Sentinel lymph node (SLN) mapping is an effective and accurate method of sampling the axillary nodal basin for metastatic disease. The SLN is the first node to receive afferent lymphatic drainage from the primary tumor. Lymphatic mapping and SLN biopsy have allowed pathologists to perform a more detailed examination of the SLN(s) and, therefore, provide more accurate staging of the regional lymphatic basin. Recently, more sensitive assays have been developed to increase the detection rate of micrometastatic to the axillary lymph nodes. Cytokeratin (CK) immunohistochemical (IHC) staining of the SLN detects micrometastatic disease, which is frequently missed on routine hematoxylin and eosin (H&E) histology. Therefore, lymphatic mapping combined with CK IHC staining of the SLN provides more accurate staging of the regional lymph nodes in patients with breast cancer. At Moffitt Cancer Center, 478 patients with newly diagnosed breast cancer underwent intraoperative lymphatic mapping using a combination of vital blue dye and technetium-labeled sulfur colloid. The excised SLNs were examined grossly, by intraoperative imprint cytology, by standard H&E histology, and by IHC stains for CK. SLNs that were only CK positive were confirmed malignant by sectioning the block, staining with H&E and finding cells with malignant cytology. Lymphatic mapping and CK IHC staining of the SLNs was successfully performed in 478 newly diagnosed breast cancer patients. Twenty-eight patients had unsuccessful lymphatic mapping for an overall failure rate of 5.5 per cent. A total of 134 (28%) patients had positive nodes (N1) detected. Ninety-three of these patients had both H&E and CK-positive lymph nodes, and an additional 41 patients had only CK-positive SLN(s). A total of 385 patients had H&E-negative SLNs, but only 344 patients had negative SLN(s) defined as both H&E and CK negative. Therefore, 41 (10.6%) of the 385 H&E-negative patients were upstaged, because of the detection of malignant cells by cytokeratin IHC staining of the SLN. Microstaging of SLNs with CK has shifted 10.6 per cent of our patient population from stage I to stage II disease. Undetected micrometastatic disease to the regional lymph nodes may account for the significant proportion of stage I breast cancer treatment failures. Furthermore, the ability to accurately stage the axilla by using lymphatic mapping techniques, SLN biopsy, and more sensitive assays may help identify a subgroup of truly node-negative patients with invasive breast cancer who can avoid the morbidity associated with a complete axillary dissection or systemic chemotherapy. Finally, those patients found to have micrometastatic disease to the regional lymph nodes can be treated appropriately in a more selective fashion.