As more biologicals are being developed and approved for therapeutic use, the pressure to maintain consistent delivery of high-quality products remains critical. Although increasing product yield is important, streamlining the aseptic process is, without a doubt, at the top of the agenda for most firms.
Human intervention is the greatest source of contamination. Modifications to a process that minimize the amount of manual input, i.e., reduce the number of operators and intervention frequency, and the proportion of components that require manual cleaning, will undoubtedly improve the sterility of the end product. Thus, increasing the use of automation and replacing traditional glassware or stainless-steel components with single-use disposable products can significantly reduce the risk of contamination, in particular the risks associated with cleaning, storage, and reuse.
Today, organizations are looking for alternatives that improve economies of scale and provide assurances to the products’ integrity and efficacy including sterility and efficiency of upstream and downstream processing. In this article, we describe how such processes can be optimized using Nunc™ and Nalgene® products from Thermo Fisher Scientific (www.thermofisher.com) for production cell culture, fluid transfer, and fluid containment.
Single-use systems can not only provide a validated sterile solution but they can also negate the need for steam/water sterilization and its associated cleaning validation, presenting cost and time savings. Furthermore, automated modular systems that assist in the filling, incubation, transport, storage, and harvesting of cell culture systems reduce the risks of contamination.
Ideally, successfully optimized processes will need to be scaled-up and validated for high-volume production. Thus the incorporation of instrumentation and consumables that can rapidly facilitate this process and reduce revalidation where possible are of high value. For example, systems that use the same surface areas in small-scale development and large-scale production can simplify the transfer process.
Versatile and Scalable Formats
The Nunc Cell Factory, a portfolio of multilayer cell-culture devices, exhibits the same growth kinetics as laboratory-scale cell-culture platforms. In addition, it is a linearly scalable device, allowing for easy process scale-up. The Cell Factory promotes the reduction of manual handling by optimizing the amount of surface area in a small footprint. A full line of automation solutions is available to facilitate the process transfer from research to industrial-scale manufacturing.
The new EasyFill™ Cell Factory features one large port to facilitate pouring and one standard port that can accommodate customized tubing sets for a more closed system. The EasyFill Cell Factory is applicable for both research and GMP-scale processes. The Active Gassing Cell Factory promotes a controlled environment within the culture trays, providing equal distribution of the user-specified gas mix. This is of particular interest when growing more challenging cells such as HEK-293 or stem cells or when air and gas exchange is critical, i.e., for interferon production.
Novel Formats and Robust Materials
Utilizing durable materials such as polyethylene terephthalate glycol (PETG) as opposed to polystyrene (PS) for the production of roller bottles reduces the risk of contamination and loss of valuable material associated with breakage. PETG can be frozen down to -40°C, which promotes the use of nonenzymatic cell-harvesting techniques such as the freeze-thaw method.
Adaptation of the cells to PETG from PS flasks is not necessary to achieve high cell yields and viability. Studies have shown that higher cell densities (per cm2) and greater total cell yield can be achieved in a PETG roller bottle in comparison to a PS roller bottle. The InVitro PETG Expanded Surface Roller Bottle provides up to 2.5 times more growth area versus the standard roller bottles (Figure 1). Utilizing bottles with greater surface area translates into using fewer bottles, therefore reducing the contamination risk in addition to overall cost.
Throughout the bioproduction process there are a number of applications that require fluid transfer including, but not limited to, seeding a cell-culture device, sample collection, harvesting, purification, bulk intermediate storage, and final filling.
Design and development of the “middleware” used for fluid transfer can be a time-consuming process, taking weeks or even months to complete with many design iterations between the end-user and the supplier. The TopWorks™ Online Custom Tubing Set Configurator enables scientists and engineers to design their own integrated fluid-transfer and bioprocess systems using Nalgene and Nunc products (Figure 2).
The TopWorks browser interface is a rules-based platform that ensures design manufacturability, i.e., the components chosen are compatible with each other. At any time during the design process, the user can create a 2-D CAD drawing to visualize the design (Figure 3). The user dictates the speed of the project. The responsibility of the system build and sterility certification is passed to the solution provider eliminating the need for in-house assembly by high-value employees.
Integrity and assurance of the container chosen for the storage of an API or bulk intermediate is paramount. The negative implication of contamination at this stage can be economically significant. Minimizing the risk of contamination not only falls on the reliability of the components used to transfer the fluid to the container but is dependent on the container itself.
The choice between flexible and rigid solutions for the storage of a valuable drug substance is a challenging consideration. There are a variety of materials, shapes, and sizes in single-use disposable formats on the market today that provide integrity, stability, and assurance in addition to chemical resistance and durability.
Nalgene brand single-use, certified, sterile, rigid, containers are available in well-characterized and validated materials. This portfolio of containers ranges from small-volume options such as vials to large carboys all manufactured from the same materials, enabling stability studies to be performed on a smaller, more economical scale. In addition, these materials are all tested to ensure biological compatibility and compliance to regulatory standards.
The use of flexible bags is ever increasing due to the space-saving features. The conversion to flexible systems, however, is not without its challenges, one being how to store your bag without compromising the contents. This is extremely important under frozen conditions.
While rigid containers offer well documented durability at conditions of -80°C and below, flexible bags are more susceptible to cracks and tears. In addition, multiple bags can be difficult to lift, transport, and store when filled. The new Nalgene Bioprocess Bag Management System addresses all of these issues. Molded of a rigid polymer, the stackable, clamshell-like design provides a durable, protective barrier offering a combination of rigid and flexible systems; it is also designed to mate with commonly available “pillow” bag containers such as the 5 L and 20 L Thermo Scientific Hyclone BioProcess Container™.
The wealth of equipment and consumables available to the bioproduction industry is constantly expanding. The opportunity to source all materials such as instrumentation (shakers, incubators, and rack systems), automation, culture ware, as well as fluid transfer and containment solutions from one source can be quite efficient.
Jill Staggert is product manager, bioproduction, at Thermo Fisher Scientific. Web: www.thermofisher.com. Email: firstname.lastname@example.org.