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710 nlo

Manufactured by Zeiss
Sourced in United States, Germany

The Zeiss 710 NLO is a high-performance confocal laser scanning microscope designed for advanced imaging applications. It features a multi-photon excitation system, enabling deep-tissue imaging and live-cell analysis. The Zeiss 710 NLO offers state-of-the-art optical components and a flexible system configuration to meet the diverse needs of researchers and scientists.

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17 protocols using 710 nlo

1

Nanoparticle Uptake and Lymph Node Imaging

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AlexaFluor647-labelled NP were conjugated to 3-Rhod, purified and concentrated, and then 30 μL of the resulting solution, containing 1.8 mg of NPs, was injected i.d. in both forearms of three 6 wk old female C57BL6/J mice. After 24 h the mice were sacrificed and the axillary and brachial LN excised, cleaned under stereoscopic observation, and plated on a glass-bottom 96-well plate for confocal imaging (Zeiss NLO 710). Using a 10x objective lens, 16-bit fluorescence imaging was taken for AlexaFluor647 and rhodamine. Z-stacks approximately 6.4 μm in depth were taken through the depth of the LN (approx. 160 μm). In order to acquire the entire LN, convex hull tiling was performed and the resulting images were stitched together.
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2

Immunofluorescence Staining in HeLa Cells

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For immunofluorescence staining, HeLa cells were fixed and immunofluorescently stained as described (Swiech et al., 2011 (link)). Images of stained cells were obtained using a Zeiss NLO 710 confocal microscope with a 40× oil objective (1,024 × 1,024 pixel resolution) as Z-stacks of the images (averaged twice per line) and next converted to single images with a maximum intensity projection function.
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3

Mitochondrial Lipid Dynamics in Liver Cells

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Crude LD from HepG2 cells, and crude LD and LDM from rat liver were isolated and stained with 1 mM MitoTracker red 579/599 nm, 1 mM BODIPY 493/503 and placed on a 1 mm glass slide and covered with cover glass. Fluorescence was measured, and image analysis was performed using Image J software. LDM and CM were isolated from adult rat liver, stained with TMRE (20 nM), and fluorescence was measured. Fluorescence was quantified using Image J. Post-treatment with or without OA (0.5 mM) PLKO.1 and Mfn2 silenced cells were washed with 1X PBS and fixed in methanol for 20 min at −20 °C. Post-fixation cells were washed twice with 1X PBS and stained with BODIPY (1 µg/mL PBS) for 30 min at room temperature. Remnants of stain were removed by washing once with 1X PBS and mounted with DAPI mounting medium (Abcam). Imaging was performed using a laser scanning confocal microscope (Model: NLO 710, CARL ZEISS). Fluorescence was measured and quantified using Image J software.
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4

Confocal Imaging of CAM Tumor Nodules

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The CAM tissues with the tumor nodules were gently rinsed with cold PBS, placed on a glass slide and mounted with 50% glycerol for confocal analysis. The nodules were scanned on a laser scanning confocal microscope (Carl Zeiss NLO-710) at an excitation 550 nm/emission 570 nm spectrum for CM-Dil labeled cells and 488 nm/509 nm for eGFP cells. The 3D images were taken at different magnifications, and the CAM was also scanned at different range of depths using the Z-stack method. Localized depths having strong fluorescent signals were selected for image analysis by ImageJ software, and parameters were adjusted using the Zen 2010 software.
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5

Cell Viability and Apoptosis Assay

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The cell viability was measured by WST-1 assay according to the manufacturer's protocol (Roche Applied Sciences). 5 × 103 cells were plated on 96-well plates and after overnight incubation treated with the indicated drugs for 72 hours. Cell viability defined as the absorbance in the treatment group compared to the control group was expressed as percent mean change ± SD (n = 4). Apoptosis was assessed using differential staining with acridine orange (green) (2 μg/ml; Sigma-Aldrich) incorporated by all cells and ethidium bromide (red) (2 μg/ml; Promega) incorporated only by apoptotic cells with compromised cell membrane. Quantification of apoptotic cells co-stained with acridine orange and ethidium bromide was performed on the images taken with confocal microscope Zeiss NLO710 as described [26 ]. Four replicate experiments were performed. Cells were counted in 5 independent random view fields.
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6

Nanoparticle Uptake and Lymph Node Imaging

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AlexaFluor647-labelled NP were conjugated to 3-Rhod, purified and concentrated, and then 30 μL of the resulting solution, containing 1.8 mg of NPs, was injected i.d. in both forearms of three 6 wk old female C57BL6/J mice. After 24 h the mice were sacrificed and the axillary and brachial LN excised, cleaned under stereoscopic observation, and plated on a glass-bottom 96-well plate for confocal imaging (Zeiss NLO 710). Using a 10x objective lens, 16-bit fluorescence imaging was taken for AlexaFluor647 and rhodamine. Z-stacks approximately 6.4 μm in depth were taken through the depth of the LN (approx. 160 μm). In order to acquire the entire LN, convex hull tiling was performed and the resulting images were stitched together.
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7

