A Novel Nanofiber Membrane to Increase Yield and Purity for Cell and Gene Therapy Applications Sara Arroyo, Ian Scanlon, Ben Wallis, Adam Pinnock, Nathaniel Hughes, Eric Schmid, Marc Hummersone, Bilal Ahmad, Astrea Bioseparations Ltd, Cambridge, CB23 7AJ, UK
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Introduction
Bioprocess adsorbents, both bead- and membrane-based, are limited by capacity, flow rate and pressure drop. These adsorbents are also expensive to manufacture and can easily become unstable under different process conditions (e.g., temperature, pH, etc). Membranes constructed from electrospun nanofibers have shown great promise when applied to the separation of biological substances, particularly for fragile vectors used in cell and gene therapies. Recently, membrane adsorption/chromatography using nanofibers have shown great promise for use in bioseparations [1, 2]. Such single component nanofibers are superior to single component microfibers, as pore sizes, affinity characteristics, and other performance criteria, can be more precisely controlled. However, single-component nanofiber structures are often less efficient in terms of stability and time requirements. There is a need to further improve stability of these single-component nanofiber structures and the purification efficiency of biological products. Here, we demonstrate that AstreAdept™, a novel composite nanofiber material, can bring fast flow rates, increased binding capacities to biological product purification, and ameliorate many of the limitations posed by single-component fiber systems.
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Combined Flexibility and Strength of AstreAdept®
AstreAdept® shows high specific strength while retaining flexibility Single component nanofibers have limitations, such as high fragility and poor mechanical strength. AstreAdept® combines the advantages of the cellulose and non-cellulose based nanofiber materials to create a nanofiber matrix that exhibits a higher specific strength while retaining flexibility, so overcoming the limitations of traditional single component nanofiber membrane. Figure 3. A comparison of specific strength and strain to failure of the cellulose, non-cellulose based nanofiber vs the composite nanofiber.
Production of AstreAdept®
A dual electrospinning technique for the fabrication of materials with superior characteristics
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Application of AstreAdept®
AstreAdept® brings improved purification efficiency to bioseparation processes
AstreAdept is a composite nanofiber structure created by electrospinning of two different polymers together, combing cellulosic and non-cellulosic materials. ®
High flow rates without compromising on dynamic binding capacity High dynamic binding capacity BSA binding of 120 mg/mL
BSA breakthrough at 60mv/min 1s residence time
A schematic of the production of this composite nanofiber structure is shown in figure 1. Here, two different polymeric solutions (one is a derivative of cellulose and the other a non-cellulose synthetic polymer) are fed through a metallic needle under an applied voltage. Both polymer nanofibers are deposited on to the same substrate material in sequential layers thus forming a composite membrane.
Figure 1. Schematic of the production of a composite nanofiber
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AstreAdept® nanofiber, functionalized with weak anion exchange, was tested using a chromatography system to characterize dynamic binding capacity and flow rate characteristics.
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Structure of AstreAdept®
Open structure allows immediate access to large surface area for binding
The Future Impact
• Lab-scale spin columns are available, enabling an increased throughput of lentiviral vector sample preparation • Development of devices incorporating AstreAdept® technology is underway to address the needs of bioseparation processes at process development, pilot, and manufacturing scales for additional cell and gene therapy applications
Scanning Electron Microscopy (SEM) of the composite nanofiber membrane, figure 2, demonstrate high surface area to volume ratio and increased permeability. These characteristics provide a greatly increased accessible binding surface area, with potential for high flow rates. The resulting composite nanofiber membrane contains fibers having two distinct mean fiber diameters belonging to each of the polymers: • Cellulose derivative, 575nm (+/- 50 nm) • Non-cellulose synthetic polymer, 200 nm (+/- 50 nm)
Figure 2. SEM of composite nanofiber membrane
Information
Easy-to-use spin column format
Schematic of devices suitable for process development, pilot and manufacturing scales
References: 1. Chia-Chi Ho, Chapter 7 - Membranes for Bioseparations, Bioprocessing for Value-Added Products from Renewable Resources, Elsevier, 2007, Pages 163-183. 2. Menkhaus, Todd & Zhang, Lifeng & Fong, Hao. (2010). Applications of electrospun nanofiber membranes for bioseparations. Applications of Electrospun Nanofiber Membranes for Bioseparations. 1-59.
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Summary The use of this novel nanofiber adsorbent addresses a critical need in downstream processing of fragile modalities by providing a fit-for-purpose membrane which results in reducing process times, increasing process efficiency, reducing buffer consumption and consequently waste treatment costs, ultimately resulting in more cost effective and greener bioprocessing.