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Extracellular vesicles (EVs) play key roles in diverse biological processes, transport biomolecules between cells and have been engineered for therapeutic applications. A useful EV bioengineering strategy is to express engineered proteins on the EV surface to confer targeting, bioactivity and other properties. Measuring how incorporation varies across a population of EVs is important for characterising such materials and understanding their function, yet it remains challenging to quantitatively characterise the absolute number of engineered proteins incorporated at single-EV resolution. To address these needs, we developed a HaloTag-based characterisation platform in which dyes or other synthetic species can be covalently and stoichiometrically attached to engineered proteins on the EV surface. To evaluate this system, we employed several orthogonal quantification methods, including flow cytometry and fluorescence microscopy, and found that HaloTag-mediated quantification is generally robust across EV analysis methods. We compared HaloTag-labelling to antibody-labelling of EVs using single vesicle flow cytometry, enabling us to measure the substantial degree to which antibody labelling can underestimate proteins present on an EV. Finally, we demonstrate the use of HaloTag to compare between protein designs for EV bioengineering. Overall, the HaloTag system is a useful EV characterisation tool which complements and expands existing methods.

In this research-based experiment, students are introduced to the interdisciplinary field of archaeological chemistry by extracting and analyzing lipid residues absorbed in pottery. Reproduction archaeological pottery sherds are prepared by soaking ceramic fragments in individual or combinations of vegetable oils. Students crush and extract the absorbed residues, transesterify the recovered lipids to fatty acid methyl esters, and analyze the product by GC or GC/MS. Recovered residues are characterized by analysis of the major fatty acid peaks, as identified using reference standards or mass spectral databases. An archaeological context that links the sherds to the Minoan civilization is provided to students and must be considered to correctly identify the absorbed residue(s). The laboratory has been used in a topical archaeological instrumentation course that has attracted second to fourth year students majoring in chemistry, biochemistry, and anthropology, and museum studies minors. Pedagogically, the laboratory introduces students to techniques currently used in the field of archaeological chemistry while reinforcing fundamental concepts in sample isolation and preparation, derivatization, gas chromatography, mass spectrometry, and multicomponent sample analysis.

ObjectiveTo assess lunchtime provision of food and drink in English primary schools and to assess both choices and consumption of food and drink by pupils having school lunches. These findings were compared with similar data collected in 2005.DesignCross-sectional data collected between February and April 2009. In each school, food and drink provision, including portion weights and number of portions of each item served at lunchtime, were recorded over five consecutive days. Caterers provided school lunchtime menus and recipes.SettingEngland.SubjectsA random selection of 6696 pupils having school lunches in a nationally representative sample of 136 primary schools in England.ResultsCompared with 2005, schools in 2009 provided significantly more fruit, fruit-based desserts, vegetables and salad, water and fruit juice, and less ketchup, sauces and gravy, starchy foods cooked in fat, snacks and confectionery (P < 0·01). Pupils were also making healthier choices, choosing an average of 2·2 portions of fruit and vegetables from their ‘five a day’, but about one-third to two-fifths of these were wasted.ConclusionsLunchtime food provision and consumption in primary schools have improved substantially since 2005, following the introduction of new standards for school food in 2008. However, improvements still need to be made to increase the Fe and Zn content and to decrease the Na content of recipes, and in encouraging pupils to eat more of the fruits and vegetables taken at lunchtime. [ABSTRACT FROM AUTHOR]

Cells form multiple, molecularly distinct membrane contact sites (MCSs) between organelles. Despite knowing the molecular identity of several of these complexes, little is known about how MCSs are coordinately regulated in space and time to promote organelle function. Here, we examined two well-characterized mitochondria-endoplasmic reticulum (ER) MCSs - the ER-mitochondria encounter structure (ERMES) and the mitochondria-ER-cortex anchor (MECA) in Saccharomyces cerevisiae. We report that loss of MECA results in a substantial reduction in the number of ERMES contacts. Rather than reducing ERMES protein levels, loss of MECA results in an increase in the size of ERMES contacts. Using live-cell microscopy, we demonstrate that ERMES contacts display several dynamic behaviors, such as de novo formation, fusion and fission, that are altered in the absence of MECA or by changes in growth conditions. Unexpectedly, we find that the mitochondria-plasma membrane (PM) tethering, and not the mitochondria-ER tethering, function of MECA regulates ERMES contacts. Remarkably, synthetic tethering of mitochondria to the PM in the absence of MECA is sufficient to rescue the distribution of ERMES foci. Overall, our work reveals how one MCS can influence the regulation and function of another., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2025. Published by The Company of Biologists.)

Extracellular vesicles (EVs) play key roles in diverse biological processes, transport biomolecules between cells, and have been engineered for therapeutic applications. A useful EV bioengineering strategy is to express engineered proteins on the EV surface to confer targeting, bioactivity, and other properties. Measuring how incorporation varies across a population of EVs is important for characterizing such materials and understanding their function, yet it remains challenging to quantitatively characterize the absolute number of engineered proteins incorporated at single-EV resolution. To address these needs, we developed a HaloTag-based characterization platform in which dyes or other synthetic species can be covalently and stoichiometrically attached to engineered proteins on the EV surface. To evaluate this system, we employed several orthogonal quantification methods, including flow cytometry and fluorescence microscopy, and found that HaloTag-mediated quantification is generally robust across EV analysis methods. We compared HaloTag-labeling to antibody-labeling of EVs using single vesicle flow cytometry, enabling us to measure the substantial degree to which antibody labeling can underestimate proteins present on an EV. Finally, we demonstrate the use of HaloTag to compare between protein designs for EV bioengineering. Overall, the HaloTag system is a useful EV characterization tool which complements and expands existing methods.