Emergence of the novel pathogenic coronavirus SARS‐CoV‐2 and its rapid pandemic spread presents challenges that demand immediate attention. Here, we describe the development of a semi‐quantitative high‐content microscopy‐based assay for detection of three major classes (IgG, IgA, and IgM) of SARS‐CoV‐2 specific antibodies in human samples. The possibility to detect antibodies against the entire viral proteome together with a robust semi‐automated image analysis workflow resulted in specific, sensitive and unbiased assay that complements the portfolio of SARS‐CoV‐2 serological assays. (...) Sensitive, specific and quantitative serological assays are urgently needed for a better understanding of humoral immune response against the virus as a basis for developing public health strategies to control viral spread. The procedure described here has been used for clinical studies and provides a general framework for the application of quantitative high‐throughput microscopy to rapidly develop serological assays for emerging virus infections. (shrink)

Pointillistic based super‐resolution techniques, such as photoactivated localization microscopy (PALM), involve multiple cycles of sequential activation, imaging, and precise localization of single fluorescent molecules. A super‐resolution image, having nanoscopic structural information, is then constructed by compiling all the image sequences. Because the final image resolution is determined by the localization precision of detected single molecules and their density, accurate image reconstruction requires imaging of biological structures labeled with fluorescent molecules at high density. In such image datasets, stochastic variations in (...) photon emission and intervening dark states lead to uncertainties in identification of single molecules. This, in turn, prevents the proper utilization of the wealth of information on molecular distribution and quantity. A recent strategy for overcoming this problem is pair‐correlation analysis applied to PALM. Using rigorous statistical algorithms to estimate the number of detected proteins, this approach allows the spatial organization of molecules to be quantitatively described. (shrink)

Fluorescence microscopy has provided a route to qualitatively analyze features of nuclear structures and chromatin domains with increasing resolution. However, it is becoming increasingly important to develop tools for quantitative analysis. Here, we present an automated method to quantitatively determine the enrichment of several endogenous factors, immunostained in pericentric heterochromatin domains in mouse cells. We show that this method permits an unbiased characterization of changes in the enrichment of several factors with statistical significance from a large number of (...) nuclei. Furthermore, the nuclei can be sorted according to the enrichment value of these factors. This method should prove useful to monitor events related to changes in the amount, rather than the presence or absence, of any factor. By adapting a few parameters, it could be extended to other nuclear structures and the benefit of using available software will permit its use in many biological labs. (shrink)

New imaging methodologies in quantitative fluorescence microscopy, such as Förster resonance energy transfer (FRET), have been developed in the last few years and are beginning to be extensively applied to biological problems. FRET is employed for the detection and quantification of protein interactions, and of biochemical activities. Herein, we review the different methods to measure FRET in microscopy, and more importantly, their strengths and weaknesses. In our opinion, fluorescence lifetime imaging microscopy (FLIM) is advantageous for detecting (...) inter‐molecular interactions quantitatively, the intensity ratio approach representing a valid and straightforward option for detecting intra‐molecular FRET. Promising approaches in single molecule techniques and data analysis for quantitative and fast spatio‐temporal protein‐protein interaction studies open new avenues for FRET in biological research. (shrink)

Phototoxicity frequently occurs during live fluorescence microscopy, and its consequences are often underestimated. Damage to cellular macromolecules upon excitation light illumination can impair sample physiology, and even lead to sample death. In this review, we explain how phototoxicity influences live samples, and we highlight that, besides the obvious effects of phototoxicity, there are often subtler consequences of illumination that are imperceptible when only the morphology of samples is examined. Such less apparent manifestations of phototoxicity are equally problematic, and can (...) change the conclusions drawn from an experiment. Thus, limiting phototoxicity is a prerequisite for obtaining reproducible quantitative data on biological processes. We present strategies to reduce phototoxicity, e.g. limiting the illumination to the focal plane and suggest controls for phototoxicity effects. Overall, we argue that phototoxicity needs increased attention from researchers when designing experiments, and when evaluating research findings. (shrink)

