The use of human epidermal growth factor receptor type 2 (HER2) gene amplification and overexpression as a molecular predictive marker has become critically important for proper selection of breast cancer patients for treatment with targeted therapeutic agents such as trastuzumab, lapatinib, pertuzumab, and T-DM1. A high level of sensitivity and specificity of molecular tests for this alteration is desirable. The American Society of Clinical Oncology and College of American Pathology have jointly established consensus guidelines to standardize characterization of this alteration in breast cancers. This chapter provides a brief overview of pre-analytic and analytical processing of breast specimens as well as subsequent molecular evaluation for HER2 status.