Mouse anti s100b

For the analysis of structure and morphology, 5-μl sample aliquots were adsorbed to carbon-coated collodion film supported on 400-mesh copper grids and negatively stained with 1% uranyl acetate. For immunogold labeling, 5-μl sample aliquots were adsorbed to carbon-coated collodion film supported on 400-mesh nickel grids for 5 min. After washing with phosphate-buffered saline (PBS), the grids were blocked in 1% bovine serum albumin (BSA) in PBS for 10 min, and then incubated with each primary antibody: rabbit anti–amyloid-β (1:50; Sigma) or mouse anti-S100B (1:50), diluted in 1% BSA/PBS; for double labeling, grids were sequentially incubated with each primary antibody. Following washing with PBS, grids were then incubated with either anti-rabbit or anti-mouse secondary antibodies conjugated to 10- or 15-nm colloidal gold, respectively, or sequentially incubated with each one, for double labeling. Following washing with PBS, samples were negatively stained with 1% uranyl acetate. The grids were visualized with a JEOL JEM-1400 transmission electron microscope equipped with an Orious Sc1000 digital camera, and exhaustively observed. As a control, we performed the immunegold labeling, incubating only the secondary antibodies, and no colloidal gold was observed.