Recently developed super‐resolution microscopy techniques are changing our understanding of lipid rafts and membrane organisation in general. The lipid raft hypothesis postulates that cholesterol can drive the formation of ordered domains within the plasma membrane of cells, which may serve as platforms for cell signalling and membrane trafficking. There is now a wealth of evidence for these domains. However, their study has hitherto been hampered by the resolution limit of optical microscopy, making the definition of their properties problematic (...) and contentious. New microscopy techniques circumvent the resolution limit and, for the first time, allow the fluorescence imaging of structures on length scales below 200 nm. This review describes such techniques, particularly as applied to the study of membrane organisation, synthesising newly emerging facets of lipid raft biology into a state‐of‐the art model.Editor's suggested further reading in BioEssays: Super‐resolution imaging prompts re‐thinking of cell biology mechanisms Abstract and Quantitative analysis of photoactivated localization microscopy (PALM) datasets using pair‐correlation analysis Abstract. (shrink)

Registration of a series of the two-dimensional electron microscope images of the brain tissue into volumetric form is an important technique that can be used for neuronal circuit reconstruction. However, complex appearance changes of neuronal morphology in adjacent sections bring difficulty in finding correct correspondences, making serial section neural image registration challenging. To solve this problem, we consider whether there are such stable "markers" in the neural images to alleviate registration difficulty. In this paper, we employ the spherical deformation model (...) to simulate the local neuron structure and analyze the relationship between registration accuracy and neuronal structure shapes in two adjacent sections. The relevant analysis proves that regular circular structures in the section images are instrumental in seeking robust corresponding relationships. Then, we design a new serial section image registration framework driven by this neuronal morphological model, fully utilizing the characteristics of the anatomical structure of nerve tissue and obtaining more reasonable corresponding relationships. Specifically, we leverage a deep membrane segmentation network and neural morphological physical selection model to select the stable rounded regions in neural images. Then, we combine feature extraction and global optimization of correspondence position to obtain the deformation field of multiple images. Experiments on real and synthetic serial EM section neural image datasets have demonstrated that our proposed method could achieve more reasonable and reliable registration results, outperforming the state-of-the-art approaches in qualitative and quantitative analysis. (shrink)

Macromolecular interactions play a central role in many biological processes. Protein‐protein interactions have mostly been studied by co‐immunoprecipitation, which cannot provide quantitative information on all possible molecular connections present in the complex. We will review a new approach that allows cellular proteins and biomolecular complexes to be studied in real‐time at the single‐molecule level. This technique is called single‐molecule pull‐down (SiMPull), because it integrates principles of conventional immunoprecipitation with the powerful single‐molecule fluorescence microscopy. SiMPull is used to count (...) how many of each protein is present in the physiological complexes found in cytosol and membranes. Concurrently, it serves as a single‐molecule biochemical tool to perform functional studies on the pulled‐down proteins. In this review, we will focus on the detailed methodology of SiMPull, its salient features and a wide range of biological applications in comparison with other biosensing tools. (shrink)

The transport of messenger RNAs (mRNAs) from the nucleus to cytoplasm is an essential step in the gene expression program of all eukaryotes. Recent technological advances in the areas of RNA‐labeling, microscopy, and sequencing are leading to novel insights about mRNA biogenesis and export. This includes quantitative single molecule imaging (SMI) of RNA molecules in live cells, which is providing knowledge of the spatial and temporal dynamics of the export process. As this information becomes available, it leads to (...) new questions, the reinterpretation of previous findings, and revised models of mRNA export. In this review, we will briefly highlight some of these recent findings and discuss how live cell SMI approaches may be used to further our current understanding of mRNA export and gene expression. (shrink)

Although negative stain electron microscopy is a wonderfully simple way of directly visualizing protein complexes and other biological macromolecules, the images are not really comparable to those of objects seen in everyday life. The failure to appreciate this has recently led to an incorrect interpretation of RecA‐family filament structures.

