Images of adipocytes expressing the glucose sensor were taken using a Zeiss LSM 780 confocal microscope with laser lines at 458nm for CFP excitation and 514nm for YFP excitation. A 20x plan-apochromat NA = 0.8 objective lens was used to generate a 571 × 571μm field of view at 210 nm/pixel. Pixel intensity was digitised into 12 bits with a scan speed taking ∼5 min per image. Laser power and gain were set to maximise signal without saturation and avoid photobleaching. Brightfield images were recorded from the transmitted light from the 514nm channel. All settings were kept constant across all experiments. Images were acquired using Zeiss ZEN10 software.
Zen10
Manufactured by Zeiss
Sourced in United States, Germany
Zen10 software is a powerful imaging and analysis platform from Zeiss. It provides a comprehensive suite of tools for acquiring, processing, and visualizing microscopy data. The software's core function is to enable users to capture high-quality images, perform image analysis, and generate informative reports.
Automatically generated - may contain errors
Lab products found in correlation
Axio microscope, by Zeiss (2 mentions)
Poly l lysine (pll), by Merck Group (2 mentions)
Fm1 43, by Thermo Fisher Scientific (2 mentions)
Lsm 780, by Zeiss (1 mentions)
Alexa fluor 488 goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
Lsm 710, by Zeiss (1 mentions)
Anti iba1, by Fujifilm (1 mentions)
Alexa fluor 546 goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Fluoroshield medium with dapi, by Merck Group (1 mentions)
Ab10062, by Abcam (1 mentions)
Donkey serum, by Merck Group (1 mentions)
Fluoromount g, by Vector Laboratories (1 mentions)
5 protocols using zen10
1
Imaging Adipocyte Glucose Sensing
2
Immunostaining of Microglia and Astrocytes
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Cells were fixed with 4% paraformaldehyde (PFA) for 10 min at 37°C, blocked and permeabilized with 5% BSA and PBS 0.1% Triton-X100 for 60 min at room temperature (RT), were washed three times and incubated O/N with primary antibody anti-Iba1 (1:500, rabbit polyclonal antibody; Wako Pure Chemical Industries, Osaka, Japan) and anti-GFAP (1:200, mouse monoclonal antibody, ab 10,062, Abcam) in 5% BSA 0.1% Triton-X100 PBS. Slides were washed three times and incubated with the secondary fluorescent antibodies, Alexa Fluor 488 goat anti-rabbit IgG (Invitrogen) and Alexa Fluor 546 goat anti-mouse IgG (Invitrogen), both diluted 1:2000 in 5% BSA 0.1% Triton-X100 PBS, for 2 h at room temperature (RT) in the dark. The slides were mounted using Fluoroshield medium with DAPI (Sigma). Labelled cells were analysed with fluorescence microscope Zeiss LSM710. The microscope pictures were processed with Zen10 software (Zeiss).
3
Comprehensive Immunohistochemistry Analysis of Pancreatic Tissues in Diabetic Mice
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Pancreatic tissues from all mice at day 30 post diabetes induction were harvested, formalin-fixed, and sliced in 5 μm sections while BMSC for immunocytochemistry were fixed in 10% formalin overnight. For histology, tissue sections were deparaffinized with grading xylene and ethanol grades and rehydrated in water. Both tissues or cells were permeabilized and blocked with 4% donkey serum (Sigma Aldrich, USA) for 1 h at RT, followed by primary antibodies (see Table 1 for details) incubation overnight at 4 °C. The next day, cells were washed and labeled with secondary antibodies (see Table 2 for details) for 30 min at RT. Nuclei were marked with DAPI and mounted with Fluoromount-G (VECTASHIELD, USA). Images were captured on LSM710 confocal microscope and analyzed using Zen10 software (Carl Zeiss, USA).
List of secondary antibodies
| Sr. no. | Antibody | Company and catalog no. | Isotype IgG | Mono/polyclonal Ab | Application | Dilution |
|---|---|---|---|---|---|---|
| 1 | Anti-Mouse-IgG-HRP | Jackson ImmunoResearch #115-035-003 | Goat | Poly | Western | 1:5000 |
| 2 | Anti-Rabbit-IgG-HRP | Jackson Immuno Research #111-035-003 | Goat | Poly | Western | 1:5000 |
| 3 | Anti-Mouse-IgG-FITC | Sigma#F8771 | Goat | Poly | IF | 1:200 |
| 4 | Anti-Rabbit-IgG-FITC | Sigma#F9887 | Goat | Poly | IF | 1:200 |
| 5 | Anti-Mouse-IgG-CF555 | Sigma#SAB4600299 | Goat | Poly | IF | 1:100 |
| 6 | Anti-Rabbit-IgG-CF555 | Sigma#SAB4600068 | Goat | Poly | IF | 1:100 |
4
Microscopic Visualization of Bacterial Biofilm
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A bacterial biofilm colony of LGG grown as described above was suspended in 200 μL 1× phosphate-buffered saline (PBS), and dispersed by pipetting. Samples were centrifuged briefly, pelleted, and re-suspended in 5 μL of 1× PBS supplemented with the membrane stain FM1-43 (Molecular Probes, Eugene, OR, USA) at 1 μg/mL. The cells were then placed on a microscope slide and covered with a poly-L-Lysine (Sigma) treated coverslip. The cells were observed by Axio microscope (Zeiss, Goettingen, Germany). Images were analyzed by Zen-10 software (Zeiss, Goettingen, Germany).
5
Bacterial Colony Membrane Staining Protocol
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A bacterial colony grown as described above was suspended in 200 μl 1x Phosphate-Buffered Saline (PBS), and dispersed by pipetting. Samples were centrifuged briefly, pelleted, and re-suspended in 5 μl of 1x PBS supplemented with the membrane stain FM1-43 (Molecular Probes, Eugene, OR, United States) at 1 μg/ml. These cells were placed on a microscope slide and covered with a poly-L-Lysine (Sigma) treated coverslip. The cells were observed by Axio microscope (Zeiss, Germany). Images were analyzed by Zen-10 software (Zeiss).
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