In order to unravel new protein activities and functions, we have expressed and purified a large number of human proteins. We have chosen to study secreted proteins and the extra-cellular domains of putative single transmembrane domain-containing proteins. In order to retain the natural protein characteristics as far as possible, we have used a mammalian expression system. Human embryonic kidney (HEK293) cells were chosen as they have been shown to possess a high protein-secretory potential. The secreted proteins were expressed with a carboxy-terminal tag and purified by affinity chromatography. Each protein was produced at a routine scale from 500 ml cell cultures, and the secreted protein was purified from the culture supernatant…
BioProcessing Journal Posts
The development and commercialization of biopharmaceutical products is paving the way for significant growth in the pharmaceutical industry. Virtual and small biotech companies are researching and developing new biotechnology-based products that help large pharmaceutical companies backfill their pipelines. Many of these products have been in development for years and are finally reaching the point of commercialization. Because of the nature of many of these substances, they are sensitive to their environment, their packaging and their delivery systems…
The viable cell concentration is of prime importance in monitoring cell culture manufacturing processes. Of the available online biomass assays, the radio-frequency impedance (RFI) method has some clear advantages for manufacturing because it is an unambiguous reflection of viable cell biovolume rather than the total number of cells. This allows RFI to be used to control feeding rates or to maintain a constant level of biomass within the bioreactor. RFI is also suitable for measuring the live cell density in bioreactors when the cells are attached to microcarriers and to inert discs…
With the continued growth of the biopharmaceutical market, the cell culture industry has seen a major shift away from the use of serum and other animal-derived supplements in the manufacture of biopharmaceuticals. Indeed, supporting guidance from the EMEA and FDA for the manufacture of biopharmaceuticals and medical devices encourages the use of “animal-free” components. The key driver for this can be attributed to the increased concern with contamination from adventitious agents such as transmissible encephalopathies…
The analytical characterization of recombinant protein therapeutic drug products has broadened to include the use of more sophisticated technologies. The expansion of technical abilities has translated into increasing the depth and breadth of our knowledge and understanding of the drug product intended for commercialization. With the availability of more precise methods, the regulatory expectations for understanding the characteristics of a protein therapeutic drug product are increasing. A thorough understanding of a therapeutic protein’s biochemical and biophysical characteristics is necessary to support investigational new drug (IND) applications and other drug regulatory filings…
Three types of heart valves are employed for replacement in patients: mechanical, xenogeneic tissue, and allogeneic human valves derived from donors post-mortem. The intention of this article is two-fold: 1) to place issues associated with the preservation of allogeneic human heart valves in perspective relative to the AATB Standards for Tissue Banking and current practices in US cardiovascular tissue-processing facilities; and 2) present potential opportunities for improvements with appropriate validation. Most patients receive either xenogeneic tissue or mechanical valves; however, the use of cryopreserved human valve allografts became established during the 1970s and 1980s for certain patient subsets…
A patent permits the patent holder to keep others out. That is, a patent holder can prevent others from making, using, selling, offering to sell, or importing an invention within the United States and its territories and possessions. In the United States, these exclusionary rights generally last for 20 years from the date on which the patent application was filed. What the patent holder can keep others out of is defined by the “claims” of the patent. Patent claims describe the boundaries of the patent, much like a real property deed outlines the borders of a parcel of land. However, having a patent does not guarantee that the patent holder is free to use the claimed invention…
Electrosprays and electrospinning are two interrelated physical phenomena which have been investigated for well over a century. Their similarity is based upon the primary driving mechanism; namely, the applied electric field. However, they have a fundamental difference that distinguishes one from the other: the former generates droplets while the latter forms continuous fibres. These two processing routes have been extensively researched in many areas. Within the realm of life sciences, these routes have ranged from novel bioanalytical approaches (DNA and biomolecules) to tissue engineering by the formation of scaffolds, which mimic extracellular matrices. Only lately have these methods been explored for the direct process handling of living cells…
Monoclonal antibodies and recombinant proteins have increased in importance and gained success as therapeutic agents in treating various diseases. Biomanufacturing of such a biopharmaceutical product by cell culture follows a main route. Upstream processing is strictly biology-driven, while on the other hand, purification is engineering driven. Fermentation is setting the pace. To some extent, that pace is a result of recent advances in cell culture, greatly increasing the densities of cells along with cell-related contaminants…