Micro-scale processing (MSP) techniques are miniaturized version of upstream and downstream conventional unit operations that are designed to accelerate the pace of bioprocess design and development. Previous ‘dead end’ filtration studies have demonstrated the usefulness of this concept for membrane filtration processes. However, these experiments were performed without stirring which is the most common strategy to control the effects of concentration polarization and fouling on filtration performance. In this work, the pressure-driven stirred conditions of a conventional stirred-cell module were integrated with a 96-well filter plate to develop a high throughput technique called ‘stirred-well filtration’ (SWF). The design allowed for up to eight constant flux filtration experiments to be conducted at once using a multi-rack programmable syringe pump and a magnetic lateral tumble stirrer. An array of pressure transducers was used to monitor the transmembrane pressure (TMP) in each well. The protein sieving behavior and fouling propensity of Omega™ ultrafiltration membranes were assessed via a combination of hydraulic permeability measurements and protein sieving tests in constant filtrate flux mode. The TMP profile during filtration of bovine serum albumin (BSA) solution was strongly dependent on the stirring conditions – for example the maximum TMP in the stirred wells were an average of 7.5, 3.8, and 2.6 times lower than those in the unstirred wells at filtrate fluxes of 12, 36, and 60 LMH (5, 15, and 25 μL/min) respectively. The consistency of the data across different wells for the same stirring condition was very good. To demonstrate the effectiveness of the SWF technique, the eight tests for a simple 2^2 factorial design-of-experiments (DOE) test with duplicates was run to evaluate the effect of solution pH and salt concentration on protein filtration. The combination of SWF with statistical methods such as DOE is shown to be an effective strategy for high-throughput optimization of membrane filtration processes. / Dissertation / Master of Applied Science (MASc)

Capacitive Deionization (CDI) is an energy-efficient desalination technology that utilizes an electric field to extract ions from water. Flow-through CDI systems show potential for superior desalination performance compared to traditional flow-by CDI; however, they face the challenge of increased occurrence of Faradaic reactions, leading to undesired by-products and reduced energy efficiency. In this study, we constructed a flow-through CDI cell and investigated the desalination performance of the two possible cell configurations: upstream anode mode and downstream anode mode. A series of experiments were conducted, measuring conductivity and pH of the effluent solution during charging and discharging phases. The results were analyzed in terms of salt adsorption capacity and charge efficiency. We used pH fluctuations in the effluent solution as indicators of Faradaic reactions. It was found that upstream anode mode yielded superior desalination, with a salt adsorption capacity of 6.79 mg/g and charge efficiency of 64.3%, compared to downstream anode mode, which displayed a salt adsorption capacity of 5.19 mg/g and charge efficiency of 50.8%. However, upstream anode mode also produced more pronounced pH oscillations, suggesting a higher occurrence of Faradaic reactions. Reconciling these conflicting results and shedding light on the complex processes within the CDI cell calls for further investigation.

The SARS-CoV2 pandemic has demonstrated on a global scale that viral infections can be highly contagious, can evolve rapidly, and are challenging to treat. The immune system provides cells with various control mechanisms to detect and prevent the spread of viral infection and further damage to the host. However, viruses have evolved methods to evade immunity, resulting in persevered viral replication and proliferation. Chronic viral infections occur when a virus evades immunity and persists in the body for an extended period, which can lead to increasingly harmful damage to the host, including increased risk of cancer. When immunity proves insufficient, alternative methods to sense virally infected cells can allow for detection and targeted elimination of the virus, which is especially necessary in cases of chronic viral infection. In this thesis, the development and characterization of RNA-editing enzymes based on adenosine deaminase acting on RNA (ADAR) that have been engineered to activate in response to viral protease is discussed. Specifically, methods for targeting ADAR editing