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26 | MAY 1, 2015 | GENengnews.com | Genetic Engineering & Biotechnology News be sure in fed-batch processes to have effcient energy metabolism via low substrate levels, but also have suffcient amounts of those sub- strates to ensure quality," Dr. Butler explains. If nutrient levels and critical parameters such as dissolved oxygen are properly optimized, it is possible to assure consistent glycosylation. The $64,000 question: Is this approach applicable to large-scale cell cultures? Dr. Butler believes so. Results at the 5 L lab scale, which he employs, are generally conserved to at least 100 L provided con- trols are maintained over pH, temperature, and oxygen transfer. Dr. Butler has used batch, fed-batch, and chemostat cultures to test his idea. Although not widely used at manufacturing scale, chemostat cultures al- low adjustment of one process variable at a time to isolate conditions affecting critical quality attributes. Overcoming Limitations Achieving high titers has become manda- tory for antibody manufacturing, yet com- panies relying on fed-batch processes still encounter production limitations. Conven- tional fed-batch culture utilizes large feed additions resulting in diluted fnal product, or incorporation of concentrated feeds that requires the addition of acidic and basic sub- groups, leading to diffcult-to-manage pro- duction at large scale. Thermo Fisher Scientifc has introduced a single-part dry-format feed media for CHO bioprocesses that hydrates in reduced liquid volumes without the need for pH adjust- ment. The product, EffcientFeed™, is en- abled by proprietary technology that allows delivery of higher concentrations of diffcult- to-solubilize components. "Our approach addresses several challeng- es associated with large-scale fed-batch manu- facturing," says Rebecca Moore, Ph.D., senior scientist for bioproduction. These challenges include bioreactor working volume restric- tions, safety concerns with large volumes of high-pH solutions, lengthy fuid transfer times, and limited storage space. "It also diminishes complications accompa- nying pH-adjusted solutions," adds Dr. Moore. The complications include complicated prepa- ration requirements, additional pH control procedures, excess osmolality from salt formed through pH adjustments, supply chain pressures from use of short-shelf-life materials, and the in- ability to combine feed solutions without com- ponent precipitation. Dr. Moore cites this feed product as an example of process intensifcation by virtue of a two- to three-fold reduction in feed vol- ume, and the potential for a substantial in- crease in production capacity resulting from the delivery of proportionately more nutri- tional content with increased volumes of super-concentrated feeds. "Eliminating pH adjustment steps also meets the defnition of intensifcation," Dr. Moore explains. These materials can achieve stable concen- trations of 150–200 g/L at neutral pH after straightforward reconstitution in water, result- ing in solutions without excess osmolality as- sociated with pH adjustment. The simplicity of reconstitution reduces the risk of errors. Treading Carefully with Single-Use Single-use technology has enhanced pro- tein-generating productivity by eliminating non-value-added activities such as cleaning and cleaning validation. While disposables are widely used for many mammalian cell lines, adoption in bacterial systems is far less com- mon due to engineering issues related to agita- tion and gas exchange. For both microbial and insect culture systems, the lack of a large body of knowledge gleaned from years of experi- ence with mammalian cell culture is telling. Biomanufacturing with insect cells faces the same challenges, and raises the same questions that were common among mam- malian culture specialists a decade ago. That insect cell cultures are already quite innova- tive tends to compound issues related to lack of experience. That is beginning to change. Protein Sci- ences has been quite successful with its Flu- Blok infuenza vaccine. Unlike traditional vaccines produced through the familiar, de- cades-old egg-based process, FluBlok employs expresSF+ ® insect cells to express (through baculovirus infection) an immunogenic virus surface protein, hemagglutinin, from antici- pated virus strains. The company maintains a 500 L culture facility at its headquarters in Meriden, CT, and a 2,500 L facility in Pearl River, NY. This process can either be used for seasonal fu vaccine production or pandemic fu vac- cine. Both sites have traditionally conducted cell culture in stainless steel bioreactors. Jamal Meghrous, Ph.D., senior scientist, tells GEN that for hemagglutinin expression, single-use bioreactors work as well as fxed- tank reactors, at least at bench scale. At the same time, he cites his company's signifcant investment in stainless steel. "Signifcant development work needs to be done before switching to single use," Dr. Meghrous advises. "Other issues exist that may affect cell growth and productivity." Nevertheless, the company recently inte- grated single-use processing for virus pro- duction. While upstream of hemagglutinin manufacture, these moves represent a posi- tive step toward single-use manufacturing for insect cell systems. 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