Many potentially therapeutic products involve the culture of stem cells. Their commercial success depends on the development of scalable good manufacturing practice (GMP) technologies that can both robustly and cost-effectively produce very large numbers of cells. Through many improvements and innovations in bioprocessing operations over the years, fed-batch suspension culture has remained the most common mode for large-scale biopharmaceutical manufacturing. However, some recent events suggest that may be changing (1,2). For the culture and expansion of stem cells, large-format adherent…
Upstream Processing
Single-Use Bioreactors and Microcarriers
Cell-based therapies hold promise for treating many acute and chronic diseases (1). Optimism surrounding that therapeutic potential has driven the initiation of multiple clinical trials in pursuit of such treatments. Procedures for preparing these therapeutic agents begin with selective isolation of cells from desired tissues. That is followed by ex vivo expansion of cells of desired phenotype and functionality. Once expanded to acceptable levels, cells are stored to preserve their viability during transportation to treatment facilities. The final step in…
North, South, East, and West
Electrophoresis is the basis of all blotting methods, and BPI Lab covered it last month (1). Electroblotting is a method for transferring electrophoretically separated proteins or nucleic acids onto a polyvinylidene fluoride (PVDF) or nitrocellulose membrane for permanence using electric current and a transfer buffer solution. This allows for analysts to further study them using probes, ligands, or stains. Capillary blotting is a variation designed to work with capillary electrophoresis. After electrophoresis the following are stacked in cathode-to-anode order: a…
Enabling Technologies
Many technological advancements in recent years have enabled companies to shorten time to market, to better understand their manufacturing processes, and to characterize their products well. In BPI’s December 2013 issue (pages 47–50), I reported on the first half of an informal reader survey about those technologies, with commentary from some survey participants and others. This month concludes with my examination of analytical, formulation/fill–finish, and facilities technologies. Analytical Technologies After writing several installments of our new “BPI Lab” series this…
Ready-to-Use Cryopreserved Primary Cells
Abiological measurement of drug activity is perhaps the most critical step in the series of tests required for product release both for clinical trials and the market. This evaluation plays an important role in the stability assessment of drug candidates. According to the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) Q6B document, measurements of biological activity can be performed in defined animal models that demonstrate a measurable physiological change in response to…
Nucleic Acid Impurity Reduction in Viral Vaccine Manufacturing
Commercial-scale viral vaccine manufacturing requires production of large quantities of virus as an antigenic source. To deliver those quantities, a number of systems are used for viral replication based on mammalian, avian, or insect cells. To overcome the inherent limitations in production outputs with serial propagation of cells, mammalian cells can be immortalized, which increases the number of times they can divide in culture. Modifications that immortalize cells are typically accomplished through mechanisms similar to those converting normal cells to…
IgM Purification with Hydroxyapatite
Hydroxyapatite (HA) has a long and successful history in the field of antibody purification, and it has worked well for immunoglobulin M (IgM) monoclonal antibodies (MAbs) (1,2,3,4,5,6,7,8). Applications range from initial capture to intermediate purification to final polishing. HA is best known for its superior ability to reduce antibody aggregates, but it also supports excellent reduction of DNA, viruses, and endotoxins. As IgM MAbs exhibit increasing potential in the fields of cancer and infectious disease and in stem-cell therapies, HA’s…
Assay Acceptance Criteria for Multiwell-Plate–Based Biological Potency Assays
For most biopharmaceuticals, potency is assessed in a bioassay by comparing dose–response curves of the test material and a reference standard. As with all analytical techniques, such assays require criteria by which their execution can be judged objectively to be valid, regardless of whether the desired or expected result is obtained for the test sample. PRODUCT FOCUS: BIOLOGICSPROCESSFOCUS: R&D, QCWHO SHOULD READ: PRODUCT AND PROCESS DEVELOPMENT, ANALYTICAL, QCKEYWORDS: IMMUNOASSAYS, POTENCY ASSAYS, PRODUCT RELEASE, REFERENCE STANDARDS, CONTROL SAMPLES, SAMPLE ACCEPTANCELEVEL: ADVANCED…
Mathematical Model for Production of Recombinant Antibody 14D9 By Nicotiana tabacum Cell Suspension Batch Culture
Transgenic plants are increasingly considered a competing system for producing high-value recombinant proteins for biomedical and industrial purposes at affordable costs (1). Researchers have shown that molecular farming (or biopharming) is a secure technology that is capable of rendering valuable recombinant proteins free of toxins and animal pathogens in a relatively short time (2,3,4,5,6). Scientists have also demonstrated that most recombinant antibodies produced in plants maintain their functional properties (substantial bioequivalence) as well as do those produced in mammalian cell…
Accounting for the Donnan Effect in Diafiltration Optimization for High-Concentration UFDF Applications
The biopharmaceutical industry is targeting high-concentration protein formulations to enable subcutaneous administrations. Such administration can provide better patient convenience than intravenous administration. One challenge associated with high-concentration formulations is increased electrostatic interaction between proteins and excipients. That is a result of increased protein-charge density at high protein concentrations. Such interactions can create an offset between excipient levels in final products and diafiltration buffers in ultrafiltration processes. The effect of such electrostatic interactions in a membrane process is known as the…