Towards defining reference materials for extracellular vesicle size, concentration, refractive index and epitope abundance

Accurate characterization of extracellular vesicles (EVs) is critical to explore their diagnostic and therapeutic applications. As the EV research field has developed, so too have the techniques used to characterize them. The development of reference materials is required for the standardization of these techniques. This work, initiated from the ISEV 2017 Biomarker Workshop in Birmingham, UK, and with further discussion during the ISEV 2019 Standardization Workshop in Ghent, Belgium, sets out to elucidate which reference materials are required and which are currently available to standardize commonly used analysis platforms for characterizing EV size, concentration, refractive index, and epitope expression. Due to their predominant use, a particular focus is placed on the optical methods nanoparticle tracking analysis and flow cytometry.Comment: 30 pages, 6 figures, 2 table

A compendium of single extracellular vesicle flow cytometry

Flow cytometry (FCM) offers a multiparametric technology capable of characterizing single extracellular vesicles (EVs). However, most flow cytometers are designed to detect cells, which are larger than EVs. Whereas cells exceed the background noise, signals originating from EVs partly overlap with the background noise, thereby making EVs more difficult to detect than cells. This technical mismatch together with complexity of EV-containing fluids causes limitations and challenges with conducting, interpreting and reproducing EV FCM experiments. To address and overcome these challenges, researchers from the International Society for Extracellular Vesicles (ISEV), International Society for Advancement of Cytometry (ISAC), and the International Society on Thrombosis and Haemostasis (ISTH) joined forces and initiated the EV FCM working group. To improve the interpretation, reporting, and reproducibility of future EV FCM data, the EV FCM working group published an ISEV position manuscript outlining a framework of minimum information that should be reported about an FCM experiment on single EVs (MIFlowCyt-EV). However, the framework contains limited background information. Therefore, the goal of this compendium is to provide the background information necessary to design and conduct reproducible EV FCM experiments. This compendium contains background information on EVs, the interaction between light and EVs, FCM hardware, experimental design and preanalytical procedures, sample preparation, assay controls, instrument data acquisition and calibration, EV characterization, and data reporting. Although this compendium focuses on EVs, many concepts and explanations could also be applied to FCM detection of other particles within the EV size range, such as bacteria, lipoprotein particles, milk fat globules, and viruses

Learning homophones in context:Easy cases are favored in the lexicon of natural languages

International audienc

Minimal information for studies of extracellular vesicles 2018 (MISEV2018):a position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines

The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles (“MISEV”) guidelines for the field in 2014. We now update these “MISEV2014” guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points

Investigating possible causes of bias in a progress test translation: an one-edged sword

PURPOSE: Assessment in different languages should measure the same construct. However, item characteristics, such as item flaws and content, may favor one test-taker group over another. This is known as item bias. Although some studies have focused on item bias, little is known about item bias and its association with items characteristics. Therefore, this study investigated the association between item characteristics and bias. METHODS: The University of Groningen offers both an international and a national bachelor's program in medicine. Students in both programs take the same progress test, but the international progress test is literally translated into English from the Dutch version. Differential item functioning was calculated to analyze item bias in four subsequent progress tests. Items were also classified by their categories, number of alternatives, item flaw, item length, and whether it was a case-based question. RESULTS: The proportion of items with bias ranged from 34% to 36% for the various tests. The number of items and the size of their bias was very similar in both programmes. We have identified that the more complex items with more alternatives favored the national students, whereas shorter items and fewer alternatives favored the international students. CONCLUSION: Although nearly 35% of all items contain bias, the distribution and the size of the bias were similar for both groups. The findings of this paper may be used to improve the writing process of the items, by avoiding some characteristics that may benefit one group whilst being a disadvantage for others

C-HEAP: A Heterogeneous Multi-processor Architecture Template and Scalable and Flexible Protocol for the Design of Embedded Signal Processing Systems

The key issue in the design of Systems-on-a-Chip (SoC) is to trade-off efficiency against flexibility, and time to market versus cost. Current deep submicron processing technologies enable integration of multiple software programmable processors (e.g. CPUs, DSPs) and dedicated hardware components into a single cost-efficient IC. Our top-down design methodology with various abstraction levels helps designing these ICs in a reasonable amount of time. This methodology starts with a high-level executable specification, and converges towards a silicon implementation. A major task in the design process is to ensure that all components (hardware and software) communicate with each other correctly. In this article, we tackle this problem in the context of the signal processing domain in two ways: we propose a modular, flexible, and scalable heterogeneous multi-processor architecture template based on distributed shared memory, and we present an efficient and transparent protocol for communication and (re)configuration. The protocol implementations have been incorporated in libraries, which allows quick traversal of the various abstraction levels, so enabling incremental design. The design decisions to be taken at each abstraction level are evaluated by means of (co-)simulation. Prototyping is used too, to verify the system's functional correctness. The effectiveness of our approach is illustrated by a design case of a multi-standard video and image codec

MIFlowCyt-EV: a framework for standardized reporting of extracellular vesicle flow cytometry experiments

Extracellular vesicles (EVs) are small, heterogeneous and difficult to measure. Flow cytometry (FC) is a key technology for the measurement of individual particles, but its application to the analysis of EVs and other submicron particles has presented many challenges and has produced a number of controversial results, in part due to limitations of instrument detection, lack of robust methods and ambiguities in how data should be interpreted. These complications are exacerbated by the field's lack of a robust reporting framework, and many EV-FC manuscripts include incomplete descriptions of methods and results, contain artefacts stemming from an insufficient instrument sensitivity and inappropriate experimental design and lack appropriate calibration and standardization. To address these issues, a working group (WG) of EV-FC researchers from ISEV, ISAC and ISTH, worked together as an EV-FC WG and developed a consensus framework for the minimum information that should be provided regarding EV-FC. This framework incorporates the existing Minimum Information for Studies of EVs (MISEV) guidelines and Minimum Information about a FC experiment (MIFlowCyt) standard in an EV-FC-specific reporting framework (MIFlowCyt-EV) that supports reporting of critical information related to sample staining, EV detection and measurement and experimental design in manuscripts that report EV-FC data. MIFlowCyt-EV provides a structure for sharing EV-FC results, but it does not prescribe specific protocols, as there will continue to be rapid evolution of instruments and methods for the foreseeable future. MIFlowCyt-EV accommodates this evolution, while providing information needed to evaluate and compare different approaches. Because MIFlowCyt-EV will ensure consistency in the manner of reporting of EV-FC studies, over time we expect that adoption of MIFlowCyt-EV as a standard for reporting EV- FC studies will improve the ability to quantitatively compare results from different laboratories and to support the development of new instruments and assays for improved measurement of EVs