Scanning electron micrographs of uncoated specimen were obtained using an FEI Nova NanoSEM 650. Crystal growth was followed using an inverted Olympus IX-70 confocal microscope. Crystal orientation was determined using a Bruker D8 Advanced diffractometer equipped with a CuKα1 X-ray source in pole configuration, using a step size 1.5° at 2.5 s (Psi 0–90, Phi 0–360). Substrates were characterized using atomic force microscopy (Bruker dimensions 3100 AFM) in tapping mode (Brucker Tespa; resonance frequency 345–385 kHz, K 20–80 Nm−1) at a scan rate of 1.98 Hz with pixel dimension of 512 × 512. Images of the internal structure of the oriented calcite crystals were obtained using high-resolution TEM imaging of thin sections prepared by FIB milling. Sample preparation was performed using an FEI Nova200 Dual Beam FIB/scanning electron microscopy. The ion beam was operated at 30 kV and at beam currents between 0.1 and 5 nA. Lift-out was performed in situ using a Kleindiek micromanipulator. The samples were then analysed with a FEI Tecnai F20 200 kV field emission gun–TEM fitted with an Oxford Instruments INCA 350 EDX system/80 mm X-Max SDD detector and a Gatan Orius SC600A Charge Coupled Device (CCD) camera.
Ix70 confocal microscope
Manufactured by Olympus
Sourced in Japan
The Olympus IX70 is a confocal microscope designed for advanced imaging applications. It features a high-resolution optical system and specialized optics for confocal imaging. The IX70 is capable of capturing detailed images of samples by scanning them point-by-point and reconstructing the image digitally.
Automatically generated - may contain errors
Lab products found in correlation
Nova nanosem 650, by Thermo Fisher Scientific (3 mentions)
Donkey serum, by Agilent Technologies (2 mentions)
Alexa fluor 488 conjugated phalloidin, by Thermo Fisher Scientific (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions)
Nova 200, by Thermo Fisher Scientific (1 mentions)
Anti th, by Merck Group (1 mentions)
Click it tunel alexa fluor imaging assay, by Thermo Fisher Scientific (1 mentions)
Paraformaldehyde (pfa), by Nacalai Tesque (1 mentions)
Prolong gold antifade reagent with dapi, by Thermo Fisher Scientific (1 mentions)
Alexa fluor conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions)
Fitc labelled phalloidin, by Merck Group (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
Triton x 100, by Merck Group (1 mentions)
Bx50 fluorescence microscope, by Olympus (1 mentions)
Alexa fluor 488 phalloidin, by Olympus (1 mentions)
Fluoview 2, by Olympus (1 mentions)
Alexa fluor 568 goat anti mouse, by Thermo Fisher Scientific (1 mentions)
Mounting medium, by Agilent Technologies (1 mentions)
Anti th antibody, by Merck Group (1 mentions)
Envision system hrp, by Agilent Technologies (1 mentions)
14 protocols using ix70 confocal microscope
1
Characterization of Oriented Calcite Crystals
2
Immunohistochemical Analysis of SN and Striatum
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
At the conclusion of experiments, NCD- and HFD-fed C57BL/6 mice were perfused with a 4% paraformaldehyde solution. Whole brains were removed, then stored in 4% paraformaldehyde for 3 days. After transferring brains to a 30% sucrose solution, coronal sections (25 µm thick) were prepared, washed with PBS, and then blocked by incubating for 1 hour with 3% donkey serum (Dako, Glostrup, Denmark) and 0.3% Triton X-100 in the dark. Thereafter, sections were washed with PBS and incubated overnight at 4℃ with anti-TH (1:500; Millipore, MA, USA) and anti-pJNK (1:500; Invitrogen, Carlsbad, CA, USA) antibodies, diluted in blocking solution. Sections were then washed with PBS and incubated with fluorescently labeled secondary antibody for 1 hour at room temperature. Sections from SN was obtained through -2.60 mm to -3.90 mm relative to bregma [39 (link)]. Sections from striatum was obtained through -1.08 mm to -0.84 mm relative to bregma [40 (link)]. TH and pJNK fluorescence (n=10 slides per condition, n=6 each groups) were visualized using an IX70 confocal microscope (Olympus, Tokyo, Japan).
3
TUNEL Assay for Apoptosis Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell apoptosis was assessed by performing the Click-iT® TUNEL Alexa Fluor® Imaging assay (Invitrogen; Thermo Fisher Scientific, Inc.). Cells were fixed with 4% paraformaldehyde for 30 min at room temperature and then washed with PBS. Subsequently, 0.3% Triton X-100 in PBS was added and incubated for 5 min at room temperature. Cells were stained with DAPI at room temperature for 10 min (Thermo Fisher Scientific, Inc.). A total of 50 µl TUNEL reaction mixture was added for 1 h at 37°C. Cells were sealed with anti-fluorescence quenched sealing solution and FITC-labeled TUNEL-positive cells were visualized in three randomly selected fields of view using an IX70 confocal microscope (Olympus Corporation) at ×20 magnification. To calculate the proportion of apoptotic cells, the number of apoptotic cells and the number of total cells were counted.