to specific mRNA transcripts and strategies in which the editing activity of engineered ADARs has been made conditional upon viral proteolytic activity are explored. The development of fluorescent and quantitative assays to characterize systems are described and the implementation of the system to control downstream transcriptional activity is discussed. This thesis explores establishing the viability of a viral protease sensor able to be self-contained in an RNA circuit, which in the future may provide a treatment method for patients with severe symptoms or chronic viral infection. The ability to sense virally infected cells and create a functional output in specific response to viral protease presence as a potential future treatment of chronic viral infection is explored through viral protease activation of engineered ADAR enzymes to enable editing of specific mRNA transcripts. / 2025-05-24T00:00:00Z

The IoT offers enormous potentials thanks to its Widespread adoption by many industries, individuals, and governments, leading explosive growth and remarkable breakthroughs that have made it a technology with seemingly boundless applications. However, the far-reaching IoT applications cum its characteristic heterogeneity and ubiquity come with a huge price for more security vulnerabilities, making the deployed IoT systems increasingly susceptible to, and prime targets of many different physical and cyber-attacks including insider attacks, thereby growing the overall security risks to the systems. This research, which focuses on addressing insider attacks on IoT, studies the likelihood of malicious insiders' activities compromising some of the security triad of Confidentiality, Integrity and Availability (CIA) of a supposedly secure IoT system with implemented security mechanisms. To further establish the vulnerability of the IoT systems to the insider attack being investigated in our research, we first produced a research output that emphasized the need for multi-layer security of the overall system and proposed the implementation of security mechanisms on components at all layers of the IoT system to safeguard the system and ensure its CIA. Those conventional measures however do not safeguard against insider attacks, as found by our experimental investigation of a working IoT system prototype. The outcome of the investigation therefore necessitates our proposed solution to the problem, which leverages the integration of distributed edge computing with decentralized Ethereum blockchain technology to provide countermeasures that preserve the Integrity of the IoT system data and improve effectiveness of the system. We employed the power of Ethereum smart contracts to perform integrity checks on the system data logically and take risk management decisions. We considered the industry use case of Downstream Petroleum sector for application of our solution. The solution was evaluated using datasets from different experimental settings and showed up to 86% accuracy rate. / Government of the Federal Republic of Nigeria through the Petroleum Technology Development Fund (PTDF) Overseas Scholarship Scheme (OSS)

Enzyme haben sich in vielen Industriezweigen zu wichtigen Werkzeugen entwickelt und könnten in Zukunft besonders aufgrund der Transformation der chemischen Industrie hin zu klimaneutralen und nachhaltigen Prozessen an Bedeutung gewinnen. Für einen flächendeckenden Einsatz fehlt es jedoch an kosteneffizienten und effektiven Reinigungsmethoden, da vor allem das Downstream Processing einen bedeutenden Anteil an den Herstellungskosten haben kann. In den vergangenen Jahren wurden eine Vielzahl neuer Verfahren etabliert, um kostenintensive, chromatografische Schritte im Downstream Processing zu ersetzen. Eine vielversprechende, jedoch wenig beachtete Technik für die Enzymreinigung ist die Schaumfraktionierung. Diese Methode beruht auf der Bildung stabiler Schaumblasen, die durch die Adsorption oberflächenaktiver Moleküle an der Gas-Flüssigkeits-Grenzfläche von Luftblasen entstehen. Dadurch können Moleküle im Schaum angereichert und gereinigt werden. Die Schaumfraktionierung zeichnet sich durch milde Betriebsbedingungen, geringe Betriebskosten und einen einfachen apparativen Aufbau aus. Ihr Einsatz wird jedoch durch die Verwendung von Tensiden zur Schaumerzeugung und -stabilisierung sowie die potenzielle Denaturierung und Inaktivierung von Enzymen an der Gas-Flüssigkeits-Grenzfläche und die Unspezifität der Methode verhindert. Fusionsproteine, abgeleitet von natürlichen oberflächenaktiven Proteinen wie (bakteriellen) Hydrophobinen, könnten aufgrund