Bioengineering2014, 1(4), 231-245; doi:10.3390/bioengineering1040231 (registering DOI) - published 27 November 2014 Show/Hide Abstract
Abstract: Recruitment of mesenchymal stromal cells (MSC) into the field of tissue engineering is a promising development since these cells can be expanded vivo to clinically relevant numbers and, after expansion, retain their ability to differentiate into various cell lineages. Safety requirements and the necessity to obtain high cell numbers without frequent subcultivation of cells raised the question of the possibility of expanding MSC in one-way (single-use) disposable bioreactors. In this study, umbilical cord-derived MSC (UC-MSC) were expanded in a disposable Z 2000 H bioreactor under dynamic conditions. Z was characterized regarding residence time and mixing in order to evaluate the optimal bioreactor settings, enabling optimal mass transfer in the absence of shear stress, allowing an reproducible expansion of MSC, while maintaining their stemness properties. Culture of the UC-MSC in disposable Z 2000 H bioreactor resulted in a reproducible 8-fold increase of cell numbers after 5 days. Cells were shown to maintain specific MSC surface marker expression as well as trilineage differentiation potential and lack stress-induced premature senescence.
Abstract: The development, optimization, and analysis of downstream processes are challenged by a high number of potentially critical process parameters that need to be investigated using lab-scale experiments. These process parameters are spread across multiple unit operations and potentially show interactions across unit operations. In this contribution, we present a novel strategy for bioprocess development that considers the risk of parameter interactions across unit operations for efficient experimental design. A novel risk assessment tool (interaction matrix) is introduced to the Quality by Design (QbD) workflow. Using this tool, the risk of interaction across unit operations is rated. Subsequently, a design of experiments (DoE) across unit operations is conducted that has the power to reveal multivariate interdependencies. The power of the presented strategy is demonstrated for protein isolation steps of an inclusion body process, focusing on the quality attribute inclusion body purity. The concentration of Triton X-100 in the course of inclusion body (IB) purification was shown to interact with the g-number of the subsequent centrifugation step. The presented strategy targets a holistic view on the process and allows handling of a high number of experimental parameters across unit operations using minimal experimental effort. It is generically applicable for process development along QbD principles.
Abstract: A steady increase of product titers and the corresponding change in impurity composition represent a challenge for development and optimization of antibody production processes. Additionally, increasing demands on product quality result in higher complexity of processes and analytics, thereby increasing the costs for product work-up. Concentration and composition of impurities are critical for efficient process development. These impurities can show significant variations, which primarily depend on culture conditions. They have a major impact on the work-up strategy and costs. The resulting “bottleneck” in downstream processing requires new optimization, technology and development approaches. These include the optimization and adaptation of existing unit operations respective to the new separation task, the assessment of alternative separation technologies and the search for new methods in process development. This review presents an overview of existing methods for process optimization and integration and indicates new approaches for future developments.
Abstract: Design and assessment activities associated with a biopharmaceutical process are performed at different levels of detail, based on the stage of development that the product is in. Preliminary “back-of-the envelope” assessments are performed early in the development lifecycle, whereas detailed design and evaluation are performed prior to the construction of a new facility. Both the preliminary and detailed design of integrated biopharmaceutical processes can be greatly assisted by the use of process simulators, discrete event simulators or finite capacity scheduling tools. This report describes the use of such tools for bioprocess development, design, and manufacturing. The report is divided into three sections. Section One provides introductory information and explains the purpose of bioprocess simulation. Section Two focuses on the detailed modeling of a single batch bioprocess that represents the manufacturing of a therapeutic monoclonal antibody (MAb). This type of analysis is typically performed by engineers engaged in the development and optimization of such processes. Section Three focuses on production planning and scheduling models for multiproduct plants.
Abstract: Biofunctional block copolymers are becoming increasingly attractive materials as active components in biosensors and other nanoscale electronic devices. We have described two different classes of block copolymers with biofuctional properties. Biofunctionality for block copolymers is achieved through functionalization with appropriate biospecific ligands. We have synthesized block copolymers of electroactive poly(3-decylthiophene) and 2-hydroxyethyl methacrylate by atom transfer radical polymerization. The block copolymers were functionalized with the dinitrophenyl (DNP) groups, which are capable of binding to Immunoglobulin E (IgE) on cell surfaces. The block copolymers were shown to be redox active. Additionally, the triblock copolymer of α, ω-bi-biotin (poly(ethylene oxide)-b-poly (styrene)-b-poly(ethylene oxide)) was also synthesized to study their capacity to bind fluorescently tagged avidin. The surface-active property of the poly(ethylene oxide) block improved the availability of the biotin functional groups on the polymer surfaces. Fluorescence microscopy observations confirm the specific binding of biotin with avidin.
Abstract: A hybrid cell sheet engineering approach was developed using ultra-thin nanofiber arrays to host the formation of composite nanofiber/cell sheets. It was found that confluent aligned cell sheets could grow on uniaxially-aligned and crisscrossed nanofiber arrays with extremely low fiber densities. The porosity of the nanofiber sheets was sufficient to allow aligned linear myotube formation from differentiated myoblasts on both sides of the nanofiber sheets, in spite of single-side cell seeding. The nanofiber content of the composite cell sheets is minimized to reduce the hindrance to cell migration, cell-cell contacts, mass transport, as well as the foreign body response or inflammatory response associated with the biomaterial. Even at extremely low densities, the nanofiber component significantly enhanced the stability and mechanical properties of the composite cell sheets. In addition, the aligned nanofiber arrays imparted excellent handling properties to the composite cell sheets, which allowed easy processing into more complex, thick 3D structures of higher hierarchy. Aligned nanofiber array-based composite cell sheet engineering combines several advantages of material-free cell sheet engineering and polymer scaffold-based cell sheet engineering; and it represents a new direction in aligned cell sheet engineering for a multitude of tissue engineering applications.