No discussions found
Found 4 matching solutions for this experiment
|This kit uses a green fluorescent probe that reacts with autophagic vacuoles and is detectable by flow cytometry|
|Incubate hepatocytes in Cyto-ID green solution dye in dark for 30 min at 37 degreeC|
|Incase of Low CYTO-ID Green dye staining in all treatments, increase the reagent concentration (500X dilution of the dye is recommended) and the incubation time.
Incase of High CYTO-ID Green staining, it could be that the cell culture medium was depleted of nutrients and so change media 4-8 hours before the experimen
Autophagic flux was assessed by increase in GFP-LC3 puncta or Western blot analysis of LC3 in hepatocytes after treatment with bafilomycin (50 nm, Sigma) for 1 h or transfecting hepatocytes with the GFP-RFP-LC3 plasmid. 24 h after transfection, cells were imaged with a Zeiss LSM 510 laser-scanning confocal microscope using a ×63 oil lens. The numbers of GFP and RFP double-positive (early autophagic vacuoles) and RFP-only (late autophagic vacuoles) puncta were counted for each cell.