (iii) ‘Initialize’ was selected; and (iv) cell number was determined. All experimental values are presented as mean ± SEM. Two-tailed Student’s t-test with unequal variance was conducted in Microsoft Excel to compare experimental groups.
5
Immunostaining of Pancreatic Islets
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Isolated islets were washed with PBS and fixed with 4% paraformaldehyde (PFA) for 15 min and permeabilized with 0.3% Triton X-100 in PBS (PBST) for 10 min at room temperature. After incubation with blocking buffer PBS with 10% normal goat serum for 1 h at room temperature, islets were incubated overnight at 4 °C with primary antibodies diluted in PBST. The next day, islets were washed, incubated with secondary antibodies for 1 h at room temperature, and washed again in PBST. DAPI provided nuclear counterstain. Islets were mounted on glass slides with Prolong Gold Anti-fade (Thermo Fisher P36930). Pancreas sections were either flash frozen in optimal cutting temperature (OCT) compound and cut into cryosections or fixed in zinc formalin (Z-Fix, Thermo Fisher NC9351419) or 4% PFA (Electron Microscopy Sciences 15710), dehydrated in 10 to 30% sucrose, and prepared as paraffin or frozen sections. OCT sections were fixed in methanol and stained using the above methods. All islet and OCT sections were imaged using an inverted Zeiss LSM880 fluorescence microscope (Nikon, Ti-E).
6
Visualizing Mitochondrial Dynamics in Cancer Cells
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CRC cells were plated in confocal dishes and then incubated with Mito-tracker (100 nM) (Thermofisher, Waltham, MA, USA) for 30 min at 37 °C. The cells were fixed with 4% paraformaldehyde for 15 min as previously described,34 (link) then washed twice with PBS and permeabilized with 0.2% Triton X-100 in PBS for 15 min at room temperature. Blocking with 5% bovine serum albumin at room temperature for 45 min and incubation with primary antibody at 4 °C overnight. Follow washing for 2 times, the dishes were incubated with Alexa Flour 647 or 488‐labeled secondary antibody, respectively for 1 h. DAPI staining (1 ug/mL,10 min) was performed after two washes and observed on a Zeiss laser scanning confocal microscope (LSM880, Germany) or an ultra-high resolution microscope (Nikon N-SIM).
7
Quantitative Analysis of Neuronal Differentiation
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Micrographs were acquired using a confocal microscope (Zeiss LSM 880) or the Nikon Model Eclipse 800 microscope and quantitative analyses were performed with Image J” (NIH). Cells were counted from twenty randomly selected fields per well for each individual experiment. At least three independent experiments were performed. The percentage of neuronal cell population was calculated against the DAPI-positive total cell number which includes undifferentiated stem cells and differentiated neurons. Cells bearing at least one neurite equal or longer than the soma diameter were considered to be differentiated. Soma size, spines and number of synaptophysin or PSD-95 containing vesicles were measured and counted manually. To differentiate dystrophic and normal neuronal populations, 100 neurons from each experiment were manually selected and analysed. The fluorescence intensity was quantified using ImageJ and displayed in corrected total cell fluorescence (CTCF) = integrated density − (area of selected cell × mean fluorescence of background readings)59 (link).
8
Immunofluorescence Staining Protocol
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Cells were plated on a glass-bottom cell culture Petri dish (NEST,801001) and grew for 24 h followed by fixation using 4% paraformaldehyde (Pierce™ 16% Formaldehyde ((w/v)), Methanol-free, ThermoFisher, 28906) in 1 × PBS for 20 min at room temperature. After washing three times in 1 × PBS, cells were blocked with 4% IgG-free Bovine Serum Albumin for 20 min at room temperature. Permeabilization of cells was performed with 0.2% Triton X-100 (Sigma Aldrich, X100) in 1 × PBS for 10 min, followed by adding primary antibody 1:500 in 1 × PBS and incubating at 37 °C for 2 h. Cells were washed with 1 × PBS three times, followed by incubation with a secondary antibody 1:1000 in 1 × PBS for one hour. After washing three times, the nuclei were stained with 10 µg/ml DAPI (Beyotime). Images were acquired by a confocal microscope (Carl Zeiss, LSM 880 with Airyscan or Nikon A1 R) with a 100× objective. At least three different dishes were quantified per treatment type.
9
Quantitative Confocal Imaging of Cells
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Immunofluorescent images were acquired at 10× and 40× on a Zeiss LSM 880 or Nikon C2 confocal microscope. A minimum of three z-stacks were acquired per animal/group/region of interest with a 2 μm step size. Cell quantification was performed on acquired images using Fiji/ImageJ (NIH) software. To calculate cell densities, the total number of cells counted was divided by the image volume expressed in 106 μm3.
10
Visualizing Lysosomal Dynamics by Immunofluorescence
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Immunofluorescence analysis was performed as described previously [77 (link)], with minor modifications. For visualization of the lysosomes, fixed samples were exposed to a blocking solution and then incubated for 30 min at room temperature with antibodies to LAMP1. The cells were washed three times with PBS and then incubated for 30 min at room temperature with Alexa Fluor 488 or 546–conjugated secondary antibodies. For DQ-BSA assay, cells were preloaded with 10 μg/ml DQ-BSA Red in prewarmed medium for 12 h. They were washed with PBS and fixed with 4% paraformaldehyde. For measurement of lysosomal acidity or activity, cells were loaded with Lysosensor Green DND-189 (1 μM) or Magic Red reagent (Immuno Chemistry Technologies) in a prewarmed medium for 1 h after treatment. Then the cells were washed three times with PBS and replaced with phenol red-free DMEM. Images were captured with a Zeiss LSM 880, Nikon A1R or Leica TCS8 confocal microscope. For live mt-Keima imaging, Hep3B cells stably expressing mt-Keima were imaged at two sequential excitations (458 nm, green; 561 nm, red) and 570–695 nm emission range.
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