In Amerindian ontologies, hallucinations or visions, rather than being dismissed as delusions or symbolic constructs, are recognized as means of perceptual access to physical reality. Lowland South American shamans claim to be able to diagnose and treat infectious diseases, and to assess the status of wildlife resources through interactions with pathogenic agents perceived in visions. This essay examines some perceptual capabilities that shamans might be employing to explore their physical reality. The structure of the eye affords a form of (...) class='Hi'>microscopy of retinal structures and of objects flowing within them, including cells and microbial agents during systemic infection. Lowland South American shamanic practices involve optical and physiological conditions that optimize entoptic microscopy. Images of those visions display the characteristic features of shadow formation, confirming their microscopic origin. This phenomenological access to the microscopic world and similarities with the panorama depicted by current microbiology indicate the commensurability of these forms of knowledge. (shrink)

This paper deals with an analysis of debris produced during the polishing of diamond. The debris is carefully collected 'as ejected' to shorten the history of the freshly removed material. Using high-resolution electron microscopy as well as electron-energy-loss spectroscopy, the structure of the material is revealed and analysed in terms of density, percentage of sp 2 hybridized carbon, and oxygen content. Debris from polishing in the so-called hard and soft directions were involved in this investigation. Overall the structure of (...) all debris is amorphous carbon. The material appears to be composed of small clusters, some nanometres in diameter, in which the graphite basal planes can be recognized. Very few and very small nanometre-sized diamond particles were found in the debris from polishing in the hard direction. The results support a polishing mechanism based on a mechanically induced transformation of diamond to graphite, after which material removal easily occurs. The well-known anisotropy observed in polishing can be explained satisfactorily on the basis of this model. Finally, in appendices, the art of polishing and the role of the black powder during preparation of the scaife are discussed. (shrink)

This paper challenges the common assumption that some phenotypic traits are quantitative while others are qualitative. The distinction between these two kinds of traits is widely influential in biological and biomedical research as well as in scientific education and communication. This is probably due to both historical and epistemological reasons. However, the quantitative/qualitative distinction involves a variety of simplifications on the genetic causes of phenotypic variability and on the development of complex traits. Here, I examine three cases from (...) the life sciences that show inconsistencies in the distinction: Mendelian traits, Mendelian diseases, and polygenic mental disorders. I show that these traits can be framed both quantitatively and qualitatively depending, for instance, on the methods through which they are investigated and on specific epistemic purposes. This suggests that the received view of quantitative and qualitative traits has a limited heuristic power—limited to some local contexts or to the specific methodologies adopted. Throughout the paper, I provide directions for framing phenotypes beyond the quantitative/qualitative distinction. I conclude by pointing at the necessity of developing a principled characterisation of what phenotypic traits, in general, are. (shrink)

SummaryThis paper analyses the development of three methods for detecting bloodstains during the first half of the nineteenth-century in France. After dealing with the main problems in detecting bloodstains, the paper describes the chemical tests introduced in the mid-1820s. Then the first uses of the microscope in the detection of bloodstains around 1827 are discussed. The most controversial method is then examined, the smell test introduced by Jean-Pierre Barruel in 1829, and the debates which took place in French academies and (...) learned societies during ensuing years are surveyed. Moving to the courtrooms a review is conducted of how the different methods were employed in criminal trials. By reviewing these cases, the main arguments against Barruel's test during the 1830s are explored as well as the changes making possible the return of the microscope to legal medicine around 1840. By reconstructing the history of these three methods, the paper reveals how the senses of smell and vision were employed in order to produce convincing evidence in both academies and courts. The paper questions two linear master narratives that are organized in terms of progress and decline: the development of forensic science as a result of continued technological progress; and the supposed decline of smell in the history of the senses, particularly in the realm of chemistry and medicine. (shrink)

