On the fourth day after transplantation, the zebrafish larvae were fixed with low melting point agarose gel for imaging under a confocal microscope using the 20× water-immersion objective (Fluoview 1000, Olympus, Japan) or the stereomicroscope (MVX10, Olympus, Japan), with spatial resolution of the images being 1,024×1,024 pixels (Fluoview 1000) or 1,600×1,200 (MVX10), respectively.
Fluoview 1000
Manufactured by Olympus
Sourced in Japan, United States, Germany, Canada, Switzerland, Panama
The Fluoview 1000 is a confocal laser scanning microscope system designed for high-resolution fluorescence imaging. It features a multi-line laser unit, advanced optics, and a sensitive detection system to capture detailed images of fluorescently labeled samples.
Automatically generated - may contain errors
Lab products found in correlation
4 6 diamidino 2 phenylindole (dapi), by Merck Group (17 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (13 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (10 mentions)
Fluoview software, by Olympus (9 mentions)
Paraformaldehyde (pfa), by Merck Group (8 mentions)
Triton x 100, by Merck Group (7 mentions)
Deadend fluorometric tunel system g3250 kit, by Promega (6 mentions)
Ix81 confocal microscope, by Olympus (5 mentions)
Hoechst 33342, by Thermo Fisher Scientific (5 mentions)
Alexa fluor 594, by Thermo Fisher Scientific (5 mentions)
Uplansapo, by Olympus (5 mentions)
Image pro plus, by Media Cybernetics (5 mentions)
Fluoview fv1000, by Olympus (5 mentions)
Mitotracker green, by Thermo Fisher Scientific (4 mentions)
Alexa fluor 488 phalloidin, by Thermo Fisher Scientific (4 mentions)
Vectashield, by Vector Laboratories (4 mentions)
Mvx10, by Olympus (4 mentions)
Image it fx signal enhancer, by Thermo Fisher Scientific (4 mentions)
Fluoview 3000, by Olympus (4 mentions)
Dabco, by Merck Group (4 mentions)
658 protocols using fluoview 1000
1
Zebrafish Larvae Imaging Protocol
2
Brain Tissue Immunostaining Protocol
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After perfusion, brain segments were postfixed in 4% paraformaldehyde for 24 h, and then permeated with 20% sucrose in 0.1 M phosphate-buffered saline (PBS) (pH 7.4) for 24 h and 30% sucrose in 0.1 M PBS for 48 h at 4 °C. Brain segments were frozen in an embedding compound (Sakura Finetek, Tokyo, Japan) on dry ice. They were cut with a cryostat (Leica CM 1100; Leica, Wetzlar, Germany) at a thickness of 30 μm and collected in PBS at 4 °C to be processed immunohistochemically as free-floating sections. The sections were incubated for 48 h at 4 °C with primary antibodies: mouse anti-GFAP (1:2000; Cat. # AB5804, RRID: AB_10062746 ), rabbit anti-BDNF (1:2000; Cat. # sc-546, RRID:AB_630940). The sections were washed six times with 0.1 M PBS (10 min each) and then incubated for 3 h at room temperature with the secondary antibody: Alexa488- and Alexa546-conjugated mouse- and rabbit-IgGs.
(Cat. # A-11034, RRID:AB_2576217 and Cat. # A11030, RRID:AB_144695). Immunohistochemical images were obtained using a confocal laser microscope (Fluoview1000; Olympus, Tokyo, Japan) and digital images were captured with Fluoview1000 (Olympus).
(Cat. # A-11034, RRID:AB_2576217 and Cat. # A11030, RRID:AB_144695). Immunohistochemical images were obtained using a confocal laser microscope (Fluoview1000; Olympus, Tokyo, Japan) and digital images were captured with Fluoview1000 (Olympus).
3
Live Cell IRM Imaging Protocol
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IRM of living cells immobilized on glycine-coated coverslips was recorded on a modified Olympus Fluoview 1000 setup equipped with 60x 1.2 NA water immersion objective (UPlanSApo; Olympus). GFP signal was excited with 20mW 488nm laser (Sapphire; Coherent) and recorded on a single-channel PMT detector equipped with a selective dichroic mirror (DM488/543/633; Olympus). For acquisition and system control the Fluoview 1000 software package (Olympus) was used. The images were processed using ImageJ/Fiji software package [50] .
4
Zebrafish Xenograft Imaging Protocol
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Cultured cancer cells were labeled with CM-DiI (Invitrogen, USA) before injection. Cultured cells were rst collected, then washed three times with HBSS. Next, the cells were labeled with CM-DiI at 37℃ for 5 min, following by 15 min at 4 ℃. Lastly, unincorporated dye was removed by rinsing three times with HBSS, then examined the cells by uorescence microscopy. 48-hpf zebra sh larvae were mounted by 1.2% lowmelting gel (Promega, USA), then about 400 CM-DiI labeled cells were injected into the PVS of each larvae under the micro-injector (Picosprizer III, USA). After injection, the xenografts were cultured at 34℃. At 24 h post injection (hpi), the zebra sh larvae with similar size of transplanted cells were picked up for the further analysis, then cultured at 34℃ until the end of experiments.
In Vivo Imaging And Quantitative Analysis At 4 days post injection (dpi), the zebra sh larvae were also mounted by 1.2% low-melting gel for the imaging experiments. Imaging experiments were performed by stereotype microscopy (MVX10, Olympus, Japan) or confocal microscope using 20X water-immersion objective (Fluoview 1000, Olympus, Japan).
The spatial resolution of the images was 1600 × 1200 (MVX10) or 1024 × 1024 pixels (Fluoview 1000).
