Rabbit anti human cd133 monoclonal antibody

To verify the GBM-SC status, immunofluorescence (IF) staining with the stem cell markers, CD133, and nestin was performed after sorting the GBM-SCs. After differentiation into astrocytic and neuronal lineages with DMEM high glucose medium containing 10% fetal bovine serum (Invitrogen), cells were stained with the astrocyte markers, glial fibrillary acidic protein (GFAP), and neuronal microtubule-associated protein 2 (MAP2) using the following steps: GBM-SCs were fixed with 4% paraformaldehyde, followed by washing and permeabilization in 0.4% Triton X-100. After sequential incubations with individual primary antibodies and the corresponding secondary antibodies, GBM-SCs were counter-stained with 4′,6-diamidino-2-phenylindole (DAPI) solution to visualize the nuclei. GBM-SCs from different groups were mounted on glass slides and analyzed for cytomorphological protein expression by fluorescence microscopy after image capture. Individual antibodies used for IF staining were: rabbit anti-human CD133 monoclonal antibody (1:200, Abcam, UK); mouse anti-human nestin monoclonal antibody (1:200, Merck Millipore, Germany); rabbit antihuman GFAP monoclonal antibody (1:5000, Abcam); mouse anti-human MAP2 monoclonal antibody (1:500, Abcam); Alexa fluor 488 goat anti-rabbit IgG (1:200, Molecular Probes, USA); and Alexa fluor 594 goat anti-mouse IgG (1:300, Molecular Probes).