Characterization of mAb aggregates in a mammalian cell culture production process

Protein aggregation is a major concern during monoclonal antibody (mAb) production [ 1 , 2 ]. The presence of aggregates can reduce the therapeutic efficacy of mAbs and trigger immunogenic responses upon administration [ 3 ]. Higher molecular weight (HMW) aggregates can be removed during downstream processing (DSP), but prevention of aggregate formation upstream could increase process yield [ 4 , 5 ]. Unfortunately, detection of aggregates upstream is challenging, since the size of aggregates ranges from small oligomers to visible particles and there is no single technique capable of measuring the broad range of aggregation phenomena [ 6 , 7 ]. For upstream detection of aggregates, all HMW species potentially present in the culture broth must be known. Therefore, we established methods to generate different types of aggregates and characterized the different HMW species using size exclusion high pressure liquid chromatography (SE-HPLC), dynamic light scattering (DLS) and UV spectroscopy. Furthermore, stability and traceability of the aggregates in cell culture medium and Chinese hamster ovary (CHO) DG44 supernatant were demonstrated. Finally, the established methods were used to monitor aggregate formation in a mAb producing CHO DG44 cell culture.