Explore the vast range of titles available.

The COVID-19 pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Similar to other coronaviruses, its particles are composed of four structural proteins: spike (S), envelope (E), membrane (M), and nucleocapsid (N) proteins. S, E, and M proteins are glycosylated, and the N protein is phosphorylated. The S protein is involved in the interaction with the host receptor human angiotensin-converting enzyme 2 (hACE2), which is also heavily glycosylated. Recent studies have revealed several other potential host receptors or factors that can increase or modulate the SARS-CoV-2 infection. Interestingly, most of these molecules bear carbohydrate residues. While glycans acquired by the viruses through the hijacking of the host machinery help the viruses in their infectivity, they also play roles in immune evasion or modulation. Glycans play complex roles in viral pathobiology, both on their own and in association with carrier biomolecules, such as proteins or glycosaminoglycans (GAGs). Understanding these roles in detail can help in developing suitable strategies for prevention and therapy of COVID-19. In this review, we sought to emphasize the interplay of SARS-CoV-2 glycosylated proteins and their host receptors in viral attachment, entry, replication, and infection. Moreover, the implications for future therapeutic interventions targeting these glycosylated biomolecules are also discussed in detail.

Antigenic characterization of emerging and re-emerging viruses is necessary for the prevention of and response to outbreaks, evaluation of infection mechanisms, understanding of virus evolution, and selection of strains for vaccine development. Primary analytic methods, including enzyme-linked immunosorbent/lectin assays, hemagglutination inhibition, neuraminidase inhibition, micro-neutralization assays, and antigenic cartography, have been widely used in the field of influenza research. These techniques have been improved upon over time for increased analytical capacity, and some have been mobilized for the rapid characterization of the SARS-CoV-2 virus as well as its variants, facilitating the development of highly effective vaccines within 1 year of the initially reported outbreak. While great strides have been made for evaluating the antigenic properties of these viruses, multiple challenges prevent efficient vaccine strain selection and accurate assessment. For influenza, these barriers include the requirement for a large virus quantity to perform the assays, more than what can typically be provided by the clinical samples alone, cell- or egg-adapted mutations that can cause antigenic mismatch between the vaccine strain and circulating viruses, and up to a 6-month duration of vaccine development after vaccine strain selection, which allows viruses to continue evolving with potential for antigenic drift and, thus, antigenic mismatch between the vaccine strain and the emerging epidemic strain. SARS-CoV-2 characterization has faced similar challenges with the additional barrier of the need for facilities with high biosafety levels due to its infectious nature. In this study, we review the primary analytic methods used for antigenic characterization of influenza and SARS-CoV-2 and discuss the barriers of these methods and current developments for addressing these challenges.

Mass mapping using high-resolution mass spectrometry has been applied to identify and rapidly distinguish SARS-CoV-2 coronavirus strains across five major variants of concern. Deletions or mutations within the surface spike protein across these variants, which originated in the UK, South Africa, Brazil and India (known as the alpha, beta, gamma and delta variants respectively), lead to associated mass differences in the mass maps. Peptides of unique mass have thus been determined that can be used to identify and distinguish the variants. The same mass map profiles are also utilized to construct phylogenetic trees, without the need for protein (or gene) sequences or their alignment, in order to chart and study viral evolution. The combined strategy offers advantages over conventional PCR-based gene-based approaches exploiting the ease with which protein mass maps can be generated and the speed and sensitivity of mass spectrometric analysis.

Aluminum-based adjuvants will continue to be a key component of currently approved and next generation vaccines, including important combination vaccines. The widespread use of aluminum adjuvants is due to their excellent safety profile, which has been established through the use of hundreds of millions of doses in humans over many years. In addition, they are inexpensive, readily available, and are well known and generally accepted by regulatory agencies. Moreover, they offer a very flexible platform, to which many vaccine components can be adsorbed, enabling the preparation of liquid formulations, which typically have a long shelf life under refrigerated conditions. Nevertheless, despite their extensive use, they are perceived as relatively ‘weak’ vaccine adjuvants. Hence, there have been many attempts to improve their performance, which typically involves co-delivery of immune potentiators, including Toll-like receptor (TLR) agonists. This approach has allowed for the development of improved aluminum adjuvants for inclusion in licensed vaccines against HPV, HBV, and COVID-19, with others likely to follow. This review summarizes the various aluminum salts that are used in vaccines and highlights how they are prepared. We focus on the analytical challenges that remain to allowing the creation of well-characterized formulations, particularly those involving multiple antigens. In addition, we highlight how aluminum is being used to create the next generation of improved adjuvants through the adsorption and delivery of various TLR agonists.

Current nutritional trends include plant-based diets as nutritional behavior of consumers who are increasingly concerned about a healthy lifestyle. Sea buckthorn (Hippophaë rhamnoides L.) is a plant with great virtues, containing more than 100 types of compounds. It is a plant with versatile properties, multiple economic advantages and a rich history, which still continues in natural medicine, and it is hence included in the daily diet by more and more people for the prevention and treatment of diet-related diseases. Its uniqueness is due to its chemical composition and the health beneficial properties that rise from its composition. This review is a detailed analytical picture of the current state of knowledge currently available regarding the Hippophaë plant, providing an overview of the qualities of sea buckthorn. This article summarizes data on sea buckthorn’s nutritional value, health beneficial properties, and its applications.

