CellLight™ Golgi-GFP BacMam 2.0 (ThermoFisher, cat. no. C10592) was used to label Golgi in U87MG cells. Golgi-GFP was added to 35-mm2 poly-d -lysine-coated glass-bottom tissue culture dishes containing U87MG cells at a concentration of 1 × 108 CellLight® particles/ml, and after incubating cells overnight at 37°C in a 5% CO2 environment, cells were washed three times with PBS and then imaged using confocal microscopy (LSM800, Zeiss) at excitation and emission wavelengths of 488 and 510 nm, respectively. Using Imaris 9.5 (Oxford Instruments) and ImageJ software, colocalization correlations for FeRhoNox-1 and Golgi-GFP-positive organelles on the merged images were determined using Pearson’s correlation coefficients, Mander’s and Lis ICQ.
Celllight golgi gfp bacmam 2
Manufactured by Thermo Fisher Scientific
CellLight Golgi-GFP BacMam 2.0 is a fluorescent protein-based labeling reagent designed to label the Golgi apparatus in live cells. It uses a baculovirus vector system to deliver the Golgi protein fused to a green fluorescent protein (GFP) marker into cells, enabling visualization of the Golgi structure.
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3 protocols using celllight golgi gfp bacmam 2
1
Golgi Labeling in U87MG Cells
2
Visualizing Golgi Localization via Fluorescence
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Fluorescent images of the cells expressing proteins linked with fluorophores were visualized with confocal fluorescence microscopy, Fluoview 10i (Olympus, Tokyo, Japan). Localization in the Golgi apparatus was visualized with CellLight Golgi-GFP BacMam 2.0 (Thermo Fisher Scientific, Waltham, MA) according to the manufacturer’s instructions.
3
HepG2-NTCP-C4 Cells Dynamics
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HepG2-NTCP-C4 cells were seeded at a density of 1.0 × 104 cells/mL in Nunc Lab-Tek chamber slides (Capitol Scientific, Austin, TX) and incubated with a baculovirus infection system encoding GFP-fused RAB5A, GFP-fused RAB7A, and CellLight Golgi-GFP BacMam 2.0 (Thermo Fisher Scientific) overnight at 37°C in a 5% CO2 atmosphere. The next day, the medium was replaced with fresh medium containing Hoechst 33342 (Thermo Fisher Scientific) for another 30 minutes. The medium was replaced by fresh growth medium containing ReAsH-labeled TC155HBV particles or ReAsH-labeled TCPreS1HBV particles purified by iodixanol density gradient. At the indicated times, the cells were fixed in 4% paraformaldehyde. Images were captured with an HM-1000 super-resolution microscope (Sysmex Corp) and Dragonfly confocal microscope (Andor, Oxford Instruments).
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