Macrophages play a critical role in the development of the tumor microenvironment (TME). Recruited macrophages in the TME differentiate into various phenotypes, each with a distinct profile of secreted cytokines. To describe and understand this large heterogeneity, we developed nested nanowell arrays for the multiplexed analysis of secreted proteins at the single cell level. The array consists of more than 100,000 wells on a cyclic olefin copolymer (COC) substrate. Each well contains seven smaller indents for cocapturing functionalized beads and can be operated with standard laboratory equipment such as pipettes and microscopes. The barcoded beads capture cytokines of interest and allow their quantification via sandwich immunoassays. We developed an image analysis tool and quantified 10 proteins secreted from single macrophages and investigated the effects of stimulation and drug treatment. We found that interleukin (IL)-1β, IL-8, and macrophage inflammatory protein 1α (MIP-1α) were highly secreted by more than 43% of the macrophages, with an increase of MIP-1α secretion under treatment with the chemotherapeutic drugs paclitaxel or docetaxel. Pairwise protein analysis confirmed cosecretion of IL-1β and IL-6 in macrophages stimulated with IL-4/IL-13, which were identified as part of a critical pathway in multiple myeloma before. We also demonstrate further multiplexing with three and four cosecreted proteins, together with assessing the cell viability as an additional parameter important for drug testing. In summary, we have shown that our nested nanowell arrays are an easy-to-use analytical tool for basic research, and we believe that it can be employed for diagnostics and personalized medicine, e.g., for investigation of cancer and immune cells from a biopsy or circulating tumor cells obtained from a liquid biopsy.

Membrane transport is fundamental to biological cells and is a major hurdle in the rational design of pharmaceuticals. To measure membrane transport in vitro, most methods focus on simple diffusion of a single analyte across nonbiomimetic interfaces. Membrane engineering has facilitated novel strategies for the reconstitution, characterization, and application of biomimetic membranes. Herein, we define drug mixture analysis, an in vitro, label-free HPLC-MS, droplet interface bilayer (DIB) method, to assess membrane transport of drug mixtures and delineate simultaneous transport mechanisms. We use our method to classify the permeability of drugs from a structurally diverse FDA-approved library. This deep analysis uncovered correlation between determined permeability classifiers and drug properties such as the hydrophobic retention time, hydrogen bond donor count, lipophilicity, and predicted gut absorption. Across higher mimetic membranes, passive transport was quantified under physiologically relevant variables such as pH, temperature, lipid composition, and, in the presence of proteins, the coexistence of facilitated diffusion and simple diffusion. Our results show that DIBs are physiologically relevant interfaces for investigating membrane transport mechanisms relevant to artificial cell systems and drug screening.

Liquids are dense repositories of information, challenged only by how well their compositions are defined, preserved, accessed, or measured. The precise spatial patterning of solutes within a bulk liquid is challenging since diffusion disperses local concentrations and thereby attenuates functionality. Herein, a new concept is introduced for writing and preserving information in the liquid state through liquid-in-liquid microdroplet array printing. This technology produces fine resolution, 2D liquid structures, composite of indexed water-in-oil droplet pixels each with a precise composition, a high spatial resolution and a tight inter-pixel pitch. With extreme control over droplet composition and by applying standard and custom encoding schemes, various forms of information are written biochemically such as images, QR codes, text characters and words. As a composite material, reversible phase transitions between dissolved liquid and crystallized solid states control information encryption and decryption. Compared to current liquid printing and chemical encoding paradigms, ours introduces a fundamentally new precedent for deterministically programming information release, exchange or decay without stimuli or physical processing. Further computational principles such as error correction and information storage are demonstrated. These micro-liquid patterns are relevant to any application based on precise liquid handling such as information theory, materials design and biological assays.

Treating bacterial infections is dependent upon their site within a biological system, where the cumulative role of membrane transport is challenging to resolve. In this work, a cultivation method based on droplet interface bilayers (DIBs) is established. The architecture of infections in both cellular and tissue contexts is crafted where individual droplets serve as artificial cells infected by intracellular bacteria, or as interconnected units in a tissue-like structure. Through spatio-temporal control over droplets, addition, withdrawal, and sequential antibiotic gradients are tailored acting upon living bacteria. With droplet networks mimicking tissues, it is showed that the treatment response is dependent on the number of the cell-like barriers, corresponding to the number of membranes from an antibiotic source, here described as the membrane depth. Through mathematical modelling a correlation is revealed between the membrane depth of each bacterial population, the antibiotic distribution and thus the treatment efficacy. Ultimately, this approach holds promise as an in vitro bioassay for understanding the response of intracellular bacteria to antibiotics, developing new antibiotics, designing biologically inspired materials, and underpinning emerging bioprinting approaches.

Droplet microarrays underpin novel experimentation across the biological and chemical sciences. This perspective explores operations and analysis with droplet microarrays, placing focus on a comparison to traditional multiwell plates.

