Glycan characterization of the NIST RM monoclonal antibody using a total analytical solution: From sample preparation to data analysis

Glycosylation is an important attribute of biopharmaceutical products to monitor from development through production. However, glycosylation analysis has traditionally been a time-consuming process with long sample preparation protocols and manual interpretation of the data. To address the challenges associated with glycan analysis, we developed a streamlined analytical solution that covers the entire process from sample preparation to data analysis. In this communication, we describe the complete analytical solution that begins with a simplified and fast N-linked glycan sample preparation protocol that can be completed in less than 1 hr. The sample preparation includes labelling with RapiFluor-MS tag to improve both fluorescence (FLR) and mass spectral (MS) sensitivities. Following HILIC-UPLC/FLR/MS analyses, the data are processed and a library search based on glucose units has been included to expedite the task of structural assignment. We then applied this total analytical solution to characterize the glycosylation of the NIST Reference Material mAb 8761. For this glycoprotein, we confidently identified 35 N-linked glycans and all three major classes, high mannose, complex, and hybrid, were present. The majority of the glycans were neutral and fucosylated; glycans featuring N-glycolylneuraminic acid and those with two galactoses connected via an α1,3-linkage were also identified.     

Experimental investigations of the human oesophagus: anisotropic properties of the embalmed mucosa–submucosa layer under large deformation

Mechanical characterisation of the layer-specific, viscoelastic properties of the human oesophagus is crucial in furthering the development of devices emerging in the field, such as robotic endoscopic biopsy devices, as well as in enhancing the realism, and therefore effectiveness, of surgical simulations. In this study, the viscoelastic and stress-softening behaviour of the passive human oesophagus was investigated through ex vivo cyclic mechanical tests. Due to restrictions placed on the laboratory as a result of COVID-19, only oesophagi from cadavers fixed in formalin were allowed for testing. Three oesophagi in total were separated into their two main layers and the mucosa–submucosa layer was investigated. A series of uniaxial tensile tests were conducted in the form of increasing stretch level cyclic tests at two different strain rates: 1% s\(^{-1}\) and 10% s\(^{-1}\). Rectangular samples in both the longitudinal and circumferential directions were tested to observe any anisotropy. Histological analysis was also performed through a variety of staining methods. Overall, the longitudinal direction was found to be much stiffer than the circumferential direction. Stress-softening was observed in both directions, as well as permanent set and hysteresis. Strain rate-dependent behaviour was also apparent in the two directions, with an increase in strain rate resulting in an increase in stiffness. This strain rate dependency was more pronounced in the longitudinal direction than the circumferential direction. Finally, the results were discussed in regard to the histological content of the layer, and the behaviour was modelled and validated using a visco-hyperelastic matrix-fibre model.

     

The use of geographic information systems to map and assess ecosystem services

In recent years, geographic information systems (GIS) have become a powerful tool for mapping and assessing the provision of ecosystem services within a landscape. GIS can help land managers and conservationists visualize spatial and temporal patterns and changes in ecosystem services and estimate the potential impact from projected changes in land use or management or climatic conditions on the provision of these services. The end-goal of ecosystem service assessment is usually to estimate marginal values of ecosystem services to inform decisions where trade-offs in ecosystem service provision will affect human well-being. Because our ability to estimate the provision of ecosystem services underlies our ability to estimate their societal values, the theoretical bases of GIS approaches and models for assessing ecosystem services need to be well understood before they are employed for decision-making purposes. This paper reviews GIS approaches and software developed for the assessment of ecosystem services and highlights their strengths and weaknesses in the context of different end uses.     

Natural genetic variation drives microbiome selection in the Caenorhabditis elegans gut

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Leishmania mexicana Induces Limited Recruitment and Activation of Monocytes and Monocyte-Derived Dendritic Cells Early during Infection

While C57BL/6 mice infected in the ear with L. major mount a vigorous Th1 response and resolve their lesions, the Th1 response in C57BL/6 mice infected with L. mexicana is more limited, resulting in chronic, non-healing lesions. The aim of this study was to determine if the limited immune response following infection with L. mexicana is related to a deficiency in the ability of monocyte-derived dendritic cells (mo-DCs) to prime a sufficient Th1 response. To address this issue we compared the early immune response following L. mexicana infection with that seen in L. major infected mice. Our data show that fewer monocytes are recruited to the lesions of L. mexicana infected mice as compared to mice infected with L. major. Moreover, monocytes that differentiate into mo-DCs in L. mexicana lesions produced less iNOS and migrated less efficiently to the draining lymph node as compared to those from L. major infected mice. Treatment of L. mexicana infected mice with α-IL-10R antibody resulted in increased recruitment of monocytes to the lesion along with greater production of IFN-γ and iNOS. Additionally, injection of DCs into the ear at the time of infection with L. mexicana also led to a more robust Th1 response. Taken together, these data suggest that during L. mexicana infection reduced recruitment, activation and subsequent migration of monocytes and mo-DCs to the draining lymph nodes may result in the insufficient priming of a Th1 response.