We evaluated the HER-2/neu status of 129 invasive breast cancer specimens for gene amplification by fluorescence in situ hybridization (FISH) and protein overexpression by immunohistochemical analysis. Each immunohistochemically stained slide was interpreted on a standard microscope independently by 10 pathologists. Separately, each pathologist reviewed the same slide set with the assistance of digital microscopy. A total of 1,258 manual immunohistochemical scores and 1,269 digital microscopy immunohistochemical scores were completed. When the same 10 pathologists scored the same immunohistochemical slides with the assistance of digital microscopy, each reviewer improved concordance with FISH, and overall concordance with immunohistochemical analysis improved significantly, to 93% (P < .001). The interrater kappa was used to compare interobserver agreement in HER-2 immunohistochemical scoring for manual and digital microscopy interpretation. Significant improvement in interobserver agreement (kappa = 0.51 vs 0.86; P < .001) was achieved when HER-2 immunohistochemical analysis was scored with the assistance of the digital microscope. The assistance of digital microscopy improves the accuracy and reliability of HER-2 immunohistochemical analysis. These data suggest that documented discrepancies between HER-2 immunohistochemical analysis and FISH reflect predominantly errors in manual immunohistochemical interpretation as opposed to immunohistochemical reagent limitations.