Antibody–drug conjugates (ADCs) are promising biopharmaceuticals. They combine the high selectivity and affinity to cancer cells with the toxicity of chemotherapeutics in one molecule. ADCs consist of a monoclonal antibody covalently bound by a linker to a highly potent cytotoxic drug. ADC-surrogates contain a nontoxic payload with similar structure and physiochemical properties as the toxic payload of an ADC. Therefore, they can be used as models to develop suitable purification processes or analytical methods. The ADC-surrogate in this work consists of adalimumab bound to fluorescein 5-isocyanate (FITC).

The purification process of ADCs is complex because of the heterogeneity of the conjugates. One of the main challenges is the separation in different drug–antibody ratios (DARs), which correlate with the potency of the ADC. High DARs are associated with high cytotoxic levels whereas low DARs affect the efficacy of therapeutics. Because of the very hydrophobic payload, a higher DAR results in a greater hydrophobicity of an ADC.

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