Antibody–drug conjugates (ADCs) combine the characteristics of large-molecule biologics and small-molecule drugs. Although a number of assays have been reported to characterize the ADCs, a new and comprehensive bioanalytical strategy to overcome the limitations of the current methods is required for better characterization of ADCs for clinical use. An appropriate bioanalytical assay is essential for characterizing ADCs, which include physicochemical and pharmacokinetic properties due to their complex and heterogeneous structures. The commonly used bioanalytical methods typically implemented to characterize various ADCs are summarized with a focus on the pharmacokinetic studies and determination of drug-to-antibody ratios. The challenges and perspectives of the assays are discussed as well.