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…
QA/QC
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…
Design of Experiments for Fed-Batch Process Development in Shaken Cultures
When designing a recombinant protein production process, a high number of parallel cultivations must be carried out. That task is typically performed using batch cultures in shake flasks or microwell plates, in which fermentation conditions are not monitored. To overcome that limitation, we combined the SensorDish Reader and Shake Flask Reader systems (from PreSens) with an enzymatic glucose delivery system (EnBase technology from BioSilta Oy) for Escherichia coli cultivations. Our objective was to determine whether SensorDish reader cultures would yield…
Responding to Life Sciences Manufacturing Industry Guidance
Increasingly, life science manufacturing companies are applying technology to meet quality by design (QbD) goals. Organizations collect overflowing volumes of process data as part of programs designed to improve manufacturing variability and outcomes. Collecting valuable data is now an everyday task thanks to available software and process analytical technology (PAT) tools. The industry today, in fact, has focused so much on gathering data that it often has lost sight of an important fact: Data collection systems are valuable only if…
Quality Risk Assessment and Management Strategies for Biopharmaceutical Companies
You’ve probably been hearing a lot about risk assessment in recent months. Indeed, some 15 times more articles have been printed referencing the concept over the past year relative to a 12-month period just three years ago. That truly represents a geometric progression. Unfortunately, very few authors have been able to disambiguate the different methods or provide insight into this time-tested, multiple-industry philosophy that at its core uses good science to make better decisions. When we undertake the challenge to…
Effects of Pressure Sensor Calibration Offset on Filter Integrity Test Values
Food and Drug Administration (FDA) and European good manufacturing practices (GMPs) require integrity testing of sterilizing-grade filters for producing injectables and other biologics. The diffusion test (also called the forward-flow test) and bubble-point test (also called the disk test) of a sterilizing-grade filter are both filter-integrity tests. The accuracy of both relies on calibration of a pressure sensor in the respective integrity test unit. Calibration of the pressure sensor of a filter-integrity testing device is an essential part of quality…
Process Improvements Increase Production Capacity of a Legacy Product
Implementation of postlicensure process improvements in the biopharmaceutical industry can benefit patients and drug manufacturers alike. Here we demonstrate through a case study how a change to the cell culture medium and process can be taken from proof of concept through scale-up to demonstration of feasibility. We further illustrate the scope and complexity of implementing a change in commercial manufacturing to realize significant benefits such as increased production capacity over an existing legacy process. The Importance of Postapproval Improvements Drug…
New Paradigms for Process Validation
Both the United States and the European Union have recently evolved guidance on how to execute process validation (1, 2) with the prospect of a more appropriate life-cycle approach. It goes beyond the traditional three to five lots run at the center point of proposed ranges for operating parameters. New approaches leverage product design and process development information. They facilitate adapting the quality by design (QbD) paradigm to allow for a science- and risk-based selection of critical process…
NIR Spectroscopy for Process Monitoring and Control in Mammalian Cell Cultivation
The quality by design (QbD) and process analytical technology (PAT) approaches have shown significant benefit in the classical pharmaceutical industry and are now strongly influencing bioprocessing. Monitoring critical process parameters (CPPs) during biotechnological cell cultivations is essential to maintaining high efficiencies and quality. Commercial sensor systems for real-time inline monitoring are available for some parameters, such as pH or the concentration of dissolved oxygen (DO). For others such as glucose concentration, total cell count (TCC), and viability no robust online…