The desire to minimize the number of individual new entities postulated is often referred to as quantitative parsimony. Its influence on the default hypotheses formulated by scientists seems undeniable. I argue that there is a wide class of cases for which the preference for quantitatively parsimonious hypotheses is demonstrably rational. The justification, in a nutshell, is that such hypotheses have greater explanatory power than less parsimonious alternatives. My analysis is restricted to a class of cases I shall refer to (...) as additive. Such cases involve the postulation of a collection of qualitatively identical individual objects which collectively explain some particular observed phenomenon. Especially clear examples of this sort occur in particle physics. 1 Introduction 2 Particle physics: a case study 3 Three kinds of simplicity 4 Explanatory power 5 Explanation and non-observation 6 Parsimony and scientific methodology 7 Conclusions. (shrink)

Two grams mass, three coulombs charge, five inches long – these are examples of quantitative properties. Quantitative properties have certain structural features that other sorts of properties lack. What are the metaphysical underpinnings of quantitative structure? This paper considers several accounts of quantity and assesses the merits of each.

Fluorescence microscopy is the primary tool for studying complex processes inside individual living cells. Technical advances in both molecular biology and microscopy have made it possible to image cells from many genetic and environmental backgrounds. These images contain a vast amount of information, which is often hidden behind various sources of noise, convoluted with other information and stochastic in nature. Accessing the desired biological information therefore requires new tools of computational image analysis and modeling. Here, we review some (...) of the recent advances in computational analysis of images obtained from fluorescence microscopy, focusing on bacterial systems. We emphasize techniques that are readily available to molecular and cell biologists but also point out examples where problem‐specific image analyses are necessary. Thus, image analysis is not only a toolkit to be applied to new images but also an integral part of the design and implementation of a microscopy experiment. (shrink)

In this paper, I motivate the view that quantitative parsimony is a theoretical virtue: that is, we should be concerned not only to minimize the number of kinds of entities postulated by our theories (i. e. maximize qualitative parsimony), but we should also minimize the number of entities postulated which fall under those kinds. In order to motivate this view, I consider two cases from the history of science: the postulation of the neutrino and the proposal of Avogadro's hypothesis. (...) I also consider two issues concerning how a principle of quantitative parsimony should be framed. (shrink)

In very recent Leibniz commentary, there has been a movement among some commentators toward the view that Leibniz was not an unwavering monadological immaterialist, committed to the substantiality only of absolutely simple, immaterial nodes of perception and appetite. It has been conceded that Leibniz was also partially sympathetic to an ontology that would concede full substantiality to composite entities. Most commentators who have been willing to concede as much have confined this alternative metaphysics to Leibniz's middle period, roughly 1676--1694. These (...) commentators, moreover, have remained fairly silent as to the reasons why Leibniz might wish to consider an ontology at such odds with his well-known immaterialism. In this dissertation, I contribute to the discussion of Leibniz's composite-substance metaphysics in three ways. First of all, I am going to argue that we should see the composite-substance metaphysics as an alternative Leibniz sustained until the very end of his career. Second of all, I provide an account of Leibniz's motivation for sustaining this alternative metaphysics, arguing that the composite-substance metaphysics is first and foremost the result of Leibniz's effort to provide a philosophical interpretation of the recent discovery of the apparent ubiquity of microscopic life. Finally, I vindicate a number of 18th-century, microorganic and physicalistic interpretations of Leibniz's theory of monads, particularly those of the Francophone natural scientists, against the view among 20th-century commentators that Leibniz was grossly misinterpreted in the decades following his death in 1716. I show that 18th-century, thinkers didn't so much misinterpret Leibniz, as they did emphasize one, currently very unfamiliar, tendency in Leibniz's thought on the nature of substance. (shrink)

An in-situ transmission electron microscopy straining technique has been used to investigate the dynamics of dislocation-defect interactions in ion-irradiated copper and the subsequent formation of defect-free channels. Defect removal frequently required interaction with multiple dislocations, although screw dislocations were more efficient at annihilating defects than edge dislocations were. The defect pinning strength was determined from the dislocation curvature prior to breakaway and exhibited values ranging from 15 to 175 MPa. Pre-existing dislocations percolated through the defect field but did not (...) show long-range motion, indicating that they are not responsible for creating the defect-free channels and have a limited contribution to the total plasticity. Defect-free channels were associated with the movement of many dislocations, which originated from grain boundaries or regions of high stress concentration such as at a crack tip. These experimental results are compared with atomistic simulations of the interaction of partial dislocations with defects in copper and a dispersed-barrier-hardening crystal plasticity model to correlate the observations to bulk mechanical properties. (shrink)

