The TSA signal amplification method was applied in a subset of 44 carcinomas with different staining patterns (16 with cytoplasmic staining in the tumor and membrane staining in adjacent thyroid; 3 with membrane staining in the tumor; 5 negative both in the in tumor and the adjacent thyroid and, finally, 20 with only cytoplasmic staining in tumor and adjacent thyroid). When we compared the slides performed with and without TSA signal amplification, we verified that only the membrane staining remained and appeared more intense with the amplification method. In these cases, the staining involved almost always the same foci of cells that already presented membrane staining (Fig. 3E, F, G and H) i.e. it did not stain additional cells. The intra-cytoplasmic staining vanished both in cancer and in normal tissues. Furthermore, we performed IHC using a homemade antibody for human NIS, pAb 795 against a peptide corresponding to the C-terminal sequence of hNIS pAb 795 (36) in 24 carcinomas (12 cPTC, 4 fvPTC, 2 micro PTC, 2 tall cell PTC, 2 FTC and 2 PDTC). The results were similar to those obtained with clone FP5A (Thermo Scientific/Lab Vision).