Advances in industry and medicine have led to the engineering of complex “designer” proteins, such as antibodies in targeted therapeutics and enzymes in process development. The ability to easily generate an almost infinite number of variants at the DNA level has increased the demand for improved protein expression methodologies to fully capture what can be produced genetically. Often, the protein of interest is eukaryotic in origin and may require posttranslational modifications specific to its native host or may be toxic to the host cells expressing them. Cell-free protein expression systems have allowed us to step beyond the limits of traditional in vivo expression methodologies by decoupling protein expression from host cell viability. Furthermore, the ability to produce complex proteins using cell-free transcription/translation systems uniquely enables high-throughput directed evolution and protein engineering efforts. Several cell-free protein expression systems have been developed in the last decade with recent advances focusing on special folding or assembly environments. Equally as important is the capability to transition from the in vitro system to larger-scale in vivo expression, while maintaining activity of the target protein…
Category: <span>Biologics Production</span>
Inductively coupled plasma mass spectrometry (ICP-MS) is a highly sophisticated multi-element analytical technique that is increasingly being used for trace analysis in various industries including analytical laboratories. The ICP-MS technique is capable of analysis down to sub parts-per-trillion (ppt) detection limits whereby the lowest detection limits can be achieved only in a cleanroom environment. Because water is used early in tests for trace elemental analysis with ICP-MS, it is obvious that any contamination from the water can compromise an entire analysis. Therefore, the water used must be of a high analytical grade quality (e.g., ASTM Type I water). The objective of the analytical test series described below is to ensure that the ultrapure water generated by the arium pro UV water system has a high purity level (in this case, free of all nondetectable metal elements) and can be used without any problems for trace analysis of elements performed with ICP-MS devices…
Tangential flow filtration (TFF) is widely used in biopharmaceutical processing for protein purification – a common application for TFF is ultrafiltration for concentration/diafiltration of proteins. In this type of application, the product protein is retained (concentrated) within the feed side of the ultrafiltration membrane, while the buffer components and other small impurities (smaller than the membrane pore size) freely pass through the membrane into the permeate side. Several scholarly articles are available in literature which discuss the ultrafiltration application as well as its optimization strategies. Another category of application where TFF finds significant use is in the clarification of cell culture bioreactor and microbial fermenter feed solutions using microfiltration membranes. In some of these microfiltration TFF applications (e.g., mammalian cell culture clarification), the product (protein) freely passes through the microfiltration membrane and is recovered on the permeate side, while the contaminating impurities (cells, cell debris, colloids) are retained on the feed side of the membrane. In certain other microfiltration TFF applications (allantoic fluid clarification in egg-based flu process), the product (flu virus) may get concentrated on the feed side of the microfiltration membrane (similar to an ultrafiltration step), while the contaminating impurities (ovalbumin, etc.) may get removed into the permeate side…
Tissue culture growth medium derives a substantial fraction of its growth-stimulating activity from the fetal bovine serum (FBS) commonly used as a supplement. In addition to the source of serum, non-essential additives such as colorized pH indicator dyes may also affect the growth stimulating properties of complete media. We show here that both of these culture medium components can dramatically affect gene expression in vitro. Using a custom gene expression chip for herpes simplex virus 1, we demonstrated significant changes of expression levels in several categories of viral genes including the immediate early viral transcription factors ICP0 (p < 0.05), ICP4 and ICP27 (both p < 0.001). This dependence of virus growth on serum source and other medium components have implications for not only in vitro virus studies, but also viral vector design and vaccine efficacy. This is especially true when examining a large DNA pathogen that potentially contains response elements that are common in the mammalian genome...