ihrer spezifischen Wechselwirkungen mit der Luft-Wasser-Grenzfläche Tenside als Schaumbildner ersetzen und eine Generalisierung der Methode ermöglichen. Vor diesem Hintergrund war es das Ziel der vorliegenden Arbeit, Fusionsproteine zu identifizieren, die eine tensidfreie, aktivitätserhaltende Schaumfraktionierung von Enzymen ermöglichen. Die Identifikation potenzieller struktureller Determinanten der Fusionsproteine, die die Aufrechterhaltung der Aktivität und die Anreicherung im Schaum bedingen, bilden eine wichtige Grundlage für künftige Untersuchungen und die Etablierung dieser Methode. Die Fusion des oberflächenaktiven Proteins Ranaspumin-2 (Rsn-2) mit dem Modellenzym β-Lactamase (Bla) resultierte in einem Fusionsprotein, das im Gegensatz zum nativen Enzym die Fähigkeit besaß, einen stabilen Schaum zu erzeugen. In Zerschäumungsversuchen konnte mit dem generierten Fusionsprotein eine tensidfreie Anreicherung in der Schaumfraktion erreicht werden. Des Weiteren gewährleistete die Fusion mit Rsn-2 einen Aktivitätserhalt während der Zerschäumung, wodurch Bla-Rsn-2 ~70% seiner Aktivität beibehielt. Vergleichende Untersuchungen mit einer Penicillin-G-Acylase (PGA) und einer Formiatdehydrogenase (RjFDH) bestätigten nicht nur die Ergebnisse, sondern außerdem die Generalisierbarkeit der Methode. Diese Untersuchungen zeigten jedoch auch den Einfluss der Enzyme auf die Schaumstabilität, sowie einen möglichen Einfluss der Fusion auf die Enzymaktivität. Zusammenfassend zeigten die Ergebnisse erstmals, dass ein Fusions-Tag (F-Tag) basierend auf einem oberflächenaktiven Protein ein veritables Mittel für eine gezielte Schaumfraktionierung von Enzymen sein könnte. Basierend auf diesen Untersuchungen wurden eine Vielzahl natürlicher, schaumstabilisierender Proteine als F-Tags in Zerschäumungsexperimenten untersucht. In diesen wurden alle Fusionsproteine mit variierender Ausbeute im Schaum angereichert und die F-Tags waren in der Lage, die Aktivität der fusionierten Enzyme in unterschiedlichem Ausmaß zu erhalten. Strukturmodell der Fusionsproteine in silico deuteten darauf hin, dass für eine optimale Aktivitätserhaltung vor allem ein großer Abstand zwischen den beiden Proteindomänen erforderlich war. Die Einführung einer künstlichen helikalen Linker-Domäne zwischen Enzym und F-Tag bestätigte diese Hypothese. Interessanterweise war der eingeführte kurze Linker R1 als F-Tag in der Lage, Enzyme aktivitätserhaltend und mit hoher Ausbeute im Schaum anzureichern. In darauffolgenden Schaumfraktionierungsexperimenten wurde versucht eine künstliche Verunreinigung (Grün fluoreszierendes Protein, eGFP) mithilfe der zuvor identifizierten drei besten F-Tags von den Fusionsenzymen abzutrennen. Die Ergebnisse zeigten, dass für eine nahezu vollständige Abtrennung der Verunreinigung ein komplexes Zusammenspiel aus Säulengeometrie, Prozessparametern und Zusammensetzung der Ausgangslösung notwendig war. Die Reinheit der Schaumfraktion wurde vor allem durch die Stabilität der erzeugten Schäume und der Drainage der interstitiellen Flüssigkeit bestimmt. Der Einsatz von Waschpuffer half dabei, die im Schaum eingeschlossene Ausgangslösung auszuspülen und die Reinheit der Schaumfraktion zu steigern. Die Anteile der Fusionsproteine im Schaum sowie deren Restaktivität wurde maßgeblich durch die spezifischen Wechselwirkungen der F-Tags mit der Grenzfläche bestimmt. Schlussendlich konnten aus den Untersuchungen jedoch keine spezifischen strukturellen Determinanten abgeleitet werden, die die Anreicherung der F-Tags im Schaum beeinflussten. Erstaunlicherweise zeigte der kleine Linker R1 unabhängig vom untersuchten Enzym in der Schaumfraktionierung die besten Schaumausbeuten und eine Reinheit von bis zu 96 %, während er in allen Experimenten eine Restaktivität der Enzyme von mindestens 80% gewährleisten konnte. Zusammenfassend wurden Proteine mit inhärent schaumstabilisierenden Eigenschaften identifiziert, die die aktivitätserhaltende Rückgewinnung und Reinigung von Enzymen durch die Schaumfraktionierung ermöglichten. Es wurden Eigenschaften der Fusionsproteine, mögliche Wechselwirkungen und Prozessparameter identifiziert, die die Schaumfraktionierung und den Aktivitätserhalt von F-Tag-Fusionsproteinen maßgeblich beeinflussten. Die Ergebnisse der vorliegenden Arbeit verdeutlichen das vielversprechende Potenzial der F-Tags bei der Etablierung einer tensidfreien, aktivitätserhaltenden Schaumfraktionierung als effiziente Methode für die Aufarbeitung von Enzymen im Downstream Processing und bilden eine Grundlage für das Verständnis und die notwendigen Weiterentwicklungen der etablierten Methode.

