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Immunohistochemistry of the brainstem Sections were washed (3 5 min) in Tris buffered saline (TBS) containing 0.3% Triton X-100 (TBST buffer), endogenous peroxidases were deactivated by incubating in peroxidase block (hydrogen peroxide) for 10 min with gentle agitation, and sections were rinsed with TBS and incubated with target retrieval solution (Dako, Glostrup, Denmark) for 30 minutes at 80 C and washed again with TBST (3 5 min)