Clone 2f11

Paraffin embedded, 7 μm thick sections of decalcified specimens were incubated with monoclonal mouse anti-human Neurofilament protein antibodies, which cross react with both human and rat neurofilament proteins (Clone 2F11, culture supernatant, 1:100; DAKO, Germany). An Isotype identical (IgG1) non-specific mouse antibody served as a negative control (eBioscience, Germany). For signal detection, an EnVision+ System-HRP (AEC) kit (Dako, Germany) was applied. Finally, a counterstain with hematoxylin was performed. Three slides per animal were analyzed using light microscopy (at 10x) and image analysis software.

The central pathology review was performed conjointly by 3 neuropathologists (ATE, PV, EL), a pathologist expert in soft tissue tumors (FL), and a pathologist expert in vascular lesions (MW). For the CNS cases, additional immunohistochemical stainings were performed on paraffin-embedded sections including: CD34 (1:40, clone QBEnd10, Dako, Glostrup, Denmark), smooth muscle actin (SMA) (1:1500, clone 1A4, Dako, Glostrup, Denmark), Desmin (1:400, clone D33, Dako, Glostrup, Denmark), h-caldesmon (1:100, clone h-CALD, Santa Cruz Biotechnology, Dallas, USA), PS100 (1:6000, polyclonal, Dako, Glostrup, Denmark), GFAP (1:200, clone 6F2, Dako, Glostrup, Denmark), neurofilament (1:25, clone 2F11, Dako, Glostrup, Denmark), STAT6 (1:200, clone YE361, Abcam, Cambridge, UK), SSTR2a (1:200, clone UMB1, Abcam, Cambridge, UK) and EMA (1:200, clone GM008, Dako, Glostrup, Denmark). All immunohistochemical stainings were performed in an automatic closed immunostainer (Omnis automate). Orcein staining was also performed.