Simple diffusion of molecular entities through a phospholipid bilayer, is a phenomenon of great importance to the pharmaceutical and agricultural industries. Current model lipid systems to probe this typically only employ fluorescence as a readout, thus limiting the range of assessable chemical matter that can be studied. We report a new technology platform, the UV-DIB, which facilitates label free measurement of small molecule translocation rates. This is based upon the coupling of droplet interface bilayer technology with implemented fiber optics to facilitate analysis via ultraviolet spectroscopy, in custom designed PMMA wells. To improve on current DIB technology, the platform was designed to be reusable, with a high sampling rate and a limit of UV detection in the low μM regime. We demonstrate the use of our system to quantify passive diffusion in a reproducible and rapid manner where the system was validated by investigating multiple permeants of varying physicochemical properties across a range of lipid interfaces, each demonstrating differing kinetics. Our system permits the interrogation of structural dependence on the permeation rate of a given compound. We present this ability from two structural perspectives, that of the membrane, and the permeant. We observed a reduction in permeability between pure DOPC and DPhPC interfaces, concurring with literature and demonstrating our ability to study the effects of lipid composition on permeability. In relation to the effects of permeant structure, our device facilitated the rank ordering of various compounds from the xanthine class of compounds, where the structure of each permeant differed by a single group alteration. We found that DIBs were stable up to 5% DMSO, a molecule often used to aid solubilisation of pharmaceutical and agrochemical compounds. The ability of our device to rank-order compounds with such minor structural differences provides a level of precision that is rarely seen in current, industrially applied technologies.

Lipid membranes are vital in a wide range of biological and biotechnical systems; underpinning functions from modulation of protein activity to drug uptake and delivery. Rapid in situ determination of phase behaviour is a key experimental objective.

Droplet microcompartments linked by lipid bilayers show great promise in the construction of synthetic minimal tissues. Central to controlling the flow of information in these systems are membrane proteins, which can gate in response to specific stimuli in order to control the molecular flux between membrane separated compartments. This has been demonstrated with droplet interface bilayers (DIBs) using several different membrane proteins combined with electrical, mechanical, and/or chemical activators. Here we report the activation of the bacterial mechanosensitive channel of large conductance (MscL) in a dioleoylphosphatidylcholine:dioleoylphosphatidylglycerol DIB by controlling membrane asymmetry. We show using electrical measurements that the incorporation of lysophosphatidylcholine (LPC) into one of the bilayer leaflets triggers MscL gating in a concentration-dependent manner, with partial and full activation observed at 10 and 15 mol% LPC respectively. Our findings could inspire the design of new minimal tissues where flux pathways are dynamically defined by lipid composition.

With the ever increasing pace of change in the Chemical and Pharmaceutical Industry, universities need to be more aware of the skill set that employers are seeking in graduates. These skills often include risk taking, creative thinking and entrepreneurship; skills that are not naturally associated with chemistry-related curriculums. This research focusses on the authors’ experiences of applying for strategic grants which underpin the dissemination of such skills, and explores how these were used as the basis for creating specific learning content for final year students. Students’ experiences after completing assignments were gathered through an online survey and subsequent analysis identified areas for improvement in chemistry curriculums. Keywords: Entrepreneurship, Chemical & Pharmaceutical industry, innovation

In previous papers attention has chiefly been paid to the properties of active nitrogen when produced. The present one deals almost entirely with the circumstances of its production by the electric discharge. The jar discharge is much the most efficient, but does not lend itself easily to quantitative investigation. It is not easily maintained steady for any length of time, but there is a more fundamental difficulty than this, for measurements of current and potential with the jar discharge do not admit of any simple interpretation. Each discharge lasts for a time which is very short compared with the intervals between discharges, and, when it does occur, it is oscillatory. For these reasons attention has been given to the steady discharge obtained from a direct current dynamo machine. This yields much less active nitrogen than the jar discharge, but still enough to admit of satisfactory observations on many points.

It is known that vacuum tubes frequently show a luminosity of the contained gas after discharge is over. In a previous paper I was able to show that this effect, as it occurs in air, is of the nature of a phosphorescent combustion, and is due to the mutual reaction of nitric oxide and ozone, each formed in the discharge. In a second paper it was shown that other phosphorescent combustions can be observed in ozone, notably of sulphur, sulphuretted hydrogen, acetylene, and iodine. Some of these give continuous spectra, but the majority band spectra. In the first paper it was stated that pure nitrogen gives no afterglow whatever, and, with the simple induction coil discharge with which I was then working, this has been frequently verified since. Mr. Percival Lewis has however, described an afterglow obtained in nitrogen when a jar discharge with spark-gap is used. I had no difficulty in obtaining this glow as soon as the jar discharge was used, and have applied to its examination the method used in the former papers. This is due to Sir James Dewar, and consists in drawing a current of gas through the vacuum tube into an observing vessel, where the glow is developed, and thence into an air pump, which must be a mechanical one of good construction, driven by power. It is thus possible to examine the properties of the glowing nitrogen much more satisfactorily than can be done by intermittent examination after successive discharges.

Professor Rutherford has given a calculation which suggests that there may be enough radium in the earth to account for the temperature gradient observed near the surface. The question is of great interest from a cosmical point of view. For if we find that the earth’s internal heat is due to radio-activity, and if we assume, as has been usual, that this heat is due to some vestiges of the cause operative in the sun and stars, it would follow that these latter are heated by radio-active changes also.

In the current number of Wiedeman’s 'Annalen,’ an experiment is described by Giesel showing that the Becquerel rays are deflected in a magnetic field. This result is of great interest, on account of the light which it throws on the nature of the rays.

In recent experiments by myself and by others upon the density of hydrogen, the gas has always been dried by means of phosphoric anhydride; and a doubt may remain whether on the one hand the removal of aqueous vapour is sufficiently complete, and on the other whether some new impurity may not be introduced.