In breast cancer amplification of the HER-2/neu oncogene and over-expression of the protein product is associated with poor prognosis, predicts response to some chemotherapeutic regimens and is the target for Herceptin treatment. To date there are several methods to assess the amplification/over-expression of HER-2/neu with each having advantages and disadvantages. We have studied amplification and over-expression of HER-2/neu in 250 consecutive cases of breast cancer (220 invasive and 30 in situ carcinomas) presenting to the Department of Pathology at Women's College Campus of Sunnybrook and Women's College Health Sciences Center. Thirty percent of the invasive carcinomas were node positive. HER-2/neu protein over-expression was assessed by immunohistochemistry (IH) using antibody CB11 and amplification of the gene by differential PCR. The percentage of tumor cells showing CB11 staining was determined and the most significant cut off point for positivity was > or =10% moderate or strong complete membranous staining. The gene was considered amplified if the density score of the product was > or =2. There was 94% concordance between the two methods (P value.0001). Both methods were positive in 16% of cases and negative in 78% of cases. Discrepant cases were examined by FISH which confirmed the IH results in 9/11 invasive carcinomas. These results show that there is excellent concordance between IH and PCR. However, immunohistochemistry is easier to perform and cheaper than PCR and could be used in routine assessment of HER-2/neu in breast cancer patients.