Antibody-Drug Conjugates (ADCs) are a rapidly growing class of biopharmaceuticals that combine the targeting specificity of monoclonal antibodies with the cytotoxic potency of small-molecule drugs. Their complex structure necessitates advanced analytical techniques for characterization, particularly in determining the drug-to-antibody ratio (DAR), which is critical for efficacy and safety. This study presents a native reversed-phase liquid chromatography high-resolution mass spectrometry (RPLC-HRMS) method for comprehensive ADC characterization using newly developed RPLC columns. A systematic column selection approach was employed to optimize chromatographic separation and MS detection of intact ADCs, including Polivy and Adcetris. The impact of stationary phase hydrophobicity on retention and selectivity of individual DAR species was evaluated across three columns: Biozen Native RP-1, RP-5, and a prototype-6 phase. Results indicate that RP-5 provides the best balance of retention and resolution, enabling full DAR species separation under MS-compatible conditions with ?25% IPA. This improved separation facilitates accurate average DAR determination and minimizes spectral complexity, leading to enhanced MS response consistency. HRMS analysis confirmed high mass accuracy (<50 ppm error) and the preservation of ADCs in their native state. The optimized method yielded average DAR values in strong agreement with literature and UV detection-based calculations. These findings demonstrate the utility of native RPLC-HRMS as a robust and efficient approach for ADC characterization, with broad applicability across various ADC modalities.