Khoury Thaer, Liu Qian, Liu Song
Departments of *Pathology †Biostatistics, Roswell Park Cancer Institute, Buffalo, NY.
Appl Immunohistochem Mol Morphol. 2014 Oct;22(9):688-95. doi: 10.1097/PAI.0000000000000018.
HER2 status is an integral part of breast cancer management. One of the variables that negatively affects HER2 test is delay to formalin fixation (DFF). The purpose of this study is to determine the effect of progressive DFF on HER2-dual color in situ hybridization (Dual-ISH).
Ten palpable invasive breast cancers were included in the study. 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; 1 section was kept in saline and stored overnight (ON) at 4°C. Two tissue microarray blocks were constructed. Then, HER2 was tested with Dual-ISH. 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 percentage of cells that had nuclear bubbling and/or background staining was also calculated.
DFF had no statistically significant effect on HER2/CEP17 ratio, absolute number of HER2, or CEP17 signals. However, 1 case became falsely amplified in ON sample. 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. Nuclear bubbling and/or haze or dust was seen in 4 cases (P=0.009). These changes started to appear at 30 minutes to 1 hour DFF (P=0.025 and 0.003, respectively).
DFF has no statistically significant effect on the HER2/CEP17 ratio tested by Dual-ISH. However, significant artifacts could occur in samples fixed after 1 hour of DFF time.
人表皮生长因子受体2（HER2）状态是乳腺癌管理的一个重要组成部分。对HER2检测产生负面影响的变量之一是延迟至福尔马林固定（DFF）。本研究的目的是确定进行性DFF对HER2双色原位杂交（Dual-ISH）的影响。
本研究纳入10例可触及的浸润性乳腺癌。对于每例病例，将获取的肿瘤分成8份，并在0、10、30分钟、1、2、4和8小时后依次固定；1份切片保存在盐水中，并在4℃下过夜保存。构建了两个组织微阵列块。然后，用Dual-ISH检测HER2。计算每个核心10个细胞中的HER2和着丝粒蛋白17（CEP17）信号以及HER2/CEP17比值。计算信号完全丧失的细胞百分比。还计算了出现核泡和/或背景染色的细胞百分比。
DFF对HER2/CEP17比值、HER2的绝对数量或CEP17信号没有统计学上的显著影响。然而，1例过夜样本出现假扩增。随着DFF时间延长，信号丧失的细胞百分比趋势具有统计学意义（P<0.001），在DFF 1小时后更为明显。4例出现核泡和/或雾状或尘状现象（P=0.009）。这些变化在DFF 30分钟至小时开始出现（分别为P=0.025和0.003）。
DFF对通过Dual-ISH检测的HER2/CEP17比值没有统计学上的显著影响。然而，在DFF 1小时后固定的样本中可能会出现明显的假象。