In this article, the co-editors of the corporate responsibility: quantitative issues section of the journal provide an overview of the quantitative CSR field and offer some new perspectives on where the field is going. They highlight key issues in developing impactful, theory-driven, and ethically grounded research and call for research that examines complex problems facing businesses and the society (e.g., big data and artificial intelligence, political polarization, and the role of CSR in generating social impact). By examining topics (...) that are under-researched, forward-looking, and socially oriented, scholars can expand the boundary of CSR’s substantive domain and produce research that helps businesses act in a long-term, socially responsible way in this quickly evolving, turbulent environment. They also discuss ways to enhance the methodological rigor of quantitative CSR research and encourage scholars to employ cutting-edge, innovative methods to shed light on the micro-level mechanisms of CSR and reveal patterns and relationships hidden in unstructured big data. (shrink)

ABSTRACT Our aim in this article is to offer a new justification for preferring theories that are more quantitatively parsimonious than their rivals. We discuss cases where it seems clear that those involved opted for more quantitatively parsimonious theories. We extend previous work on quantitative parsimony by offering an independent probabilistic justification for preferring the more quantitatively parsimonious theories in particular episodes of theory choice. Our strategy allows us to avoid worries that other considerations, such as pragmatic factors of (...) computational tractability and so on, could be the driving ones in the historical cases under consideration. _1_ Introduction _2_ Three Desiderata _2.1_ Limiting _2.2_ Robustness _2.3_ Breadth _2.3.1_ A limited success for Baker _2.3.2_ Rejecting Baker’s analysis _2.4_ The proposal _3_ Probabilistically Additive Hypotheses and a Bayesian Account: The Limpid Rationale Relativized and Reconsidered _3.1_ Neutrinos and beta decay _3.2_ Avogadro’s hypothesis _3.3_ Postulation of Neptune _4_ Conclusion. (shrink)

The current revolution in biological microscopy stems from the realisation that advances in optics and computational tools and automation make the modern microscope an instrument that can access all scales relevant to modern biology – from individual molecules all the way to whole tissues and organisms and from single snapshots to time‐lapse recordings sampling from milliseconds to days. As these and more new technologies appear, the challenges of delivering them to the community grows as well. I discuss some of (...) these challenges, and the examples where openly shared technology have made an impact on the field. (shrink)

We chart the ways in which closure properties of consequence relations for uncertain inference take on different forms according to whether the relations are generated in a quantitative or a qualitative manner. Among the main themes are: the identification of watershed conditions between probabilistically and qualitatively sound rules; failsafe and classicality transforms of qualitatively sound rules; non-Horn conditions satisfied by probabilistic consequence; representation and completeness problems; and threshold-sensitive conditions such as `preface' and `lottery' rules.

Microscopy allows us to observe objects we cannot see with our eyes alone. With a light microscope, we can distinguish objects at the scale of the wavelengths of visible light just under a micrometer. Around 1870 Ernst Abbe, who laid the foundation of modern optics, suggested that the resolution of a microscope would improve by using some yet-unknown radiation with shorter wavelengths than visible light, that is, below 390 nanometers (1 nm = 10−9 m). Electrons can have wavelengths near (...) 1 picometer (1 pm = 10−12 m) and should therefore allow atoms to be distinguished since they are typically at least a few hundred pm apart. In this oversimplified view, further decreasing the wavelength of the radiation should allow us to increase the resolution of electron microscopy even more. However, this can only be done by increasing the energy of the radiation, which would ultimately destroy the samples. Other factors also affect the resolution of electron microscopes, among them non-ideal imaging properties of electromagnetic lenses, which result in false images. It took more than half a century to understand and control these aberrations and separate objects less than 1 Ångstrom (1 Å = 10−10 m) apart. (shrink)