In vivo analysis of glial cell phenotypes during a viral demyelinating disease in mice

At 1, 2, 3, and 4 WPI, some infected mice as well as their matching controls were injected intraperitoneally 2 h before killing with [methyl-3H]thymidine (6.7 Ci/mmoi; New England Nuclear, Boston, MA), 10 #Ci/g body weight. Subsequent tissue preparation and the immunolabaling was performed as described in the previous sections. After immunolabeling, the sections were dehydrated in 70% ethanol and air dried. They were then dipped in Kodak NTB-2 photographic emulsion diluted 1/2 in distilled water, dried, and stored 1-3 wk at 4°C before being developed. The developing procedure was performed at 14°C: sections were first immersed in Kodak D-19 developer (1:2 in water) for 4 min, rinsed in distilled water for 1 min, treated with Kodak fixer for 5 min, rinsed in distilled water, and coverslipped with tris-glycerol as described above.