In Vivo Imaging And Quantitative Analysis At 4 days post injection (dpi), the zebra sh larvae were also mounted by 1.2% low-melting gel for the imaging experiments. Imaging experiments were performed by stereotype microscopy (MVX10, Olympus, Japan) or confocal microscope using 20X water-immersion objective (Fluoview 1000, Olympus, Japan).
The spatial resolution of the images was 1600 × 1200 (MVX10) or 1024 × 1024 pixels (Fluoview 1000).
5
Visualizing S. mutans Biofilm Disruption
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S. mutans biofilms for CLSM (FluoView 1000, OLYM-PUS, Japan) were generated by placing 100 µl aliquots of an overnight culture (diluted to 10 7 CFU/ml) on each of the chambered cover glasses (Nagle Nunc International, Rochester, NY) and allowing the attachment of the bacteria for 30 min at room temperature. BHI (2 ml) supplemented with 1% sucrose was added to each well, and the chambers were incubated for 24 h at 37°C anaerobically. The biofilms were then washed twice with sterile deionized water and treated with chrysophsin-3 at concentrations of 64 µg/ml and 128 µg/ml. Biofilms treated with 0.12% chlorhexidine (CHX) for 1 h served as a positive control, and biofilms left untreated served as a negative control. To examine the effects of chrysophsin-3 against S. mutans biofilms, biofilms were stained in the dark for 15 min using a LIVE/DEAD® BacLightTM Bacterial Viability kit (L13152, Invitrogen Inc. USA). The stained biofilms were observed under a confocal laser scanning microscope (CLSM) (FluoView 1000, Olympus, Japan), and stacks of images were obtained at 60× magnification. The "Image to Stack" and "Reslice" commands of Image J were used to generate Z-axis images. The fluorescence intensity (FI) of the images was analyzed using Image Pro-Plus software 6.0 (34) .
6
Confocal Microscopy of DAPI-Stained Cells
7
Confocal Microscopy Analysis of TRPM4 and Cholesterol in mpkCCDc14 Cells
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Confocal microscopy experiments were performed as previously reported (Wu et al., 2016 (link)). Briefly, after fixation with 4% paraformaldehyde at room temperature for 10 min, the cells were permeabilized with 0.2% Triton X-100 in PBS for 10 min and blocked with 5% BSA/PBS-T for 30 min. Rabbit polyclonal anti-TRPM4 antibody (alomone ACC-044; 1:100 dilution) was added in 1% BSA/TBS-T for overnight at 4°C. The sections were washed in TBS-T and incubated with Alexa Fluor 488 conjugated donkey anti-rabbit IgG (Invitrogen A21206, 1:1000 dilution) and Alexa-594-conjugated cholera toxin B (CTB) (Invitrogen C34777) for 1 h. All slides were imaged using a confocal microscope (Olympus, Fluoview1000, Japan). To detect cholesterol levels in the plasma membrane of mpkCCDc14 cells, the cells were incubated with 5 μg/ml filipin (Sigma, Cat#: F9765) for 30 min. filipin staining was viewed by confocal microscope using DAPI filter. The control fluorescent intensity is used as a calibrator, and relative fluorescent intensity is calculated against this calibrator. All slides were imaged using a confocal microscope (Olympus, Fluoview1000, Japan) and analyzed using Olympus Fluoview FV1000 version 3.1 software. Identical acquisition settings were used for all images. To quantify colocalizations, the image analysis program ImageJ was used. Both Pearson and Manders coefficients were calculated.
8
Live Cell IRM Imaging Protocol
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IRM of living cells immobilized on glycine-coated coverslips was recorded on a modified Olympus Fluoview 1000 setup equipped with 60x 1.2 NA water immersion objective (UPlanSApo; Olympus). GFP signal was excited with 20mW 488nm laser (Sapphire; Coherent) and recorded on a single-channel PMT detector equipped with a selective dichroic mirror (DM488/543/633; Olympus). For acquisition and system control the Fluoview 1000 software package (Olympus) was used. The images were processed using ImageJ/Fiji software package [50] .
9
Visualizing Mitochondria and Protein Localization in Arabidopsis
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To obtain transformed Arabidopsis lines, T2 transgenic plants showing a restored phenotype were genetically analyzed and homozygous transgenic lines were selected for in-depth analysis. Staining of mitochondrial-specific dye was performed as described previously65 (link), and the GFP signal and fluorescence of MitoTracker Red were examined under a confocal microscope at an excitation wavelength of 488 nm and 543 nm, respectively (FluoView 1000, Olympus). To carry out the transient expression analysis of MOD1-GFP, Arabidopsis mesophyll protoplasts were isolated and transformed as described previously66 (link), and GFP signals were detected by a confocal fluorescence microscope (FluoView 1000, Olympus).
10
Clearing and Imaging Thy1-YFP Mouse Brains
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The brain samples from Thy1-YFP (line H) mice were placed in a hand-made chamber filled with clearing agents. For SeeDB-treated samples, 130% (wt/vol) fructose was not used for immersion due to the fact that its extreme viscosity would cause an uneven RI distribution after evaporation of water during imaging and might impair image quality. Although previously suggested as an immersion agent (Ke et al., 2013 (link)), 2, 2′-thiodiethanol was also not used, due to lack of availability. As a result, 100% FRUIT (wt/vol, RI = 1.48) or 115% FRUIT (wt/vol, RI = 1.50) was used for immersion for FRUIT (35:100)- or SeeDB-cleared samples. An upright multiphoton microscope (FV10MP-BXD4CH, Olympus) was used for two-photon imaging with 920 nm excitation. Images at a resolution of 512 × 512 pixels were collected with a 25 × objective (Olympus, NA = 1.0, working distance = 4.0 mm). Image analysis and presentation were performed with FluoView 1000 software (Olympus).
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