Immunohistochemical Analysis of Glial and Inflammatory Markers

Immunohistochemical staining was performed as described previously [45 (link)]. The sections were incubated overnight with a rabbit/mouse polyclonal antibody against GFAP; SC-33673 (1:300, Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA), IBA-1; NB100-1028, iNOS; NB300-605 (1:300; Novus Biologicals, Inc., Littleton, CO, USA), COX-2; #12282 (1:300; Cell Signaling Technology, Inc., Beverly, MA, USA). To prevent nonspecific staining, a blocking step was included. Sections were incubated at room temperature for 2 h with 5% bovine serum albumin [46 (link)] (in PBS), and incubated for overnight at 4 °C with the primary antibody in blocking solution (5% BSA). Immunohistochemical staining was performed on 8 mice per group (3 sections per each mouse).