A high content microscopy-based platform for detecting antibodies to the nucleocapsid, spike and membrane proteins of SARS-CoV-2

The strong humoral immune response produced against the SARS-CoV-2 nucleocapsid (N) and spike (S) proteins has underpinned serological testing but the prevalence of antibody responses to other SARS-CoV-2 proteins, which may be of use as further serological markers, is still unclear. Cell-based serological screening platforms can fulfil a crucial niche in the identification of antibodies which recognise more complex folded epitopes or those incorporating post-translation modifications which may be undetectable by other methods used to investigate the antigenicity of the SARS-CoV-2 proteome. Here, we employed automated high content immunofluorescence microscopy (AHCIM) to assess the viability of such an approach as a method capable of assaying humoral immune responses against full length SARS-CoV-2 proteins in their native cellular state. We first demonstrate that AHCIM provides high sensitivity and specificity in the detection of SARS-CoV-2 N and S IgG. Assessing the prevalence of antibody responses to the SARS-CoV-2 structural membrane protein (M), we further find that 85% of COVID-19 patients within our sample set developed detectable M IgG responses (M sensitivity 85%, N sensitivity 93%, combined N + M sensitivity 95%). The identification of M as a serological marker of high prevalence may be of value in detecting additional COVID-19 cases during the era of mass SARS-CoV-2 vaccinations, where serological screening for SARS CoV-2 infections in vaccinated individuals is dependent on detection of antibodies against N. These findings highlight the advantages of using cell-based systems as serological screening platforms and raise the possibility of using M as a widespread serological marker alongside N and S.

Protein Appetite Drives Macronutrient-Related Differences in Ventral Tegmental Area Neural Activity.

Control of protein intake is essential for numerous biological processes as several amino acids cannot be synthesized de novo, however, its neurobiological substrates are still poorly understood. In the present study, we combined in vivo fiber photometry with nutrient-conditioned flavor in a rat model of protein appetite to record neuronal activity in the VTA, a central brain region for the control of food-related processes. In adult male rats, protein restriction increased preference for casein (protein) over maltodextrin (carbohydrate). Moreover, protein consumption was associated with a greater VTA response, relative to carbohydrate. After initial nutrient preference, a switch from a normal balanced diet to protein restriction induced rapid development of protein preference but required extensive exposure to macronutrient solutions to induce elevated VTA responses to casein. Furthermore, prior protein restriction induced long-lasting food preference and VTA responses. This study reveals that VTA circuits are involved in protein appetite in times of need, a crucial process for animals to acquire an adequate amount of protein in their diet.SIGNIFICANCE STATEMENT Acquiring insufficient protein in one's diet has severe consequences for health and ultimately will lead to death. In addition, a low level of dietary protein has been proposed as a driver of obesity as it can leverage up intake of fat and carbohydrate. However, much remains unknown about the role of the brain in ensuring adequate intake of protein. Here, we show that in a state of protein restriction a key node in brain reward circuitry, the VTA, is activated more strongly during consumption of protein than carbohydrate. Moreover, although rats' behavior changed to reflect new protein status, patterns of neural activity were more persistent and only loosely linked to protein status.

Limited Aggregation and E-Cigarettes.

INTRODUCTION: Nonconsequentialist ethicists have noted that small harms, goods, or claims should not count against large claims. For example, given a choice between saving one life and a large group of people with minor headaches, we ought to save the one life, no matter how large the group is. This principle has been called limited aggregation. The principle of limited aggregation might have implications on public health policy, given that public health policy involves weighing the claims of individuals against one another. AIMS AND METHODS: I aim to show that limited aggregation has implications for policies on e-cigarettes and alternative nicotine delivery systems. The methodology used in this study involves critical analysis of existing literature and pre-theoretical moral views. RESULTS: This study does not use empirical research. CONCLUSIONS: In deciding to allow or encourage the use of e-cigarettes or alternative nicotine delivery systems, we sometimes must weigh benefits to each existing smoker affected by the policy against risks to each nonsmoker affected. I argue in this paper that when these risks, to each individual nonsmoker, are sufficiently small, we ought not to count them against more significant benefits to smokers. This applies even when the number of nonsmokers affected by a policy exceeds the number of smokers. IMPLICATIONS: This paper implies that policymakers ought to be sensitive to the scale of benefits or risks introduced by a policy on individuals. If the negative side effects, on each affected individual, of a proposed policy are sufficiently small, they do not count against the beneficial goals of that policy. Depending on the expected effects of each given e-cigarette policy, this may give policymakers defeasible reasons to prioritize the needs of current smokers, who may each gain a lot from various means of smoking cessation, over nonsmokers, who each may only have a small chance of picking up smoking.

The role of dopamine in the pursuit of nutritional value.

Acquiring enough food to meet energy expenditure is fundamental for all organisms. Thus, mechanisms have evolved to allow foods with high nutritional value to be readily detected, consumed, and remembered. Although taste is often involved in these processes, there is a wealth of evidence supporting the existence of taste-independent nutrient sensing. In particular, post-ingestive mechanisms arising from the arrival of nutrients in the gut are able to drive food intake and behavioural conditioning. The physiological mechanisms underlying these effects are complex but are believed to converge on mesolimbic dopamine signalling to translate post-ingestive sensing of nutrients into reward and reinforcement value. Discerning the role of nutrition is often difficult because food stimulates sensory systems and post-ingestive pathways in concert. In this mini-review, I discuss the various methods that may be used to study post-ingestive processes in isolation including sham-feeding, non-nutritive sweeteners, post-ingestive infusions, and pharmacological and genetic methods. Using this structure, I present the evidence that dopamine is sensitive to nutritional value of certain foods and examine how this affects learning about food, the role of taste, and the implications for human obesity.

Age matters.

The age of an experimental animal can be a critical variable, yet age matters are often overlooked within neuroscience. Many studies make use of young animals, without considering possible differences between immature and mature subjects. This is especially problematic when attempting to model traits or diseases that do not emerge until adulthood. In this commentary we discuss the reasons for this apparent bias in age of experimental animals, and illustrate the problem with a systematic review of published articles on long-term potentiation. Additionally, we review the developmental stages of a rat and discuss the difficulty of using the weight of an animal as a predictor of its age. Finally, we provide original data from our laboratory and review published data to emphasize that development is an ongoing process that does not end with puberty. Developmental changes can be quantitative in nature, involving gradual changes, rapid switches, or inverted U-shaped curves. Changes can also be qualitative. Thus, phenomena that appear to be unitary may be governed by different mechanisms at different ages. We conclude that selection of the age of the animals may be critically important in the design and interpretation of neurobiological studies.