Goat anti doublecortin dcx

Sections were washed with Na-PB and mounted onto charged slides. Sections stained for BrdU were pretreated with 2 N HCl at 37℃ for 30 min and neutralized with PBS before incubation with primary antibody. Sections were blocked with PBS containing 5% horse serum and 0.4% Triton X-100 in PBS (PBST) for 60 min followed by incubation with primary antibody in the same buffer solution at 10℃ overnight. Primary antibodies were used at the following concentrations: rat anti-BrdU (1:200; Abcam, Cambridge, UK), mouse anti-GFAP (1:200; Chemicon, Temecula, CA, USA), mice anti-neuronal nuclear antigen (NeuN, 1:100; Chemicon), rabbit anti-ionized calcium-binding adapter molecule 1 (Iba-1, 1:100; Wako Chemicals USA, Inc., Richmond, VA, USA), and goat anti-Doublecortin (DCX, 1:100; Cell signaling, Danvers, MA, USA). Sections were washed three times with PBST for 10 min at room temperature and blocked in PBST containing 5% horse serum for 30 min. Samples were then incubated with secondary antibodies conjugated to FITC (green, Jackson Immuno-Research, West Grove, PA, USA) or CY3 (red, Jackson Immuno-Research) at room temperature for two h, washed three times with PBST, stained with 10 mg/ml 4′, 6′-diamidino-2-phenylindole (DAPI) (Sigma) for 10 min, and coverslipped.

Sections were washed in Na-PB and mounted on charged slide glasses for IHC. The sections immunostained for BrdU were pretreated with 2 N HCl for 30 minutes at 37℃ and neutralized with PBS before incubation with primary antibodies. Sections were incubated for 60 minutes with 5% normal horse serum in 0.4% Triton X-100 in PBS (PBST). The sections were incubated overnight at 4℃ with primary antibodies in the same buffer solution. Primary antibodies were used at the following concentrations: rat anti-BrdU (1:200; Abcam, Cambridge, UK), goat anti-doublecortin (DCX, 1:100; Cell Signaling, Danvers, MA), mouse anti-neuronal nuclear antigen (NeuN, 1:100; Chemicon, Temecula, CA), and rabbit anti-ionized calcium-binding adapter molecule 1 (Iba-1, 1:100; Wako Chemicals USA, Inc., Richmond, VA). Sections were washed three times with PBST for 10 minutes at room temperature and blocked in PBST containing 5% horse serum for 30 minutes. Sections were then incubated for 2 hours with secondary antibodies conjugated to FITC (Jackson Immuno-Research, West Grove, PA) or CY3 (Jackson Immuno-Research, West Grove, PA). The sections were washed three times with PBST, and stained with 10 mg/ml 4'6'-diamidino-2-phenylindole (DAPI) (Sigma-Aldrich, St. Louis, USA) for 30 minutes before mounted.