Paraffin-embedded tonsil, 4 normal peripheral nerves (sural nerve), 23 NF1-DNFs (13 males, 10 females) and 55 sp-DNFs (31 males, 24 females) were included in the immunohistochemical analyses. All DNFs were incubated with neurofibromin (Santa Cruz sc-67) antibodies, while and all DNFs, tonsil and normal sural nerve sections were incubated with KIR2DL5 (Abcam; ab175895) antibodies. Biotinylated secondary rabbit antibodies (Vector Laboratories) were used in combination with Vectastain Elite ABC development and hematoxylin counterstaining. Normal human tonsil was used as the reference control tissue for the KIR2DL5 antibody. Images were acquired on a Nikon Eclipse E600 microscope conjugated with a Nikon Plan Fluor 10x/0.30 DIC L objective and a Leica EC3 camera. Normal human sural nerve, sp-DNF sections, as well as primary cell cultures of normal human Schwann cells and HEK293T cells were analyzed by immunofluorescence using S100β (Millipore CB1040), c-Kit (Millipore; MAB1164), tryptase (Abcam; ab2378), vimentin (DSHB; AMF-17b) and KIR2DL5 (Abcam; ab129751) antibodies conjugated to appropriate Alexa Fluor secondary antibodies. Images were acquired on a Nikon Eclipse TE300 fluorescence microscope conjugated with a Nikon Plan Fluor 20x/0.45 ELWD objective and a Leica DFC3000G camera.
Anastasaki C., Dahiya S, & Gutmann D.H. (2017). KIR2DL5 mutation and loss underlies sporadic dermal neurofibroma pathogenesis and growth. Oncotarget, 8(29), 47574-47585.
+ Open protocol