NaOCl proved an effective inactivating agent for the GMM cells suspended in protein-containing media. NaOCl concentration,
culture density, and length of contact between the inactivating agent and culture were significant factors in determining
the efficacy of cell inactivation (Figures 3a and 3b). The level of inactivation was proportional to both the concentration
of NaOCl (p-value < 0.0001) and duration of contact between the inactivating agent and culture (p-value = 0.0010). Cell inactivation was inversely proportional to the density of the culture (p-value = 0.0215).
Figure 3. NaOCl Inactivation of Recombinant CHO Cells CHO cells are suspended in protein-containing medium at high (3x107 cells/ml (a)) and low (3x106 cells/ml (b)) cell densities. Following exposure to 0%, 0.25%, and 0.5% (active chlorine) NaOCl,
average cell kill (expressed relative to the VCD at time 0) is graphed against time.
At high densities (approximately 3x107 cells/ml), increasing but incomplete cell inactivation was observed with increasing time and NaOCl concentrations, up to
0.5 percent active chlorine (Figure 3a). At lower densities (3x106 cells/ml), complete cell inactivation was seen within one minute at NaOCl concentrations ≥0.25 percent active chlorine, i.e.,
any residual viable cells were below the level of detection of the system( Figure 3b).
The addition of NaOH to a culture of GMM cells also resulted in cell inactivation. The final concentration of NaOH, the contact
time between the cells and inactivation solution, and the protein content of the media were significant factors in determining
the extent of cell kill (Figures 4a, 4b, 4c, and 4d). Increased cell kill was observed with increasing concentrations of NaOH
(p-value < 0.0001). Recombinant CHO cells suspended in protein-free media appeared more susceptible to inactivation than cells
in protein-containing media (p-value < 0.0001). However, NaOH-induced inactivation was independent of the initial culture density (p-value = 0.1236).
Figure 4. NaOH Inactivation of Recombinant CHO Cells. CHO cells are suspended in protein-containing (a, b) and protein-free
medium (c, d), at high (5x107 cells/ml (a, c)) and low (5x106 cells/ml (b, d)) cell densities. The average cell kill (expressed relative to the VCD at time 0) is graphed against time for
increasing concentrations of NaOH.