Transfection of rat HER2, ER and PgR genes into 4T1 cells produces a population of TNBC and TPBC. (A) Parental 4T1 cells (106), TNBC (106), or TPBC (106) were placed on ice and stained with anti-ER-FITC (top panels) or anti-PgR-PE (bottom panels). The surface intensity for these antibodies were analyzed by flow cytometry using a FACSort with a Lysys II software program (Becton & Dickinson). Individual cells were gated based on forward (FSC) and orthogonal scatter (SSC). Cell debris was excluded by raising the FSC-height PMT threshold. Histograms are mean relative counts and are a representative experiment of three independently performed experiments with similar results. (B) Parental 4T1 cells (106) (lane 1), TNBC (106) (lane 2), or TPBC (106) (lane 3) were lysed and the amount of ErbB2 (HER2) was determined by Western blot analysis using specific Mab as described in detail in the Materials and Methods section. β-actin was used as control for equal loading. The intensity of the bands were analyzed by densitometry with a video densitometer (Chemilmager™ 5500; Alpha Innotech, San Leandro, CA) using the AAB software (American Applied Biology). Data is a representative experiment from three independently performed experiments with similar results.