Evaluation and optimization of hepatocyte culture media factors by design of experiments (DoE) methodology

Abstract

Optimization of cell culture media based on statistical experimental design methodology is a widely used approach for improving cultivation conditions. We applied this methodology to refine the composition of an established culture medium for growth of a human hepatoma cell line, C3A. A selection of growth factors and nutrient supplements were systematically screened according to standard design of experiments (DoE) procedures. The results of the screening indicated that the medium additives hepatocyte growth factor, oncostatin M, and fibroblast growth factor 4 significantly influenced the metabolic activities of the C3A cell line. Surface response methodology revealed that the optimum levels for these factors were 30 ng/ml for hepatocyte growth factor and 35 ng/ml for oncostatin M. Additional experiments on primary human hepatocyte cultures showed high variance in metabolic activities between cells from different individuals, making determination of optimal levels of factors more difficult. Still, it was possible to conclude that hepatocyte growth factor, epidermal growth factor, and oncostatin M had decisive effects on the metabolic functions of primary human hepatocytes.

Profiling the Impact of Medium Formulation on Morphology and Functionality of Primary Hepatocytes in vitro

“…The importance of identifying a culture medium which promotes hepatotrophic support, viability and differentiated function in vitro for BALs, is well recognised6203239. The aim of this study was to assess in vitro retention of synthetic, detoxification and metabolic parameters of high-density hepatocytes cultured in fully defined WE, Modified L-15, M199 and RPMI 1640 media formulations; integrate these datasets using the HBAI to assess global functional capacity, with parallel morphological parameters to elucidate hepatotrophic support suitability.…”

Identification of process conditions influencing protein aggregation in Chinese hamster ovary cell culture

“…There are only a few studies focusing on protein aggregation during USP (Fischer et al, 2015;Franco, Daniela, Fabrizio, Ilaria, & Detlev, 1999;Gomez et al, 2012;Jing et al, 2012;Paul et al, 2015;Paul, Schwab, & Hesse, 2014). Statistical Design of experiments (DoE) strategies have often been used for bioprocess optimization of cell culture processes producing mAbs, since DoE enables process characterization of parameter interactions and classification of parameters based on their impact to the bioprocess and product quality (Abu-Absi et al, 2010;Dong et al, 2008;Mandenius & Brundin, 2008;Montgomery, 2013;Moran et al, 2000). Astonishingly, the formation of large aggregate particles in cell culture was not investigated before, although such particles are known to induce nucleation-dependent aggregation processes (Wang, Nema, & Teagarden, 2010).…”

High-Throughput Nano-Biofilm Microarray for Antifungal Drug Discovery

“…We applied statistical analysis to these results using the Design of Experiments software (24) (Minitab Inc., State College, PA) to detect the interaction effects between the factors. The effects of the interactions between the four independent design variables on robust attachment and formation of true biofilms are shown in Fig.…”