A 4% paraformaldehyde solution was used for fixation of the cells, 0.5% Triton X-100 for permeabilization, and 3% bovine serum albumin for blocking. Anti-LC3 primary antibody (Cell Signaling Technology, Danvers, MA, USA) was added, the solution was incubated overnight, and the secondary antibody was added and incubated in dark for 30 min. After staining the nucleus with 4’,6-diamidino-2-phenylindole, the sections were sealed with an anti-fluorescence quenching agent and visualized using a Ts2-FC fluorescence microscope (Nikon, Tokyo, Japan).
Ts2 fc fluorescence microscope
Manufactured by Nikon
Sourced in Japan
The Ts2-FC fluorescence microscope is a laboratory instrument designed for the observation and analysis of fluorescently labeled samples. It features a dedicated fluorescence illumination system and optical components tailored for fluorescence imaging. The core function of the Ts2-FC is to enable high-contrast, high-resolution visualization of fluorescent signals within prepared specimens.
Automatically generated - may contain errors
3 protocols using ts2 fc fluorescence microscope
1
Immunofluorescent Staining of LC3 in Cells
2
Glucose Uptake Fluorescence Assay
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Glucose uptake was detected using a fluorescence assay. HepG2 cells (1 × 104 cells/well) were seeded in a 6-well plate and treated as described above. Then, the medium was removed, and 100 μmol/Lof 2-NBDG was added and coincubated in serum-free DMEM medium at 37°C for 30 min. Then, the relative fluorescence images were observed under a Ts2-FC fluorescence microscope (Nikon, Japan).
3
Immunofluorescence Assay of p-AKT and p-mTOR
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HepG2 cells were prepared as described above and fixed with 10% formalin in PBS for 1 h. Then, the fixed cells were washed with PBS for three times, permeabilized in 0.5% Triton X-100, and then blocked with 5% normal goat serum. After incubation with primary p-AKT (ab23875, 1 : 250, Abcam, USA), p-mTOR (ab131538, 1 : 250, Abcam, USA), and secondary antibody IgG H&L (Alexa Fluor® 594) (ab150076, 1 : 250, Abcam, USA), cell nuclei were stained with DAPI (Beyotime Institute of Biotechnology, China). Then, the relative fluorescence images were obtained using a Ts2-FC fluorescence microscope (Nikon, Japan).
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