Anti-Beclin 1 antibody [2A4] (ab114071)

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	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.

Protocol tips

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.

Publication protocol

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.

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