The Patient Protection and Affordable Care Act (PPACA), which was passed in 2010, included the Biologics Price Competition and Innovation Act (BPCIA) authorizing the United States Food and Drug Administration (FDA) to establish an abbreviated regulatory approval pathway for complex macromolecules produced in living cells or organisms. The BPCIA implements many new provisions relating to the abbreviated review and approval of follow-on biopharmaceutical products. These include: (1) evaluating the biochemical properties of reference and follow-on macromolecules; (2) establishing procedures for the exchange of information between product sponsors, regulatory agencies, and the federal court system; and (3) establishing periods of regulatory exclusivity which complement patent rights typically awarded to the sponsor of a reference molecule. Important aspects of the BPCIA, particularly issues relating to the exchange of information between product sponsors and the establishment of periods of regulatory exclusivity protecting a novel or follow-on macromolecule are discussed in this article. This is the second in a series of three articles describing key provisions of the Leahy-Smith America Invents Act (AIA) and the BPCIA that affect intellectual property rights of academic and corporate institutions having an interest in the life sciences…
A fairly extensive body of literature exists on the inactivation of the circovirus by physical (e.g., heat, irradiation) and chemical means (e.g., disinfection). This is probably due to the fact that the circovirus is an economically significant pathogen within the swine and poultry industries. Sufficiently high temperatures afford substantial circovirus inactivation as do certain disinfectants and chemicals. However, there have been relatively few reports on the use of irradiation as an inactivation approach for this family of viruses. There is empirical evidence that gamma irradiation, at the doses normally employed for risk mitigation (25–45 kGy), is relatively ineffective on circoviruses. Fortunately, data on the effectiveness of ultraviolet radiation in the C range (UV-C, 248–254 nm, max) for the inactivation of other single-stranded DNA viruses (e.g., parvovirus) have been used in two orthogonal prediction models which estimate that the circovirus should be similarly sensitive to UV-C irradiation. Overall, appropriately-selected inactivation approaches may provide a high degree of risk mitigation for the circoviruses, whether present as contaminants of facility surfaces, animal-derived raw materials, or other process additives…
The manual preparation of cell banks in screw-capped cryovials is a slow and labour-intensive process which, due to the sensitivity of cells to the cryopreservation solutions, limits the sizes of batches that can be produced. Manual pipetting and vial-capping tasks increase contamination risks and quality control costs, and raise the likelihood of strain and repetitive motion injuries amongst laboratory staff. TAP Biosystems set an objective to create an automated system for reducing manual processing steps while increasing throughput, and maintaining the same or better sample quality over existing manual processes. Significant improvements have been achieved in tube processing times, reproducibility of filling volumes, and QC costs — all of which are described in this paper…
More and more companies are encountering added pressures to reduce costs, risks, and time-to-market while maximizing efficiencies. They have converged on a common strategy of outsourcing non-core functions. A trend is developing where companies as small as start-up biotechs to the major pharmaceutical and biopharmaceutical producers are outsourcing their peptide production needs. Whether a company needs research or GMP-grade materials, they are realizing enhanced cost-effectiveness and efficiency by doing so…
As clinical trials become progressively more complex, pharmaceutical and biotech companies face a multitude of logistical challenges. The age of personalized medicine and small-molecule drug development has made it imperative that clinical trial samples reach their destinations safely, securely, and at the right temperatures. The heightened demand is reflected by the fact that transportation and logistics services that can account for as much as 30% of a study budget. This has made managing the biomaterial cold chain a critical component in the research and development of biopharmaceutical products. Cold chain management defines how temperature-sensitive products and biomaterials such as clinical trial samples, cell banks, and active pharmaceutical ingredients are packaged, transported, and stored throughout the research and development process. Any weak link in this chain can compromise sample or product integrity, breach security, delay shipments, and ultimately result in financial loss or liability…
Government policies affecting intellectual property rights and the review of food and health care products dramatically influence investments in research leading to the development and sale of products that serve unmet medical needs or provide consumers with safe sources of food and drug products at a low cost. When statutes that affect several regulatory agencies are revised within a short time period, institutions that rely on exclusive rights offered by those agencies in exchange for obligations of disclosure and compliance must alter their business plans to adjust to new rules leading to the benefit conferred by the government. In 2011, the Leahy-Smith America Invents Act (AIA) was passed, changing many aspects of the federal statutes relating to the United States Patent and Trademark Office (PTO), and in 2010, the Patient Protection and Affordable Care Act (PPACA) was passed, which included the Biologics Price Competition and Innovation Act (BPCIA), requiring the United States Food and Drug Administration (FDA) to establish an abbreviated regulatory approval pathway for complex macromolecules produced in living cells or organisms. This series of articles briefly reviews key aspects of the AIA and the BPCIA, plus recent court cases relating to complementary periods of exclusivity offered by the FDA and the PTO, which should be of great interest to academic and corporate institutions having an interest in the life sciences. Important aspects of the AIA will be discussed in the first article in this series…