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


Materials and Methods

Cell Culture and Bioreactor Operation

The cells used for protein production were Chinese hamster ovary (CHO) cells, and all data presented are from one protein, product 1. All cell culture was performed in custom media and feed developed by Irvine Scientific. Atypical seed train was used for manufacturing runs. The original seed train consisted of WAVE reactors up to the 100-L and 500-L working volume scale for manufacturing. The current seed train in the new facility uses WAVE bioreactors for scale-up and SUBs for production vessels at the 50-L, 250-L, and 1,000-L scale. Development runs were performed in four 2-L Applikon glass vessel reactors with ADI 1030 controllers and two 5-L Sartorius Stedim bioreactors with A+ and B+ controllers. Scaled-up runs were performed in 50-, 250-, and 1,000-L SUBs. The 50-L SUB was integrated with an Applikon I controller, and the 250- and 1,000-L SUBs were integrated with a Finesse TruLogic RDPD controller. The parameters that were logged and optimized included feeding strategy (i.e., schedule, volumes), pH, temperature, DO, and agitation set points, and flow rates. Partial pressure of carbon dioxide (pCO2) levels also were monitored and controlled. Viable cell density and viability were measured using a Cedex automatic cell counter (Innovatis) or a BioProfile Flex (Nova Biomedical). Protein production was determined using a Protein A assay on an Agilent 1100 series high performance liquid chromatography (HPLC) instrument. Some nutrients, metabolites, and electrolytes (L-glutamine, glucose, glutamate, lactate, ammonium, and sodium, calcium, and potassium ions) were monitored offline using the BioProfile Flex.

Multiple scale-up parameters were analyzed to transfer the optimized process from the glass vessels to the SUBs. Initial values for gas flow rates were calculated using vessel volumes per minute. For agitation speed, tip speed, power per volume, and torque, calculations were performed and evaluated. Power per volume was chosen to scale up the agitation speed. After the flow rates and agitation speed had been approximated, oxygen mass transfer coefficient (kLa) studies were performed in the 50-LSUB to determine the efficiency of the calculated values compared with small-scale studies.

Fiber Optic Autoclavable and Disposable Probes

Different configurations of fiber optic technology were evaluated. Autoclavable oxygen probes for in-line measurement from PreSens and Finesse (TruDO Optical) were studied in the 5-L and 2-L glass vessels, respectively. Measurements were taken every minute and compared directly to the Mettler-Toledo (InPro6800) polarographic probes integrated with the controllers. Preliminary studies also were performed in shaker flasks using the PreSens non-invasive oxygen sensors based on the same fiber optic technology. The values from these disposable probes were compared to offline measurements of the BioProfile Flex. Lastly, the disposable option available from Finesse (TruFluor DO) was evaluated in the 50-L SUB and compared directly with autoclavable polarographic DO probes. The TruFluor DO is available in a sleeve that is manufactured with HyClone's bioprocess container (bag) for the SUBs. The sleeve contains the disposable sensor, and inserted into the sleeve is a non-invasive optical reader connected to a transmitter.


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