The American Society of Clinical Oncology/College of American Pathologist (ASCO/CAP) guidelines on breast cancer currently recommend a cold ischemic time of <1 h, although data are limited. Breast resection specimens were subjected to variable cold ischemic time periods (0.5, 1, 2, 3, 4, 24, and 48 h) within the refrigerator and at room temperature. The study included 25 tumors, all of which had refrigerated samples. Non-refrigerated samples (samples at room temperature) were present on 23 cases. Hormone receptors were semi-quantitatively scored using the H-score method. Human epidermal growth factor receptor 2 gene ERBB2 (HER2) was scored using the ASCO/CAP guidelines. The results were compared with the core biopsy scores, which have negligible cold ischemic time period before fixation. Mild reduction in staining for hormone receptors was judged present if the H-score on the resection specimen was between one-half and three-fourth of the H-score at core biopsy. Significant reduction was judged present when the H-score on resection was less than one-half of the core biopsy H-score. Mild reduction in HER2 staining was judged present if there was one-step reduction; and significant reduction was judged present if there was two-step reduction in staining. A true reduction was judged present only when the reduction was consistently present for the increasing time interval. A focal reduction for a particular time sample was attributed to the heterogeneity of the tumor sample. Non-refrigerated samples are affected more by prolonged cold ischemic time than refrigerated samples. Cold ischemic time period of as short as one-half hour may occasionally impact the immunohistochemical (IHC) staining for progesterone receptor. Significant reduction in IHC staining for hormone receptors, and HER2, however, generally does not result until 4 h for refrigerated samples and 2 h for non-refrigerated samples. The ASCO/CAP guideline of cold ischemic time period of <1 h is a prudent guideline to follow.