High-Throughput Process Development for the Chromatographic Purification of Viral Antigens

Chromatography is a widely used method in the biotechnology industry and functions to separate the desired product from process and product related impurities. There is a multitude of resins available based on different modalities (such as charge, hydrophobicity, and affinity) to provide a spectrum of approaches to meet the separation challenges of the diverse products. The challenge of developing viral antigen purification processes is addressed in this method. A unique feature of this product class is that in order to protect against more than one strain of an antigen, vaccines are often multivalent. This entails multiple production processes for each antigen, all of which will require separate development and validation. Ideally, a universal purification method is sought, but differences in the protein subunits (frequently used as the antigens) make this challenging and often-bespoke purification steps are required. This means process development for the chromatographic stages of these products can be particularly challenging and labour intensive. With the numerous choices available, making critical process decisions that are usually unique to each product, process, and strain, can be costly and time-consuming. To address this, scale down purification at <1.0 mL column volume and automation approaches are increasingly applied to increase throughput. In this work, a method is described wherein a Tecan Freedom EVO® automated liquid handler is deployed for the evaluation of different resin chemistries and buffer conditions to find a suitable purification strategy. This method allows for the rapid evaluation of the separation viral antigens where limited information on chromatography behavior is known at the early stages of process development. Here, we demonstrate the methodology firstly by explaining the automated purification script and secondly by applying the script for an efficient purification development for different serotypes of rotavirus antigens

Prüfung verschiedener Erbsentypen auf deren Krankheitsanfälligkeit, Biomasse und Kornertrag

Generally, purple-flowered peas are considered to be less susceptible to soil-borne diseases and pests than white-flowered peas. Besides, forage peas usually produce more aboveground biomass and less grain yields in comparison to grain peas. Tolerance to soil-borne diseases, above- and belowground biomass and grain yields of two white-flowered (forage and grain pea, respectively) and one purple-flowered (forage pea) pea were examined in a field experiment in Bavaria, Germany. Purple-flowered peas had neither obvious higher tolerance to soil-borne diseases nor more biomass yield compared to white-flowered peas. Furthermore, the grain yields in this experiment were extremely low and there was no significant difference between the two types. A possible cause could be the unsuitable weather conditions, which caused an increased disease pressure and resulted in low grain and biomass yields. Hence, it can be assumed purple-flowered peas are generally not less susceptible to soil-borne pests than white-flowered peas

Evaluation of three medicinal plant extracts against Plasmodium falciparum and selected microganisms

Background: A great revival of scientific interests in drug discovery has been witnessed in recent years from medicinal plants for health  maintenance. The aim of this work was to investigate three Nigerian  medicinal plants collected in Nigeria for their in vitro antiplasmodial and antimicrobial activities.Materials and Methods: Extracts obtained from parts of Persea americana, Jatropha podagrica and Picralima nitida and their fractions wereevaluated for in vitro antiprotozoal and antimicrobial activity.Result: The methanol extract of P. nitida demonstrated activity against chloroquine-sensitive and chloroquine-resistant P. falciparum clones with IC50 values of 6.3 and 6.0 µg/mL, respectively. Methanol and chloroform extracts of P. americana seed showed antifungal activity against Cryptococcus neoformans IC50 less than 8 and 8.211 µg/mL respectively. Finally, the petroleum ether extract of P. americana had activity against methicillinresistant Staphylococcus aureus (MRSA) with an IC50 value of 8.7 µg/mL.Conclusion: The study revealed the antibacterial and antiplasmodial activities of the plants extracts at the tested concentrations.Keywords: Antifungal, Antibacterial, Persea americana, Picralima nitida, Jatropha podagrica, Plasmodium falciparu

Evolution of Heart Transplantation Surgical Techniques

Organ transplantation has kindled the human imagination since the beginning of time. Prehistorically, transplantation appeared as mythological stories: from creatures with body parts from different species, the heart transplant between two Chinese soldiers by Pien Ch’iao, to the leg transplant by physician Saints Cosmas and Damian. By 19th century, the transplantation concept become possible by extensive contributions from scientists and clinicians whose works had taken generations. Although Alexis Carrel is known as the founding father of experimental organ transplantation, many legendary names had contributed to the experimental works of heart transplantation, including Guthrie, Mann, and Demikhov. The major contribution to experimental heart transplantation before the clinical era were made by a team lead by Richard Lower and Norman Shumway at Stanford University in the early 1960s. They played the vital role in developing experimental and clinical heart transplantation as it is known today. Using Shumway biatrial technique Christiaan Barnard started a new era of clinical heart transplantation, by performing the first in man human-to-human heart transplantation in 1967. The techniques of heart transplant have evolved since the first heart transplant. This chapter will summarize the techniques that have been used in clinical heart transplantation

Development and Application of Synthetic Affinity Ligands for the Purification of Ferritin-Based Influenza Antigens

