Immunofluorescent double labelling for GFAP and C3 of human postmortem AD tissue (see Supplemental Data Table 4) and PD tissue (see Supplemental Data Table 5) was completed on 16 μm thick snap-frozen sections, fixed with ice-cold 1:1 methanol:acetone for 5 minutes at room temperature. Endogenous protein activity was blocked with 10% serum of species in which secondary antibodies were raised, diluted in 1× PBST for 2 hours at room temperature. Primary antibody incubations were made overnight at 4 °C – for AD, rat anti-mouse GFAP (1:1000, Sigma G3893) and for PD, mouse anti-porcine GFAP (1:1000, Millipore MAB360), and rabbit anti-human C3D (1:1000, DAKO A0063). Detection was achieved with appropriate Alexa fluorescent secondaries for 3 hours at room temperature (diluted 1:5000 in PBST, Invitrogen).
