Data on ER, PR and HER2 was available from previous studies. The status of ER and PR was assessed retrospectively with immunohistochemistry (IHC) using the VENTANA® automated slide stainer (Ventana Medical Systems, Inc.). The primary monoclonal antibodies used were CONFIRM™ mouse anti-ER antibody (clone 6F11) and CONFIRM™ mouse anti-PR antibody (clone 16) (Ventana Medical Systems, Inc.). The cut-off level was set to 10% positively stained tumor cells (9 (link)). HER2 was analyzed with IHC as previously described (10 (link)). The Nottingham Histological Grade (NHG) was analyzed retrospectively by the same investigator for all tumor samples.
The protein expression of RAB6C was analyzed with IHC, and the staining pattern was evaluated independently by two investigators (JS and TB). The polyclonal rabbit antibody ab200396 (Abcam) was used. The intensity of RAB6C in the nucleus was analyzed and scored as 0, 1, 2 or 3. If the nuclei had an intensity ≥2, the tumor was considered to have high expression of RAB6C (RAB6C+). Otherwise, it was considered to have low RAB6C expression (RAB6C-).
The protein expression of RAB6C was analyzed with IHC, and the staining pattern was evaluated independently by two investigators (JS and TB). The polyclonal rabbit antibody ab200396 (Abcam) was used. The intensity of RAB6C in the nucleus was analyzed and scored as 0, 1, 2 or 3. If the nuclei had an intensity ≥2, the tumor was considered to have high expression of RAB6C (RAB6C+). Otherwise, it was considered to have low RAB6C expression (RAB6C-).
