Scale-Up and Comparison Studies Evaluating Disposable Bioreactors and Probes - Process performance was comparable across all scales, and fiber optic sensors appeared interchangeable with conventional

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Scale-Up and Comparison Studies Evaluating Disposable Bioreactors and Probes
Process performance was comparable across all scales, and fiber optic sensors appeared interchangeable with conventional probes.


BioPharm International Supplements


Process Transfer


Figure 3. Normalized protein concentration versus time in culture for 50-L stirred-tank single-use bioractor runs pre- and post-optimization
The final process was transferred to the 50-L SUB integrated with an Applikon I controller. Initially, the culture was run pre-optimization without the temperature shift, pH drift, or the final feed schedule. The optimized process was run and resulted in an increase in both the normalized protein concentration (Figure 3) and the fractional viability over the pre-optimization run (Figure 4).


Figure 4. Fractional viability versus time in culture for the 50-L and 250-L stirred tank single-use bioractor runs pre- and post-optimization
The 250-L SUB was integrated with the Finesse TruLogic RDPD controller. An engineering run was performed to test the SUB and the controller. The engineering run was performed without the temperature shift or final feed schedule. The fractional viability for this run is atypical, and the rapid drop seen in Figure 4 is likely a result of the fact that this was the pilot run at Acceleron. The optimized process was then run with the fractional viability and normalized protein concentration curves seen in Figures 3 and 4. The optimized process maintained higher viabilities and increased the normalized protein concentration. The pH drift was not necessary for the optimized run because the dual sparger option was used. This run was performed in the new manufacturing facility.


Figure 5. Normalized viable cell concentration (VCD, closed symbols) and fractional viability (FV, open symbols) versus time in culture for the 50-L, 250-L, and 1,000-L stirred-tank single-used bioreactors (SUBs) using the optimized process
Finally, the process was transferred to the 1,000-L SUB. The optimized process was compared across the three scales. Figures 5 and 6 show the normalized viable cell density, the cell viability, and the normalized protein concentration. The normalized viable cell density and viability were similar across scales. However, the 50-LSUB shows slower growth, which has been observed consistently over multiple 50-L runs. The cause of this difference is still under investigation, but it has been hypothesized that it is related to the seed train. The 50-L bioreactor is seeded directly from a WAVE bag, whereas the 250- and 1,000-L bioreactors are seeded from other SUBs. For seeding the 50-L SUB, the inoculum cell culture has to remain longer in a less controlled vessel (WAVE) to reach the densities required for seeding compared to the inoculum culture required to seed the 250- and 1,000-L SUBs (seeded from 50- and 250-L SUBs, respectively).


Figure 6. Normalized protein concentration versus time in culture for 50-L, 250-L, and 1,000-L stirred-tank single-use bioreactors (SUBs) using the optimized process
On the other hand, the normalized viable cell density for the 1,000-L bioreactor appears higher than the data for the 50- and 250-L bioreactors. The cell counts for the 1,000-L reactor were performed using the Cedex, whereas cells in the 50- and 250-L SUBs were counted using the BioProfile Flex. The Cedex and the Flex consistently drift apart, and the Cedex tends to overestimate viable cell density toward the end of the culture. The normalized protein concentration is consistent across all scales (Figure 6).

Comparing Traditional and Fiber Optic Probes


Figure 7. Comparison of disposable versus traditional dissolved oxygen (DO) probes. A). PreSens in-line fiber optic dissolved oxygen probe compared to the traditional polarographic probe. B). Finesse TruDO Optical DO probe compared to the traditional polarographic probe. C). Finesse TruFluor DO disposable DO probe compared to the traditional polarographic probe
The PreSens in-line fiber optic DO probe was compared directly with the polarographic DO probe integrated with the 5-L controller. When the data are overlaid, the trends for the two technologies are identical. The data from one run are shown in Figure 7A. This comparability has proven to be reproducible over many runs.

Finesse's TruDO Optical uses technology similar to that of the PreSens probe. However, the trends between the traditional probe and the fiber optic probe do not overlay exactly. As shown in Figure 7B, data from the TruDO Optical show a 5–8% upward shift in the pO2 level compared to the trend captured by the polarographic sensor. Based on discussions with Finesse, this shift is caused by the temperature at which the two different probes were calibrated. The TruDO Optical comes with a factory calibration at room temperature, whereas the polarographic probe is calibrated in-house at 37 C. In addition, the TruDO Optical data appear to be more noisy. This may come from the number of readings (reading are taken every minute) compared with the number of readings taken from the traditional probe (every three minutes). Additional studies must be run to explain or correct for these differences.

Finally, the TruFluor DO was evaluated in the 50-L SUB. Figure 7C shows the pO2 levels over time in culture for both the disposable TruFluor DO and the traditional probe. The trends are identical between the two probes. Further studies are being performed to confirm direct comparability. These preliminary data show that disposable probe options available are interchangeable with the traditional technology.


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