<p>Ribosomes are the universal ribonucleoprotein organelles that translate the genetic message from mRNA to protein. In prokaryotes, the ribosomal subunits are 30S and 50S subunit, which bind together during the translation process forming 70S ribosome. The ribosome is a highly dynamic structure, and acts as a working platform for the different factors involved in the process of converting the genetic information into protein.</p><p>Cryo-electron tomography (cryo-ET) is an emerging imaging technology that combines the potential of three-dimensional (3D) reconstruction at molecular resolution with a close-to-native preservation of the specimen. Here, we have applied this method to reconstruct rifampicin-treated <i>Escherichia coli</i> individual 30S subunits in vitro and in situ, and individual 50S subunits in situ. In the 30S subunit, the head, the platform and the body show large conformational movements relative to each other. The particles are grouped into three conformational groups according to the width/height ratios. Also, an S15 fusion protein derivative has been used as a physical reporter to localize S15 in the 30S subunit. In the 50S subunit, the L1 stalk, the L7/L12 stalk, the central protuberance (CP), and the peptidyl transferase center (PTC) cleft are the most dynamic and flexible parts in the reconstructed structures with clear movements indicated. Different locations of the tunnel in the central cross-sections through the in situ 50S subunits indicate a flexible pathway inside the large subunit. In addition, gross morphological changes were also been observed in our reconstructions. Our results demonstrate a considerable conformational flexibility among individual ribosomal subunits, both in vitro and in situ.</p><p>Translation is an essential process for all cells and organisms. Translation initiation is the rate-limiting step and the most highly regulated phase of translation process. Several regions along the mRNA have been reported to influence translation initiation. The Shine-Dalgarno (SD) sequence located 5-9 bases upstream of the initiation codon supports translation initiation by complementary binding to the Anti-Shine-Dalgarno (ASD) sequence on the 16S rRNA.</p><p>We have here compared how an SD⁺ sequence influences gene expression, if located upstream or downstream of an initiation codon. The positive effect of an upstream SD⁺ is confirmed. A downstream SD⁺ gives decreased gene expression. If an SD⁺ is placed between two potential initiation codons, initiation takes place predominantly at the second start site. The first start site is activated if the distance between this site and the downstream SD⁺ is enlarged and/or if the second start site is weakened. Upstream initiation is eliminated if a stable stem-loop structure is placed between this SD⁺ and the upstream start site. The results suggest that the two start sites compete for ribosomes that bind to an SD⁺ located between them. A minor positive contribution to upstream initiation resulting from 3’ to 5’ ribosomal diffusion along the mRNA is suggested. Since the location of SD⁺or SD-like sequences can strongly influence gene expression, this should be of significant evolutionary importance.</p>

Ribosomes are the universal ribonucleoprotein organelles that translate the genetic message from mRNA to protein. In prokaryotes, the ribosomal subunits are 30S and 50S subunit, which bind together during the translation process forming 70S ribosome. The ribosome is a highly dynamic structure, and acts as a working platform for the different factors involved in the process of converting the genetic information into protein. Cryo-electron tomography (cryo-ET) is an emerging imaging technology that combines the potential of three-dimensional (3D) reconstruction at molecular resolution with a close-to-native preservation of the specimen. Here, we have applied this method to reconstruct rifampicin-treated Escherichia coli individual 30S subunits in vitro and in situ, and individual 50S subunits in situ. In the 30S subunit, the head, the platform and the body show large conformational movements relative to each other. The particles are grouped into three conformational groups according to the width/height ratios. Also, an S15 fusion protein derivative has been used as a physical reporter to localize S15 in the 30S subunit. In the 50S subunit, the L1 stalk, the L7/L12 stalk, the central protuberance (CP), and the peptidyl transferase center (PTC) cleft are the most dynamic and flexible parts in the reconstructed structures with clear movements indicated. Different locations of the tunnel in the central cross-sections through the in situ 50S subunits indicate a flexible pathway inside the large subunit. In addition, gross morphological changes were also been observed in our reconstructions. Our results demonstrate a considerable conformational flexibility among individual ribosomal subunits, both in vitro and in situ. Translation is an essential process for all cells and organisms. Translation initiation is the rate-limiting step and the most highly regulated phase of translation process. Several regions along the mRNA have been reported to influence translation initiation. The Shine-Dalgarno (SD) sequence located 5-9 bases upstream of the initiation codon supports translation initiation by complementary binding to the Anti-Shine-Dalgarno (ASD) sequence on the 16S rRNA. We have here compared how an SD+ sequence influences gene expression, if located upstream or downstream of an initiation codon. The positive effect of an upstream SD+ is confirmed. A downstream SD+ gives decreased gene expression. If an SD+ is placed between two potential initiation codons, initiation takes place predominantly at the second start site. The first start site is activated if the distance between this site and the downstream SD+ is enlarged and/or if the second start site is weakened. Upstream initiation is eliminated if a stable stem-loop structure is placed between this SD+ and the upstream start site. The results suggest that the two start sites compete for ribosomes that bind to an SD+ located between them. A minor positive contribution to upstream initiation resulting from 3’ to 5’ ribosomal diffusion along the mRNA is suggested. Since the location of SD+ or SD-like sequences can strongly influence gene expression, this should be of significant evolutionary importance.

