DNA-dependent protein kinase (DNA-PK) has an important role in DNA double-strand break repair. We previously demonstrated the association of DNA-PK activity in peripheral blood lymphocytes (PBL) with incidence of chromosomal aberrations and risk of cancer. In this study, we examined the expression of Ku70 and Ku86 in breast cancer tissue and normal breast tissue with immunohistochemistry. We also measured the DNA-PK activity in PBL of the same patient. One hundred and ten breast cancer patients were included in this study. The expression of Ku70, and Ku86 in normal mammary epithelial cells and breast cancer cells obtained from surgical specimens was immunohistochemically examined. DNA-PK activity of PBL was measured by DNA-pull-down assay. The expression of Ku70 and that of Ku86 tended to parallel each other in normal and cancer tissues. There was also a relationship in the expression of Ku70 and Ku86 between cancer tissues and normal tissues in the same samples. Lower expression of Ku70 or Ku86 tended to be associated with higher malignant nuclear grade of cancer cells and higher frequency of axillary lymph node metastasis. The staining score of Ku70 or Ku86 of normal mammary epithelial cells or breast cancer cells had no significant relationship with DNA-PK activity of PBL. In conclusion, breast cancer cells inherited the characteristics of expression of Ku proteins from original mammary epithelial cells. The staining score of Ku70 or Ku86 of normal or cancer cells had no significant relationship with DNA-PK activity of PBL. This may be due to limitations in the assay sensitivity of immunohistochemistry.