[2040] Validation of Dual Color HER2/Chromosome 17 ISH (In-Situ Hybridization) – Comparison with Single Color CISH (Chromogenic In-Situ Hybridization) and Inter-Observer Reproducibility

Mary P Branscomb, John R Ross, Debra Horton, Marva S Rasco, Thomas S Winokur, Vishnu V Reddy, Shi Wei, Shuko Harada. University of Alabama at Birmingham, Birmingham, AL

Background: Evaluation of HER2 status is important in breast cancer management. Fluorescent in-situ hybridization (FISH) is considered the standard method, although brightfield in-situ hybridization (ISH) has several advantages including certainty of recognizing tumor cells and stability of the signals. Recently, dual color brightfield ISH has become available. In this study, dual color HER2 ISH testing was compared with HER2 single color CISH testing, currently offering clinically in our institution.
Design: Forty-seven cases with invasive breast carcinoma were selected based on HER2 CISH test (Invitrogen SPOT-LIGHT) results (50 cases were selected originally and 3 cases were excluded due to lack of tumor cells on the section; 22 amplified, 22 non-amplified, and 3 equivocal). Dual color HER2/Chromosome 17 ISH (Ventana INFORM) were performed on these cases. Dual color ISH results were evaluated by four pathologists independently who were blinded to any information and the results were compared with CISH results. For discrepant cases, final calls were made at the QA conference.
Results: All 22 cases called non-amplified by CISH were called non-amplified by Dual color ISH assay and there was no inter-observer variation. Among 22 cases called amplified by CISH, one case was called non-amplified because the areas showing amplification were not present in the section used for Dual color ISH. The remaining 21 cases were called amplified. Two of these cases showed inter-observer variation. Two of three equivocal cases showed inter-observer variation with one called amplified and the other non-amplified. Third case was called non-amplified due to polysomy, which cannot be evaluated by CISH.

Table 1: Concordance between single color CISH and Dual color ISH
 Single Color CISH
Dual Color ISHAmplifiedEquivocalNon-Amplified
* The areas showing amplification in CISH were not present in the section used for Dual color ISH

Overall, concordance rate between two assays was 97.7%. Inter-observer concordance rate was 91.5% with Cohen's kappa 0.907. Four cases showing inter-observer variation appear to have tumor heterogeneity.
Conclusions: In conclusion, Dual-color HER2/Chromosome 17 ISH assay showed an excellent performance with added advantage of ability to distinguish true amplified cases from polysomy without amplification. This assay may be a better alternative to the currently most commonly used FISH assay.
Category: Techniques

Wednesday, March 6, 2013 9:30 AM

Poster Session V # 264, Wednesday Morning


Close Window