CONTEXT.-: Breast cancer with human epidermal growth factor receptor 2 (HER2) fluorescence in situ hybridization (FISH) group 2 (HER2:CEP17 ratio ≥2.0 and HER2 signals <
4.0) and group 4 (HER2:CEP17 ratio <
2.0 and HER2 signals ≥4.0 and <
6.0) patterns represent a small and challenging subset of cases. These special categories were previously considered positive and equivocal, respectively. However, following the 2018 update of the American Society of Clinical Oncology/College of American Pathologists guidelines, these groups were deemed negative unless concurrent immunohistochemistry was 3+. The guidelines indicate repeated HER2 testing on excisional specimens may be appropriate, but retesting is not definitively recommended. OBJECTIVE.-: To determine if group 2 and group 4 cases change after repeated testing on additional specimens. DESIGN.-: A retrospective review of breast biomarker synoptic reports was conducted from 2016 to 2023. Cases identified as HER2 FISH group 2 and group 4 were recorded, along with the results of any repeated testing. RESULTS.-: A total of 5695 cases with HER2 FISH testing were identified: 101 cases of group 2 (1.8%) and 194 of group 4 (3.4%). There were 42 initial breast biopsies from group 2, and 110 from group 4 with repeated testing on excision. Of these, 5 group-2 (11.9%) and 19 group-4 (17.3%) cases became HER2 positive. None demonstrated positive staining (3+) by immunohistochemistry. There was no association with receipt of neoadjuvant therapy or triple-negative status in either group. CONCLUSIONS.-: We recommend that breast biopsies with group 2 and 4 FISH results have HER2 testing repeated on the excisional specimen.