Fluorescence in situ hybridization (FISH) is increasingly recognized as the most accurate and predictive test for both HER2 gene amplification or expression and response to Herceptin™ therapy in breast cancer. Diagnostic procedures for FISH require rigorous quality control, as with all diagnostic procedures, which rely on standardized methodologies. We describe the use of FISH for HER2 in our own laboratory, based on more than 4000 diagnostic assays, and the optimal approaches to scoring HER2 gene amplification in breast cancer.