Subset #8 CLL BCR binds to autoantigens, including vimentin, on breast epithelial cells. (A) CLL 657 (S#8,1) IgG binding to breast epithelium. (B) SDS-PAGE electrophoresis of breast tissue protein extracts probed with CLL mAbs, CLL657 (S#8,1), and P326 (S#2,3), or with secondary Ab only (anti-human IgG). The arrows indicate breast tissue proteins specifically recognized by subset #8 CLL657 (S#8,1) mAb. The subset #2 P326 (S#2,3) mAb did not exhibit differential protein recognition relative to control (α-IgG). (C) CLL657 (S#8,1) and P326 (S#2,3) CLL mAbs were used to immunoprecipitate proteins from breast tissue. The immunoprecipitated proteins were resolved by SDS-PAGE electrophoresis, and thereafter the gel was subjected to Coomassie blue staining. The arrows indicate differentially immunoprecipitated proteins between CLL657 (S#8,1) and P326 (S#2,3) mAbs that were subjected to nLC-ESI-MS/MS analysis (supplemental Table 7). (D) IP and Sup samples from breast tissue extracts using CLL657 (S#8,1) and P326 (S#2,3) CLL mAbs were electrophoresed in 10% SDS-PAGE along with lysate of HeLa cells, and subsequently immunoprobed with anti-vimentin and anti–β-actin antibodies. Vimentin was recognized and immunoprecipitated by the subset #8 CLL657 (S#8,1) mAb. IP, immunoprecipitate; Sup, supernatant; Neg CTL, negative control.