The indirect (ELISA) assays were used in the present study to assay the levels of pro-inflammatory cytokines in AD mice brain. The lysates obtained from cortex and hippocampal CA1 region in 5XFAD mice following four weeks of anti-GMF antibody injection were used for pro-inflammatory cytokines assay (Gan and Patel, 2013 (link); Ahmed et al., 2016 (link)). Briefly, equal amount of (30 μg) protein samples from cortex and hippocampus were diluted up to 50 μl with coating buffer and incubated overnight at 4 °C in PVC ELISA microplates (Corning). Thereafter, following 3 washes, each well was blocked with 1 % BSA (blocking buffer) for 1 h at room temperature. Then, each well was incubated with respective antibodies against TNF-α anti-rat (1:200, R&D Systems, Minneapolis, MN), IL-1β anti-rabbit (1:250, CST) and IL-6 anti-goat (1:200, R&D Systems, Minneapolis, MN) for 2 h at 37 °C. Thereafter, following three washes each well was incubated with HRP-conjugated secondary antibodies for 1 h at room temperature. Finally, wells were washed three times and color was developed by TMB solution (3, 3, 5, 5–tetramethyl benzidine, Thermo Fisher) for 30 min at room temperature. The reaction was eventually stopped by using sulfuric acid and the optical density was read at 450 nm on a spectrophotometer (Molecular Devices, Sunnyvale, CA). Data were analyzed as percent change compared with the WT mice.