Cell Line Development Knowledge Center

Reagents Help Clones Survive The Expansion Process And Boost Their Peak Viable Cell Density

In this case study we will examine a customer’s evaluation of the ACF InstiSHAKE CHO cell culture supplement. The customer’s objective was to reduce the number of clones lost during static to shaking expansion and improve the performance of their mAb expressing CHO-K1 cell line during fed-batch.

The project started using the VIPS™ instrument to automate the dispense of single cells into 96 well plates. BalanCD medium was supplemented with 5% InstiGRO CHO, in addition to the customer’s routine medium supplements. After 14 days culture, clones were assessed for IgG productivity (mg/L), and the top 96 clones were expanded to 24 well plates. To assess InstiSHAKE CHO’s ability to support clonal expansion, 48 of the 96 clones were expanded with InstiSHAKE CHO, while the remaining 48 were expanded using the customer’s standard protocol.

After 6 days in 24 well plates, clone productivity was further assessed, and the top 48 clones were expanded to 6 well plates (24 with and 24 without InstiSHAKE). Productivity was again assessed, and the top 37 clones were expanded from 6 well to 50mL spin tubes (18 clones from the control and 19 clones from the InstiSHAKE group). Once the clones had been expanded from 50mL spin tubes to shake flasks, clones were evaluated for productivity during a 14-day fed-batch experiment with Dynamis media (Gibco) and 3% Cell Boost 7a/7b feeds (HyClone).

Results

The customer was very pleased to find that the InstiSHAKE CHO clones excelled during the fed batch experiment with both higher viability (%) and higher viable cell density (cells/mL), see Figures 1 and 2.

InstiSHAKE CHO also increased the number of clones which survived expansion from static to shaking culture. 12 out of 19 clones which were expanded to shaking culture with InstiSHAKE CHO successfully transitioned with high viable cell densities. By comparison, of the 12 control clones which survived expansion, 6 suffered with very low viable cell densities (see Figure 3).

Figure 1. Shows the average viability of the 18 clones expanded from the control group and 19 clones expanded from the InstiSHAKE CHO group at each time point. Error bars show the standard deviation between clone viabilities.

Figure 2. Shows the average viable cell density (cells/mL) for the 18 clones expanded from the control group and 19 clones expanded from the InstiSHAKE CHO group. Error bars show the standard deviation between clone viable cell densities.

Figure 3. Shows the number of clones which were successfully expanded from static to shaking culture with and without InstiSHAKE CHO. Of the successfully expanded clones, those which suffered with low viable cell density (<0.7×106cells/mL) are presented.

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