Mouse anti mbp

Slides were washed three times with PBS for 5 minutes before being blocked for 1 hour at room temperature in blocking solution (10% normal goat serum, 1% BSA, 0.25% Triton X-100 in PBS). Slides were then incubated with primary antibodies overnight at 4° C in antibody solution (1% BSA, 0.25% Triton X-100 in PBS). Slides underwent three washes with PBS at room temperature. Secondary labeling with Alexa Fluor antibodies (Invitrogen; 488, 594, 680; 1:1000 dilution) raised in goat was carried out for 1 hour at room temperature. Slides were then washed three times in PBS and incubated in Hoechst 33258 (Invitrogen) for 5 minutes diluted 1:5000 in PBS. Slides were washed three times in PBS and three more times in distilled water, before being cover-slipped using Fluoroshield (Sigma). Slides were dried overnight in the dark prior to imaging.
Primary antibodies used in these experiments included: rat anti-CD11b (BD Biosciences) 1:400, mouse anti-GFAP (Sigma) 1:400, rabbit anti-GFAP (Dako) 1:400, rabbit anti-GFP (Invitrogen) 1:1000, chicken anti-GFP (Invitrogen) 1:750, rabbit anti-RFP (Rockland Immunohistochemicals) 1:750 which recognizes DsRed, mouse anti-MBP (Millipore) 1:500, and rabbit anti-TMEM119 (gift from M. Bennett and B. Barres, Stanford)