Assessment of anti-drug antibodies (ADAs) for neutralizing activity is important for the clinical development of biopharmaceuticals. Two types of neutralizing antibody (NAb) assays (competitive ligand-binding assay [CLBA] and cell-based assay [CBA]) are commonly used to characterize neutralizing activities. To support the clinical development of benralizumab, a humanized, anti-interleukin-5 receptor α, anti-eosinophil monoclonal antibody, we developed and validated a CLBA and a CBA. The CLBA and CBA were compared for sensitivity, drug tolerance, and precision to detect NAbs in serum samples from clinical trials. The CLBA was more sensitive (27.1 and 37.5 ng/mL) than the CBA (1.02 and 1.10 μg/mL) in detecting NAbs to benralizumab for the polyclonal and monoclonal ADA controls, respectively. With the same polyclonal ADA control, the CLBA detected 250 ng/mL of ADA in the presence of 100 ng/mL of benralizumab, whereas the CBA detected 1.25 μg/mL of ADA in the presence of 780 ng/mL of benralizumab. In 195 ADA-positive samples from 5 studies, 63.59% (124/195) and 16.9% (33/195) were positive for NAb as measured by the CLBA and the CBA, respectively. ADA titers were strongly correlated (Pearson's correlation coefficient r = 0.91; n = 195) with CLBA titers. Moreover, the CLBA titer correlated with CBA percentage inhibition in the CBA-positive samples (Spearman's coefficient r = 0.50; n = 33). Our data demonstrated advantages of the CLBA in various aspects and supported the choice of the CLBA as a NAb assay for the phase III trials.