Fitc conjugated mouse monoclonal anti human cd74 antibody m b741

Melanoma cells were cultured with or without IFN-γ (100 IU/ml) for 48 hours. Cells were subsequently treated with chondroitinase ABC (0.1 U/ml; Sigma-Aldrich) for 4 hours at 37°C, washed with ice-cold phosphate-buffered saline, detached by gentle pipetting, and subsequently fixed with 1% paraformaldehyde in phosphate-buffered saline. Cells were stained with FITC-conjugated mouse monoclonal anti-human CD74 antibody (M-B741) or its isotype control antibody (IgG2a, κ; BD Biosciences, San Jose, CA) and analyzed using a FACSCalibur flow cytometer (BD Biosciences). For immunofluorescence staining, cells were incubated with mouse anti-CD74 antibody (M-B741) or isotype control antibody (BD Biosciences) for 1 hour and then incubated with Alexa Fluor 488-labeled goat anti-mouse IgG antibody (Life Technologies) for 1 hour. Slides were photographed using a confocal fluorescence microscope (FV1000; Olympus).