title: Fluorescence dye-based detection of mAb aggregates in CHO culture supernatants.
abstract: Product yields, efficacy, and safety of monoclonal antibodies (mAbs) are reduced by the formation of higher molecular weight aggregates during upstream processing. In-process characterization of mAb aggregate formation is a challenge since there is a lack of a fast detection method to identify mAb aggregates in cell culture. In this work, we present a rapid method to characterize mAb aggregate-containing Chinese hamster ovary (CHO) cell culture supernatants. The fluorescence dyes thioflavin T (ThT) and 4-4-bis-1-phenylamino-8-naphthalene sulfonate (Bis-ANS) enabled the detection of soluble as well as large mAb aggregates. Partial least square (PLS) regression models were used to evaluate the linearity of the dye-based mAb aggregate detection in buffer down to a mAb aggregate concentration of 2.4 μg mL(-1). Furthermore, mAb aggregates were detected in bioprocess medium using Bis-ANS and ThT. Dye binding to aggregates was stable for 60 min, making the method robust and reliable. Finally, the developed method using 10 μmol L(-1) Bis-ANS enabled discrimination between CHO cell culture supernatants containing different levels of mAb aggregates. The method can be adapted for high-throughput screening, e.g., to screen for cell culture conditions influencing mAb product quality, and hence can contribute to the improvement of production processes of biopharmaceuticals in mammalian cell culture.
authors: Institute of Applied Biotechnology (IAB), Biberach University of Applied Sciences, Hubertus-Liebrecht-Strasse 35, 88400, Biberach, Germany, paul@hochschule-bc.de.
authors: Albert Jesuran
authors: AJ
authors: Paul
authors: Karen
authors: K
authors: Schwab
authors: Nina
authors: N
authors: Prokoph
authors: Elena
authors: E
authors: Haas
authors: René
authors: R
authors: Handrick
authors: Friedemann
authors: F
authors: Hesse
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D003470:Culture Media; D005456:Fluorescent Dyes / Q000737:chemistry*
chemical_list: D000911:Antibodies, Monoclonal; D003470:Culture Media; D005456:Fluorescent Dyes