Exosomes are extracellular vesicles that share components of their parent cells and are attractive in biotechnology and biomedical research as potential disease biomarkers as well as therapeutic agents. Crucial to realizing this potential is the ability to manufacture high‐quality exosomes; however, unlike biologics such as proteins, exosomes lack standardized Good Manufacturing Practices for their processing and characterization. Furthermore, there is a lack of well‐characterized reference exosome materials to aid in selection of methods for exosome isolation, purification, and analysis. This review informs exosome research and technology development by comparing exosome processing and characterization methods and recommending exosome workflows. This review also provides a detailed introduction to exosomes, including their physical and chemical properties, roles in normal biological processes and in disease progression, and summarizes some of the on‐going clinical trials. The capability to manufacture high quality exosomes is essential for fully realizing their potential as disease biomarkers as well as therapeutic agents. This review informs exosome research and technology development by comparing exosome processing and characterization methods and recommending exosome workflows. The figure shows relative yield, purity, and throughput of different exosome isolation techniques.

The spike protein of SARS-CoV-2, the virus responsible for the global pandemic of COVID-19, is an abundant, heavily glycosylated surface protein that plays a key role in receptor binding and host cell fusion, and is the focus of all current vaccine development efforts. Variants of concern are now circulating worldwide that exhibit mutations in the spike protein. Protein sequence and glycosylation variations of the spike may affect viral fitness, antigenicity, and immune evasion. Global surveillance of the virus currently involves genome sequencing, but tracking emerging variants should include quantitative measurement of changes in site-specific glycosylation as well. In this work, we used data-dependent acquisition (DDA) and data-independent acquisition (DIA) mass spectrometry to quantitatively characterize the five N-linked glycosylation sites of the glycoprotein standard alpha-1-acid glycoprotein (AGP), as well as the 22 sites of the SARS-CoV-2 spike protein. We found that DIA compared favorably to DDA in sensitivity, resulting in more assignments of low-abundance glycopeptides. However, the reproducibility across replicates of DIA-identified glycopeptides was lower than that of DDA, possibly due to the difficulty of reliably assigning low-abundance glycopeptides confidently. The differences in the data acquired between the two methods suggest that DIA outperforms DDA in terms of glycoprotein coverage but that overall performance is a balance of sensitivity, selectivity, and statistical confidence in glycoproteomics. We assert that these analytical and bioinformatics methods for assigning and quantifying glycoforms would benefit the process of tracking viral variants as well as for vaccine development.

The pandemic of the novel coronavirus disease 2019 (COVID-19) has caused severe harm to the health of people all around the world. Molecular detection of the pathogen, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), played a crucial role in the control of the disease. Reverse transcription digital PCR (RT-dPCR) has been developed and used in the detection of SARS-CoV-2 RNA as an absolute quantification method. Here, an interlaboratory assessment of quantification of SARS-CoV-2 RNA was organized by the National Institute of Metrology, China (NIMC), using in vitro transcribed RNA samples, among ten laboratories on six different dPCR platforms. Copy number concentrations of three genes of SARS-CoV-2 were measured by all participants. Consistent results were obtained with dispersion within 2.2-fold and CV% below 23% among different dPCR platforms and laboratories, and Z′ scores of all the reported results being satisfactory. Possible reasons for the dispersion included PCR assays, partition volume, and reverse transcription conditions. This study demonstrated the comparability and applicability of RT-dPCR method for quantification of SARS-CoV-2 RNA and showed the capability of the participating laboratories at SARS-CoV-2 test by RT-dPCR platform.

PurposeTo develop an analytical platform for the estimation as well as characterization of aggregates over the complete size spectrum (from invisible monomer to visible precipitates).MethodsTwo mAb samples were incubated at 30°C in different buffer systems of protein A chromatography for observing degradation due to aggregation. The aggregation in these samples was quantified by size exclusion chromatography (SEC), dynamic light scattering (DLS), and micro flow imaging (MFI).ResultsThe results obtained from various characterization tools were analysed in various size ranges - size exclusion chromatography (SEC) (1 nm - 25 nm), dynamic light scattering (DLS) (10 nm - 5 μm), and micro flow imaging (MFI) (2 μm - 300 μm). Since each characterization tool covers a particular size range, data from multiple tools was collected in the “handover” regions to demonstrate accuracy of the platform.ConclusionsBased on the observations from the experiments, an analytical platform has been proposed covering the whole size spectrum that would be of utility to those engaged in formulation development as well as other aspects related to stability of biotherapeutic products.

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) presents a serious threat to human health all over the world. The development of effective vaccines has been focusing on the spike (S) glycoprotein, which mediates viral invasion to human cells through its interaction with the angiotensin-converting enzyme 2 (ACE2) receptor. In this work, we perform analytical characterization of N- and O-linked glycosylation of the SARS-CoV-2 S glycoprotein. We explore the novel use of dual-functionalized titanium (IV)-immobilized metal affinity chromatography (Ti-IMAC) material for simultaneous enrichment and separation of neutral and sialyl glycopeptides of a recombinant SARS-CoV-2 S glycoprotein from HEK293 cells. This strategy helps eliminate signal suppression from neutral glycopeptides for the detection of sialyl glycopeptides and improves the glycoform coverage of the S protein. We profiled 19 of its 22 potential N-glycosylated sites with 398 unique glycoforms using the dual-functional Ti-IMAC approach, which exhibited improvement of coverage by 1.6-fold compared to the conventional hydrophilic interaction chromatography (HILIC) glycopeptide enrichment method. We also identified O-linked glycosylation site that was not found using the conventional HILIC approach. In addition, we reported on the identification of mannose-6-phosphate (M6P) glycosylation, which substantially expands the current knowledge of the spike protein’s glycosylation landscape and enables future investigation into the influence of M6P glycosylation of the spike protein on its cell entry.