Comparison of the behavior of CHO cells during cultivation in 24-square deep well microplates and conventional shake flask systems

Abstract

In biopharmaceutical production, the optimization of cell culture media and supplementation is a vital element of process development. Optimization is usually achieved through the screening of multiple media, feed and feeding strategies. However, most screening is performed in shake flasks, which makes the screening process very time consuming and inefficient. The use of small scale culture systems for the screening process can aid in the ability to screen multiple formulations during process development. In order to assess the suitability of 24 deep well (24DW) plates with the Duetz sandwich-covers as a small scale culture system for process development, we have tested growth and production performance of CHO cells in 24DW plates and conventional shake flask cultures. Multiple studies were performed to assess well-to-well and plate-to-plate variability in 24DW plates. Additional studies were performed to determine the applicability of 24DW plates for cell culture medium and supplement screening in batch and fed batch processes. Cultures in 24DW plates exhibited similar kinetics in growth, viability and protein production to those cultured in shake flasks, suggesting that 24DW plates with Duetz sandwich-covers can be effectively used for high throughput cell culture screening.

Keywords: %CV, percent coefficient of variation; 24DW, 24 deep well; Batch; CD, chemically defined; CHO cells; CHO cells, Chinese Hamster Ovarian cells; Cell culture; Deep well plates; Duetz sandwich-cover system; Fed batch; VCD, viable cell density.

Fig. 1

Comparison of cell growth and mAb production of four CHO lines (CHO line 1–4) in 24DW plate with Duetz sandwich-cover system and conventional shake flask system. Lines represent VCD, while bars represent % of mAb production on Day 10. Each data point represents the mean ± SD of six replicates for plates and duplicates for shake flasks.

Fig. 2

Comparison of viability of four CHO cell lines (CHO line 1–4) in 24DW plate with Duetz sandwich-cover system and conventional shake flask system. Each data point represents the mean ± SD of six replicates for plates and duplicates for shake flasks.

Fig. 3

Multivariate charts showing plate-to-plate variation in a peptone titration study. VCD (upper panel) and mAb production (lower panel) responses of CHO line 5 in TCY peptone titration study. Open and closed circles represent duplicate samples at each TCY peptone concentration and blue squares represent the means of duplicates. The orange solid line connects the titration level means and green diamonds represent overall mean for each plate. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 4

Representative batch culture study in 24DW plate with Duetz sandwich-cover system and conventional shake flask system. A concurrent batch study was performed in 24DW plates and shake flasks. CHO line 4 was exposed to various concentrations of PP3 on Day 0 of culture and samples were collected on various days of culture for assessment of growth and mAb production. Lines represent VCD, while columns represent % mAb production. Each data point represents mean of duplicate samples.

Fig. 5

Representative fed batch culture study in 24DW plate with Duetz sandwich-cover system and conventional shake flask system. A concurrent fed batch study was performed in 24DW plates and shake flasks. CHO line 1 was fed with a CD supplement on day 0, 2, 4 and 6 of culture. Lines represent VCD, while columns represent mAb production. Each data point represents mean of duplicate samples.