Fig. 9.

Binding studies of Z_mab25 or protein G with fetal bovine serum and species selective affinity recovery of two monoclonal antibodies. a 10% FBS was flowed over a Z_mab25 (gray solid trace) or protein G C2–C3 fragment (SPG) (black dashed trace) BIAcore biosensor surface. For comparison, a sample containing 10% FBS spiked with mouse IgG₁ mAb3 was also flowed over the Z_mab25 surface (black solid trace). b: SDS-PAGE analysis (reducing conditions) of samples, flow-through (FT), and eluate (E) fractions from affinity chromatography experiments using either a protein G (SPG) column or a Z_mab25 column, and samples of 10% FBS only (lanes 1–6) or 10% FBS with in-spiked mouse IgG₁ mAb3 (lanes 7–12) or 10% FBS with in-spiked mouse IgG₁ 8F11 (lanes 13–15). M LMW-SDS molecular weight marker; lane 1: 10% FBS sample; lane 2: SPG column, FT; lane 3: SPG column, E; lane 4: 10% FBS sample; lane 5: Z_mab25 column, FT; lane 6: Z_mab25 column, E; lane 7: 10% FBS + mAb3 sample; lane 8: SPG column, FT; lane 9: SPG column, E; lane 10: 10% FBS + mAb3 sample; lane 11: Z_mab25 column, FT; lane 12: Z_mab25 column, E; lane 13: 10% FBS + mAb 8F11 sample; lane 14: Z_mab25 column, FT; lane 15: Z_mab25 column, E. The black arrows to the right indicate nominal molecular weights of antibody heavy and light chains, respectively. The numbers to the left indicate molecular weights in kilo Dalton (kDa). c Results from a biosensor analysis of pH neutralized eluates from the SPG and Z_mab25 columns, originating from the use of the 10% FBS sample containing in-spiked mAb3 (1–4) or mAb 8F11 (5, 6). On separate flow cell surfaces, anti-cow IgG and anti-mouse Ig antibodies were immobilized, and samples were injected. (1) black solid trace SPG column eluate, anti-cow IgG surface; (2) black dashed trace SPG column eluate, anti-mouse Ig surface; (3) gray solid trace Z_mab25 column eluate, anti-cow IgG surface; (4) gray dashed trace Z_mab25 column eluate, anti-mouse Ig surface; (5) black dotted trace Z_mab25 column eluate, anti-cow IgG surface; (6) gray dotted trace Z_mab25 column eluate, anti-mouse Ig surface