Confocal Imaging of Fluorescent Embryos

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Embryos were anaesthetized and mounted in 1% low-melting agarose on a 35-mm diameter glass-base dish (627870 or 627861 Greiner). Confocal images were obtained using a Leica TCS SP8 confocal microscope (Leica Microsystems) equipped with water immersion 25× (Fluotar VISR, 0.95 NA) objective, water immersion 40× (HC PL APO CS2, 1.1 NA) objective and glycerol immersion 63× (HC PL APO CS2, 1.3 NA) objective or FluoView FV1000/FV1200/FV3000 confocal upright microscope (Olympus) equipped with a water immersion 20× (XLUMPlanFL, 1.0 NA) lens. The 473 nm (for GFP), 559 nm (for mCherry), and 633 nm (for Qdot 655) laser lines in FluoView FV1000/FV1200/FV3000 confocal microscope and the 488 nm (for GFP) and 587 nm (for mCherry) in Leica TCS SP8 confocal microscope were employed, and 488 and 651 nm on the Zeiss NLO710, respectively.
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8

Mitochondria Localization and Immunostaining

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The 250,000 cells were plated in glass coverslips in 6-well plates and allowed to attach for 24h at 37°C (5% CO2). The cells were incubated in 1:5000 concentration of Mitotracker Deep Red (Invitrogen) for 15 minutes, washed with prewarmed media, then fixed and permeabilized in 100% methanol for 5 minutes at 4°C. Next, the cells were blocked with 5% BSA in PBS-TT (0.5% Tween and 0.1% Triton) for 1h and immunostained for mGPDH using monoclonal antibody anti-mGPDH (Abcam, Cambridge, MA) for 2h then secondary antibodies conjugated with Alexa Fluor 488 or Alexa Fluor 569 for 1h (Life Technologies, Thermo Fisher Scientific). DNA was stained with DAPI (Vector Laboratories, Burlingame, CA). Fluorescence images were detected by confocal microscope NLO 710 Zeiss, and collected using Carl Zeiss Zen Software (Zeiss, Germany).
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9

Immunocytochemistry Protocol for Quantifying Mitotic Lox Levels

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Forty-eight hours after transfection with siControl or siLOX, 250,000 cells were plated in glass coverslips in 6-well plates and allowed to attach for 24 hr at 37°C and in a 5% CO2 atmosphere. Cells were then fixed in 4% paraformaldehyde (PFA) for 15 min. After permeabilization with 70% ethanol, cells were blocked with 5% BSA in PBS-TT (0.5% Tween and 0.1% Triton) for 1 hr at RT. The cells were immunostained for LOX using monoclonal antibody anti-LOX (Abcam), anti-tubulin (Cell Signaling Technology or DSHB Univ. Iowa), and anti-pH3(Ser10) (Cell Signaling Technology). Then the cells were incubated with appropriate secondary antibodies conjugated with Alexa Fluor 488 or Alexa Fluor 569 for 1 hr (Life Technologies, Thermo Fisher Scientific). DNA was stained with DAPI (Vector Laboratories, Burlingame, CA). For the immunofluorescence staining of the mitotic fractions the samples were fixed in PFA 4%, washed and reconstituted in PBS before overnight incubation on poly-lysine coated cover slips (BD Biosciences) at 4°C. The samples were then stained with Hoechst 33342 (Sigma Aldrich) to detect DNA and with tubulin and LOX antibodies as described above. Fluorescence images were detected by confocal microscope NLO 710 Zeiss, and images were collected using Carl Zeiss Zen Software (Zeiss, Germany).
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10

Multiscale Characterization of Ligament Microstructure

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Upon completion of biaxial mechanical testing, the tissue samples were imaged using the SHG technique at the unloaded state. We utilized a Zeiss 710 NLO inverted confocal microscope (Carl Zeiss Microscopy, LLC, Thornwood, NY, USA), equipped with a mode-locked Ti:Sapphire Chameleon Ultra laser (Coherent Inc., Santa Clara, CA), a non-descanned detector (NDD), and a Plan-Apochromat 40× oil immersion objective. The laser was set to 800 nm and emission was filtered from 380–430 nm. Samples were kept hydrated with saline solution during imaging to prevent drying artifacts and covered with #1.5 coverslips. Samples were imaged inside the area delimited by the graphite markers, and 2D image slices were collected in the thickness direction from the smooth side of each sample. A 2D slice has 512×512 pixels to 1024×1024 pixels, and for each sample the number of slices was varied to cover the thickness. In total, we obtained 3D SHG images (size from 512×512×N to 1024×1024×N) of 48 tissue samples from different animal subjects, and the corresponding mechanical testing data. Representative SHG images of a GLBP sample are shown in Figure 2, with a total of 18 slices (N=18) through the thickness. It can be seen that geometry variation (e.g. fiber waviness) in each imaging plane is much larger than that in the thickness direction as shown in Figure 2.
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