Immunohistochemical (IHC) staining for BRAFV600E and Ki67 were performed in 34 and 32 radiogenic, and in 38 and 36 sporadic MPTCs, respectively, for which the additional tissue sections were available.
IHC staining for BRAFV600E was performed as described before (34 (link), 35 (link)). In brief, a mouse monoclonal anti-BRAF (mutated V600E) antibody (VE1) ab228461(Abcam) at a 1:100 dilution and the Novolink Polymer Detection System (250T) (Leica RE7140-K) were used to detect IHC reaction product. In our hands, the BRAFV600E positivity on IHC was concordant with the presence of the BRAFV600E mutation (36 (link)).
The proliferative activity of tumors was evaluated by IHC using a Ki67 antibody (clone MIB-1; DAKO, Glostrup, Denmark, 1:100 dilution) in a Ventana BenchMark ULTRA instrument. The Ki67 labeling index (Ki67 LI) was determined with the image-analyzing software (CountσCell, Ki67 antigen Semi-Auto Counter, Seiko Tec LTD, Fukuoka, Japan) in a total of approximately 1,000 PTC cells per case (LZ). Image analysis was performed in a blind for the BRAFV600E status manner.
IHC staining for BRAFV600E was performed as described before (34 (link), 35 (link)). In brief, a mouse monoclonal anti-BRAF (mutated V600E) antibody (VE1) ab228461(Abcam) at a 1:100 dilution and the Novolink Polymer Detection System (250T) (Leica RE7140-K) were used to detect IHC reaction product. In our hands, the BRAFV600E positivity on IHC was concordant with the presence of the BRAFV600E mutation (36 (link)).
The proliferative activity of tumors was evaluated by IHC using a Ki67 antibody (clone MIB-1; DAKO, Glostrup, Denmark, 1:100 dilution) in a Ventana BenchMark ULTRA instrument. The Ki67 labeling index (Ki67 LI) was determined with the image-analyzing software (CountσCell, Ki67 antigen Semi-Auto Counter, Seiko Tec LTD, Fukuoka, Japan) in a total of approximately 1,000 PTC cells per case (LZ). Image analysis was performed in a blind for the BRAFV600E status manner.