ESR1 Amplification in Breast Cancer by Optimized RNase FISH: Frequent but Low-Level and Heterogeneous. PLoS ONE 8(12): e84189. Optical Dataset S1

Moelans CB, Holst F, Hellwinkel O, Simon R, van Diest PJ (2013) ESR1 Amplification in Breast Cancer by Optimized RNase FISH: Frequent but Low-Level and Heterogeneous. PLoS ONE 8(12): e84189. doi:10.1371/journal.pone.0084189

Optical Dataset S1 (without RNase) of S1+S2

OD S2: http://dx.doi.org/10.6084/m9.figshare.864525

Combined Description for Optical Dataset S1 and S2:

The optical datasets illustrate the FISH signal appearance by full size ESR1 FISH photographs (not all of them representative) of a total of 35 breast cancer cases (Case#1-35) including 24 tumors with increased ESR1 copy numbers by FISH (15 ESR1 amplified and 9 ESR1 gained, Case#01-#24) and 11 tumors without ESR1 copy number increase (Case#25-#35), selected from a consecutive subset of 90 formalin fixed, paraffin embedded grade 3 breast cancers from the archives of the Department of Pathology in Hamburg and previously used in an ESR1 FISH mapping study [12]. To illustrate the results of RNase A treatment, for all 24 cases with ESR1 copy number increase different pictures (Fig.01-19) with (+RNase A: optical dataset S2; “OD2”) and without RNase A treatment (optical dataset S1; “OD1”) are available.

FISH data are optical patterns visualized by imaging technology and translated by subjective interpretation into numerical information. To support the confirmability of optical FISH “raw data”, the pictures document by three color photographs the ESR1 FISH signal appearance in nuclei with ESR1 copy number increase and nuclei without ESR1 copy number increase on 4µm conventional large section FFPE tissue slides, showing ESR1 signals (green), CEN6 signals (orange) and cell nuclei (blue) at 100x or 63x magnification. Pictures are labeled “CNI” for observed gene “copy number increase” and “NO” for “normal” or “no copy number increase”. Gene loci with additional allelic copies (CNI) are marked exemplarily in some cases (white arrows and edges), partially indicating occurrence of nuclei with “CNI” appearing sporadically distributed in a pattern of mosaic heterogeneity intermingled with nuclei without “CNI”.

Due to intensity variations, nucleus truncation during slide sectioning and the three dimensional distribution of signals, individual gene copies may not always become visible. To illustrate the distribution of signals within the z-axis, some pictures are taken with different z-layers (Z-Stack A-D).

To illustrate the difference between cases or tissue areas showing ESR1 copy number increase (CNI) and cases or tissue areas without ESR1 copy number increase, pictures of the 11 cases and other tissue areas with clearly normal copy number or “no copy number increase” (NO) are shown in addition.

12. Holst F, Stahl PR, Ruiz C, Hellwinkel O, Jehan Z, et al. (2007) Estrogen receptor alpha (ESR1) gene amplification is frequent in breast cancer. Nat Genet 39: 655-660.