Delay to Formalin Fixation Negatively Affects on HER2 Test by Dual-Color Silver-Enhanced In Situ Hybridization (SISH)
Thaer Khoury, Qian Liu, Song Liu. Roswell Park Cancer Institute, Buffalo, NY
Background: A woman is decided to receive an effective target therapy of anti-HER2 antibody (trastuzumab) based on HER2 test. Therefore, accurate determination of HER2 is essential. One of the variables that negatively affects HER2 test is delay to formalin fixation (DFF). We and others have demonstrated the effect of DFF on HER2-FISH and HER2-IHC. The purpose of this study is to determine the effect of progressive DFF on HER2-SISH.
Design: Ten palpable invasive breast cancers were resected and underwent immediate gross evaluation. For each case, the procured tumor was divided into 8 parts and consecutively fixed after 0, 10, 30 minutes, 1, 2, 4, and 8 hours; one section was kept in saline and stored overnight at 4C°. Two tissue microarray blocks were constructed. Then, HER2 was tested SISH (Ventana). HER2 and cep17 signal and HER2/Cep17 ratio was calculated in 10 cells for each core. The percentage of cells that had complete loss of signal was calculated. The core was considered affected by artifact when nuclear bubbling and/or haze or dust was present in more than 50% of the cells. Statistical analyses Page's L test, Fisher's exact test and Chi square test were used.
Results: All ten cases were invasive ductal carcinomas. One case had HER2 amplification and the rest were not amplified. DFF had no statistically significant effect on HER2/Cep17 ratio, absolute number of HER2, or Cep17 signals. The trend of the percentage of cells that lost signal with time of DFF was statistically significant (p<0.001), more notably after 1 hour of DFF (figure 1). Nuclear bubbling and/or haze or dust was seen in 4 cases (p=0.009, Chi square test). These changes started to appear at 30-minute to 1-hour DFF (p=0.025 and 0.003, Fisher's exact test respectively).
Conclusions: DFF has no effect on the HER2-SISH results. However, the interpretation could be limited due to the artifact caused by DFF. Therefore, the tissue should be fixed within 1 hour of tissue harvesting and should not be stored overnight.
Tuesday, March 5, 2013 1:00 PM
Poster Session IV # 39, Tuesday Afternoon