4
Colocalization of CD26 and SSTR4 in MESO1 and JMN Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For detection of colocalisation between CD26 and SSTR4 in MESO1 and JMN cells, cells were preincubated in collagen-coated 8-well chamber slide glass cells (Iwaki, Tokyo, Japan), and fixed with 4% paraformaldehyde in PBS (Nakarai Tesque, Inc.). After being washed with ice-cold PBS, cells were blocked with normal goat and rabbit IgG (Santa Cruz Biotechnology Inc.), followed by incubation with Alexa Fluor 488-conjugated anti-CD26 pAb and Alexa Fluor 594-conjugated anti-SSTR4 pAb (each at a concentration of 5 μg ml−1). After being washed with ice-cold PBS, slides were mounted with Prolong Gold antifade reagent with DAPI (Molecular Probes, Eugene, OR, USA). Confocal microscopy was performed with an Olympus IX70 confocal microscope with 60 objective lenses (Olympus), using laser excitation at 496 and 568 nm. The widths of Alexa Fluor 488 and 594 emission channels were set to maximise specificity.
5
Cryostat Immunofluorescence Staining of Nerves
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Nerves were transferred to 0.1 m PBS prior to fixing in 4% paraformaldehyde for 30 min at room temperature. The nerves were then cryoprotected (20–30% sucrose for 5 min), transferred to Tissue-Tek medium (Sigma), and frozen using ethanol and dry ice. 20-μm sections were cut by cryostat and mounted onto microscope slides. The freshly cut sections were then submerged in 0.1 m PBS for 5 min and blocked in 0.1 m PBS containing 10% goat serum and 0.5% Triton-X for 120 min at room temperature prior to overnight exposure to primary antibody at 4 °C in the same solution. Monoclonal GFAP (Molecular Probes, 1:200) and rabbit anti-V-ATPase F (sc-20947, Santa Cruz Biotechnology, 1:100) were used. Slides were then washed (3 × 5 min) and incubated in the appropriate Alexa Fluor-conjugated secondary antibody (Molecular Probes, 1:1000) for 60 min at room temperature. The sections were then washed sequentially in 0.5, 0.1, and 0.05 m PBS containing 10% normal serum. Images were collected using an Olympus IX70 confocal microscope (60× objective). In all cases, controls treated as above but without primary antibody were blank.
6
SEM Analysis of CaCO3 Precipitation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
CaCO3 precipitates at different growth and dissolution stages were characterised by Scanning Electron Microscopy (SEM). Electron micrographs of uncoated specimen were obtained using a FEI Nova NanoSEM 650. Crystal growth and dissolution rates of outgrown individual calcite crystal were obtained using an inverted Olympus IX-70 confocal microscope.
7
Immunofluorescence Imaging of Ad-MVFs and Islets
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
ad-MVFs and Langerhans islets in 2D conditions and 3D collagen hydrogels containing in coculture ad-MVFs and islets were fixed in 3.7% formaldehyde (Sigma) for 10 min and permeabilized with 0.1% Triton X-100 (Sigma) for 5 min. The target proteins were recognized by the following monoclonal primary antibodies: Rabbit CD31 (1:500) (Abcam, Cambridge, UK), Mouse CD90 (1:300) (Abcam), and Mouse αSMA FITC conjugated (Sigma). In the same cases, αActin was detected using FITC-labelled phalloidin (1:300) (Sigma). The cell nuclei were stained by DAPI (1:20.000) (Sigma). Cells in 2D and 3D conditions were respectively observed under an OLYMPUS BX50 Fluorescence microscope and OLYMPUS IX70 confocal microscope.
8
SEM Analysis of CaCO3 Precipitation
9
Visualizing Actin Cytoskeleton Dynamics via Microscopy
10
Quantifying Synapses and Neurodegeneration
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Preparations were visualized using an Olympus IX70 confocal microscope. For body wall preparations, 1μm optical sections were taken. Stacks from hemisegments were used to quantify the number of Bruchpilot-containing active zones, with a macro-function developed in ImageJ that automatically selects the HRP-labeled motor terminal in each section and counts individual Bruchpilot-positive punctae while avoiding repeatedly counting the same synapse in contiguous sections. Number of boutons (ellipsoid enlargements of the HRP-labeled axon terminal) and branches (defined as number of HRP-labeled axonal branch tips per hemisegment) was manually quantified based on the anti-HRP signal using the ImageJ point picker tool.
Autofluorescence from whole brains was recorded with an argon 488 nm laser in 2μm optical sections. To quantify brain neurodegeneration, a macro function was developed in Fiji-win32 to automatically detect black circular areas (holes of neurodegenerated tissue) in each autofluorescent section of the brain and measure the area of neurodegeneration relative to the total brain area.
Autofluorescence from whole brains was recorded with an argon 488 nm laser in 2μm optical sections. To quantify brain neurodegeneration, a macro function was developed in Fiji-win32 to automatically detect black circular areas (holes of neurodegenerated tissue) in each autofluorescent section of the brain and measure the area of neurodegeneration relative to the total brain area.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!