A recently developed novel recombinant influenza antigen vaccine has shown great success in preclinical studies in ferrets and mice. It provides broader protection, and is efficient to manufacture compared to the conventional trivalent influenza vaccines (TIV). Each strain of the recombinant antigen has a constant self-assembled bacterial ferritin core which, if used as a target for affinity chromatography, could lead to a universal purification method. Ferritin in silico models were used to explore potential target binding sites against ligands synthesized by the four-component Ugi reaction. Two ligands, SJ047 and SJ055, were synthesized in solution, characterized by 1H, 13C, and 2D NMR spectroscopy, and subsequently immobilized on the PEG-functionalized beads. Ligands SJ047 and SJ055 displayed apparent Kd values of 2.04 × 10–7 M and 1.91 × 10–8 M, respectively, against the ferritin. SJ047 and SJ055-functionalized resins were able to purify hemagglutinin (New Caledonia)-ferritin expressed in a crude Human Embryonic Kidney (HEK) cell supernatant in a single step to a purity of 85 ± 0.5% (97 ± 1% yield) and 87.5 ± 0.5% (95.5 ± 1.5% yield), respectively. Additionally, SJ047 and SJ055-functionalized resins purified the recombinant antigens when spiked at known concentrations into HEK supernatants. All three strains, hemagglutinin (New Caledonia)-ferritin, hemagglutinin (California)-ferritin, and hemagglutinin (Singapore)-ferritin were purified, thereby offering an ideal alternate platform for affinity chromatography. Following elution from the affinity adsorbents, absorbance at 350 nm showed that there was no aggregation of the recombinant antigens and dynamic light scattering studies further confirmed the structural integrity of the recombinant antigen. The use of Ugi ligands coupled to a PEG-spacer arm to target the ferritin core of the strain is entirely novel and provides an efficient purification of these recombinant antigens. This approach represents a potentially universal method to purify any ferritin-based vaccine

Effect of volcanic dykes on coastal groundwater flow and saltwater intrusion : a field-scale multiphysics approach and parameter evaluation

Acknowledgments This research was primarily based on research grant‐aided by the Irish Department of Communications, Energy and Natural Resources under the National Geoscience Programme 2007–2013. It also benefited from complementary funding from the Scottish Alliance for Geoscience, Environment and Society (SAGES). We acknowledge the contribution in data acquisition of the MSc students in Environmental Engineering at Queen's University Belfast, the landowner for access to the inland fields and the Department of Geography, Archaeology and Paleoecology at QUB for provision of the tidal model of Belfast Lough. The data used are listed in the references, tables, and figures and are available from the corresponding author upon demand. We acknowledge the constructive comments by the Associate Editor and three reviewers, which helped in improving the final manuscript.Peer reviewedPublisher PD

MHC class I loci of the Bar-Headed goose (Anser indicus)

MHC class I proteins mediate functions in anti-pathogen defense. MHC diversity has already been investigated by many studies in model avian species, but here we chose the bar-headed goose, a worldwide migrant bird, as a non-model avian species. Sequences from exons encoding the peptide-binding region (PBR) of MHC class I molecules were isolated from liver genomic DNA, to investigate variation in these genes. These are the first MHC class I partial sequences of the bar-headed goose to be reported. A preliminary analysis suggests the presence of at least four MHC class I genes, which share great similarity with those of the goose and duck. A phylogenetic analysis of bar-headed goose, goose and duck MHC class I sequences using the NJ method supports the idea that they all cluster within the anseriforms clade

Development of a new version of the Liverpool Malaria Model. II. Calibration and validation for West Africa

<p>Abstract</p> <p>Background</p> <p>In the first part of this study, an extensive literature survey led to the construction of a new version of the <it>Liverpool Malaria Model </it>(LMM). A new set of parameter settings was provided and a new development of the mathematical formulation of important processes related to the vector population was performed within the LMM. In this part of the study, so far undetermined model parameters are calibrated through the use of data from field studies. The latter are also used to validate the new LMM version, which is furthermore compared against the original LMM version.</p> <p>Methods</p> <p>For the calibration and validation of the LMM, numerous entomological and parasitological field observations were gathered for West Africa. Continuous and quality-controlled temperature and precipitation time series were constructed using intermittent raw data from 34 weather stations across West Africa. The meteorological time series served as the LMM data input. The skill of LMM simulations was tested for 830 different sets of parameter settings of the undetermined LMM parameters. The model version with the highest skill score in terms of entomological malaria variables was taken as the final setting of the new LMM version.</p> <p>Results</p> <p>Validation of the new LMM version in West Africa revealed that the simulations compare well with entomological field observations. The new version reproduces realistic transmission rates and simulated malaria seasons are comparable to field observations. Overall the new model version performs much better than the original model. The new model version enables the detection of the epidemic malaria potential at fringes of endemic areas and, more importantly, it is now applicable to the vast area of malaria endemicity in the humid African tropics.</p> <p>Conclusions</p> <p>A review of entomological and parasitological data from West Africa enabled the construction of a new LMM version. This model version represents a significant step forward in the modelling of a weather-driven malaria transmission cycle. The LMM is now more suitable for the use in malaria early warning systems as well as for malaria projections based on climate change scenarios, both in epidemic and endemic malaria areas.</p