<p>Abstract</p><p>The aim of the present study was to increase the understanding of how the cell surface properties affect the performance of unit operations used in primary protein purification. In particular, the purpose was to develop, set up and apply methods for studies of cell surface properties and cell interactions.</p><p>A method for microbial cell surface fingerprinting using surface plasmon resonance (SPR) is suggested. Four different <em>Escherichia coli </em>strains were used as model cells. Cell surface fingerprints were generated by registration of the interaction between the cells and four different surfaces, with different physical and chemical properties, when a cell suspension was flown over the surface. Significant differences in fingerprint pattern between some of the strains were observed. The physical properties of the cell surfaces were determined using microelectrophoresis, contact angle measurements and aqueous two-phase partitioning and were compared with the SPR fingerprints. The generated cell surface fingerprints and the physical property data were evaluated with multivariate data analysis that showed that the cells were separated into individual groups in a similar way using principal component analysis plots (PCA).</p><p>Studies of the behaviour of the model cells on stirred cell filtration and in an interaction test with different expanded bed adsorption (EBA) adsorbents were performed. It could be concluded that especially one of the strains behaved differently. Differences in the properties of the model cells were indicated by microelectrophoresis and aqueous two-phase partitioning which to some extent correlated with observed differences in behaviour during filtration and in an interaction test with EBA adsorbents.</p><p>The impact of high-pressure homogenisation of <em>E. coli </em>cell extract was examined, with a lab scale and a pilot scale technique. The DNA-fragmentation, visualised with agarose gel electrophoresis, and the resulting change in viscosity was analysed. A short homogenisation time resulted in increased viscosity of the process solution that correlated with increased concentration of released non-fragmented DNA. With longer homogenisation time the viscosity decreased with increasing degree of DNA-fragmentation.</p><p>The results show that strain dependant cell surface properties of<em> E. coli</em> may have an impact on several primary steps in downstream processing.</p>

Les gobies amphidromes et, en particulier, ceux de la sous famille des Sicydiinae représentent une part importante de la diversité et de l'abondance des peuplements de poissons d'eau douce des îles tropicales. Ces espèces se reproduisent dans les rivières et ont une phase larvaire marine pendant plusieurs mois avant de retourner croitre et maturer en eau douce. L'objectif de cette thèse est d'acquérir des connaissances concernant la phénologie des dynamiques de migration en eau douce de deux espèces de Sicydiinae de La Réunion, Sicyopterus lagocephalus et Cotylopus acutipinnis et les mécanismes qui peuvent en être à l'origine. Ces études montrent que la dévalaison des larves vers la mer immédiatement après leur éclosion avait principalement lieu pendant l'été austral et en début de nuit dans les zones aval. Le débit des cours d'eau et ses fluctuations saisonnières et journalières jouent un rôle prépondérant dans cette dynamique de dévalaison en termes d'abondances de larves dérivant, de temps de dévalaison jusqu'à la mer et de survie des larves. Après leur arrivée en eau douce, les juvéniles vont coloniser l'ensemble des zones des bassins versants. Les plus fortes abondances de juvéniles en migration vers les zones amont sont observées en fin d'étiage et pendant l'après-midi. Enfin, S. lagocephalus présente des performances locomotrices supérieures à celles de C. acutipinnis soutenues par des morphologies plus diverses. Ces meilleures performances, soutenues par des morphologies plus diverses, sont l'un des facteurs qui peuvent expliquer la large aire de répartition de S. lagocephalus, présent dans les océans Indien et Pacifique, par rapport à C. acutipinnis qui est endémique de l’archipel des Mascareignes. Des recommandations de gestion, permettant principalement de restaurer la continuité biologique au niveau des barrages, sont faites au regard des résultats obtenus durant cette