| Popis: |
Background Assessing HER2 status accurately and reliably is of great importance in optimizing treatment of HER2-positive breast or gastric cancer. In gastric cancer, for which trastuzumab treatment has been recently approved in Japan, immunohistochemistry (IHC) is the standard method for evaluating the HER2 level. In this study, we examined the impact of formalin fixation conditions on HER2 staining using xenografted tumor samples from mouse models. Methods Mice (BALB-nu/nu) were subcutaneously inoculated with two human gastric cancer cell lines, SCH (score 2+) and SNU-16 (score 1+). Xenograft tumor tissues were collected and left at room temperature for 0, 6 or 24 h before being fixed with 10% neutral buffered formalin for 24 h, 5, 7 or 10 days and then embedded in paraffin. To examine the effect of fixing solution on HER2 IHC, we used 10 and 20% neutral-buffered or non-buffered formalin or Ufix (Sakura Finetek Japan). HER2 IHC was carried out according to the Hercep test. We assessed the effect on IHC by examining the atrophy of tumor cells, autolysis of tumor tissues, immunostaining intensity and immunostaining area. Results Continuous staining of HER2 on the cell membrane with moderate intensity was observed in SCH tumors, whereas partly localized staining on the cell membrane with weak immunostaining intensity was seen in SNU-16. Leaving samples for 6 h before fixation at room temperature decreased immunostaining intensity and induced atrophy of peripheral tumor tissues in both SCH and SNU-16 specimens. Leaving the specimens for 24 h before fixation induced autolysis of tumor tissues and reduced the immunostaining intensity. On the other hand, fixation in 10% neutral buffered formalin for more than 5 days diminished the HER2-stained area in SNU-16 samples. Fixing in 20% neutral buffered or in 10 or 20% non-buffered formalin reduced the staining intensity by prolonged fixation. Ufix did not influence the staining area, but atrophied the tumor cells. Conclusion Our results suggest that the timing of fixation, including when it starts and how long it takes, will influence the HER2 staining. Non-buffered or highly concentrated formalin also might affect the IHC result. It is important to optimize the conditions of sample collection and fixation. |
