Fig. 2

HER2 assessment in cell lines and tissues by using automatic and quantitative IF-FISH analysis. a HER2 overexpression (given by the cell-by-cell ratio between the HER2 and the CK signals) vs. HER2 amplification (given by the cell-by-cell ratio between the number of HER2 loci and CEP17) for HER2 positive (+) and HER2 negative (−) cell lines. Data are plotted as a mean ± standard deviation. b Assessment of HER2 overexpression and amplification in HER2 + and HER2- tissues with the same methodology as in a. The HER2 status is obtained from assessment by a pathologist using a standard FISH technique. The error bars represent the variation of these scores among cells in the tissue. c IF image of a HER2-equivocal tissue. d Cell-by-cell representation of HER2/CK. and HER2/CEP17 signals. Cells are represented by dots. The contours of the dots (in yellow scale) denote the normalized HER2/CK ratio, while the inside of the dots (in red scale) indicate the HER2/CEP17 ratio of the cells in (c). Normalization of the HER2/CK ratio (yellow scale) is obtained by allocating the value 1 to the maximum HER2/CK ratio that was obtained from all tissues. The maximum of the red scale is chosen as 8 for easy distinction between positive and negative cells. e IF image of a HER2-positive tissue. f Cell-by-cell representation of the tissue in (e) with the same methodology as in (d). Colors in IF images: Blue = DAPI, Green = CK, Red = HER2