thèse. / Amphidromous gobies, especially those of the Sicydiinae subfamily, represent most of the diversity and abundance of the freshwater fish assemblages in tropical islands. These species spawn in rivers, spend months in the ocean as pelagic larvae and return to freshwater to grow and reproduce. This doctoral thesis aims at describing the phenology and some mechanisms of the dynamics of migration in freshwater of two Sicydiinae species in Réunion Island: Sicyopterus lagocephalus and Cotylopus acutipinnis. These studies highlighted that larval downstream migration to the sea, immediately after hatching, occurs mainly during austral summer and a few hours after sunset in downstream reaches. Seasonal and daily fluctuations of the flow regime also greatly influence the abundances, the transport duration from spawning site to the sea and the survival of larvae during their downstream migration. After their return to the freshwater, juveniles settle in the watersheds from the estuary to the most upstream reaches. The highest abundances of juveniles migrating to upstream reaches are observed during low flow conditions at the end of the afternoon. Finally, locomotor performances are better for S. lagocephalus compared to C. acutipinnis. These better locomotor performances, supported by more diverse morphologies, are among the factors explaining the presence of S. lagocephalus from Eastern Pacific Ocean to Western Indian Ocean when C. acutipinnis is endemic to the Mascarenes Archipelago. Based on the results obtained in this doctoral thesis, management and conservation recommendations are suggested especially for the restoration of fish passage at several migration barriers such as dams and weirs.

Human mesenchymal stem cells (hMSCs) have been identified as a promising cell-based therapy candidate to treat a number of unmet clinical indications, however, in vitro expansion will be required to increase the available number of cells and meet this demand. Scalable manufacturing processes, amenable to closed, single-use and automated technology, must therefore be developed in order to produce safe, effective and affordable hMSC therapies. To address this challenge, a controlled serum-free end-to-end microcarrier process has been developed for hMSCs, which is amenable to large-scale manufacture and therefore increasing economies of scale. Preliminary studies in monolayer culture assessed the level of variability in growth between five hMSC donors, which was found to have a variance of 25.3 % after 30 days in culture. This variance was subsequently reduced to 4.5% by the development of a serum-free monolayer culture process with the maintenance of critical hMSC characteristics and an increased number of population doublings. In order to transfer this into a scalable system, the serum and serum-free expansion processes were transferred into suspension by the addition of plastic microcarriers in 100 mL spinner flasks without control of pH or dissolved oxygen (DO). This achieved a maximum cell density of 0.08 ± 0.01 · 106 cells.mL-1 in FBS-based medium, 0.12 ± 0.01 · 106 cells.mL-1 in HPL-based medium and 0.27 ± 0.03 · 106 cells.mL-1 in serum free medium after six days. In order to drive consistency and yield into the manufacturing process, a process control system was developed for the FBS-based microcarrier expansion process in a 100 mL DASbox bioreactor platform to control DO, pH, impeller rate and temperature. Reduced impeller rates and DO concentrations were found to be beneficial, with a final cell density of 0.11 ± 0.02 · 106 cells.mL-1 and improved post-harvest outgrowth and colony-forming unit (CFU) potential compared to uncontrolled microcarrier and monolayer culture. This controlled bioreactor expansion process was then applied to the previously developed serum-free microcarrier process, eventually achieving a final cell density of 1.04 ± 0.07 · 106 cells.mL-1, whilst retaining key post-harvest hMSC characteristics. Following the controlled serum-free expansion and harvest of hMSCs, a downstream and cryopreservation process was developed to assess the impact of prolonged holding times and subsequent unit-operations on hMSC quality characteristics. This showed that hMSCs are able to maintain key characteristics throughout the entire end-to-end process, demonstrating their potential for commercial scale manufacture.