pH is an important and commonly researched parameter that affects the production and quality of monoclonal antibodies (mAbs). Our previous study showed that culture at high pH (7.15 +/- 0.05) yielded a high cell density, at a low pH (6.85 +/- 0.05) yielded a high titer and low byproduct. However, the effects of a pH shift on the production process, quality, and biological potency of mAbs are seldom reported in perfusion culture. In this study, we performed a pH shift perfusion culture (from 7.15 +/- 0.05 to 6.85 +/- 0.05 at day 9) and a control group (constant at 7.15 +/- 0.05) in a 15-L bioreactor and compared the production process, critical attributes and biological efficacy (complement-dependent cytotoxicity, CDC; antibody-dependent cell-mediated cytotoxicity, ADCC) of an anti-CD52 mAb. The cells maintained a higher density and viability, and productivity was significantly enhanced with the pH shift. The charge and size heterogeneity of the mAb were comparable to the control group. However, the glycosylation content, especially that of galactosylation and afucosylation, was significantly higher with the pH shift, which lead to increased biological efficacy. Thus, a pH shift in perfusion culture had a noticeably positive effect on process robustness: it prolonged the culture duration, enhanced productivity, improved product quality and increased biological efficacy.