A Reproducible Approach to the Assembly of Microcapillaries for Double Emulsion Production

Double emulsions attract considerable interest for their potential utility in applications as diverse as drug delivery, contrast agents, and compartmentalizing analytes for fluorescence-activated cell sorting (FACS). Microfluidic platforms provide a particularly elegant approach to generating these structures, but the construction of devices to provide reproducible and stable production of double emulsions remains challenging. PDMS-based systems require specialized surface treatments that are difficult to implement and lack long-term stability, and current glass microcapillary systems, while offering some advantages, lack flexible and reproducible methods for capillary alignment. This article describes a microcapillary-based approach that addresses these key challenges. Our approach utilizes translational stage elements and alignment end caps that are fixed in place once configured, rather than tightly fitting capillaries. This new approach enables alignment to within ± 10 µm and allows greater flexibility in choosing the dimensions of the capillary, which contributes to the size and stability of formation of the double emulsion. Importantly, it also allows the user to compensate for the deviations from ideal shape that occur in pulled glass capillaries, which has been a source of failure with previous methods. A detailed description of the critical design and operational parameters that affect double emulsion generation in these capillary microfluidic devices is provided

3D Visualisation of Additive Occlusion and Tunable Full-Spectrum Fluorescence in Calcite

From biomineralization to synthesis, organic additives provide an effective means of controlling crystallisation processes. There is growing evidence that these additives are often occluded within the crystal lattice, where this promises an elegant means of creating nanocomposites and tuning physical properties. Here, we use the incorporation of sulfonated fluorescent dyes to gain new understanding of additive occlusion in calcite (CaCO3), and to link morphological changes to occlusion mechanisms. We demonstrate that these additives are incorporated within specific zones, as defined by the growth conditions, and show how occlusion can govern changes in crystal shape. Fluorescence spectroscopy and lifetime imaging microscopy also show that the dyes experience unique local environments within different zones. Our strategy was then extended to simultaneously incorporate mixtures of dyes, whose fluorescence cascade creates calcite nanoparticles that fluoresce white. This offers a simple strategy for generating biocompatible and stable fluorescent nanoparticles whose output can be tuned as required

Evaluation of microflow configurations for scale inhibition and serial X-ray diffraction analysis of crystallization processes

The clean and reproducible conditions provided by microfluidic devices are ideal sample environments for in situ analyses of chemical and biochemical reactions and assembly processes. However, the small size of microchannels makes investigating the crystallization of poorly soluble materials on-chip challenging due to crystal nucleation and growth that result in channel fouling and blockage. Here, we demonstrate a reusable insert-based microfluidic platform for serial X-ray diffraction analysis and examine scale formation in response to continuous and segmented flow configurations across a range of temperatures. Under continuous flow, scale formation on the reactor walls begins almost immediately on mixing of the crystallizing species, which over time results in occlusion of the channel. Depletion of ions at the start of the channel results in reduced crystallization towards the end of the channel. Conversely, segmented flow can control crystallization, so it occurs entirely within the droplet. Consequently, the spatial location within the channel represents a temporal point in the crystallization process. Whilst each method can provide useful crystallographic information, time-resolved information is lost when reactor fouling occurs and changes the solution conditions with time. The flow within a single device can be manipulated to give a broad range of information addressing surface interaction or solution crystallization

Rapid Mapping of Protein Interactions Using Tag‐Transfer Photocrosslinkers

Analysing protein complexes by chemical crosslinking‐mass spectrometry (XL‐MS) is limited by the side‐chain reactivities and sizes of available crosslinkers, their slow reaction rates, and difficulties in crosslink enrichment, especially for rare, transient or dynamic complexes. Here we describe two new XL reagents that incorporate a methanethiosulfonate (MTS) group to label a reactive cysteine introduced into the bait protein, and a residue‐unbiased diazirine‐based photoactivatable XL group to trap its interacting partner(s). Reductive removal of the bait transfers a thiol‐containing fragment of the crosslinking reagent onto the target that can be alkylated and located by MS sequencing and exploited for enrichment, enabling the detection of low abundance crosslinks. Using these reagents and a bespoke UV LED irradiation platform, we show that maximum crosslinking yield is achieved within 10 seconds. The utility of this “tag and transfer” approach is demonstrated using a well‐defined peptide/protein regulatory interaction (BID80‐102/MCL‐1), and the dynamic interaction interface of a chaperone/substrate complex (Skp/OmpA)

Dynamic Crystallization Pathways of Polymorphic Pharmaceuticals Revealed in Segmented Flow with Inline Powder X-ray Diffraction

Understanding the transitions between polymorphs is essential in the development of strategies for manufacturing and maximizing the efficiency of pharmaceuticals. However, this can be extremely challenging: crystallization can be influenced by subtle changes in environment, such as temperature and mixing intensity or even imperfections in the crystallizer walls. Here, we highlight the importance of in situ measurements in understanding crystallization mechanisms, where a segmented flow crystallizer was used to study the crystallization of the pharmaceuticals urea: barbituric acid (UBA) and carbamazepine (CBZ). The reactor provides highly reproducible reaction conditions, while in situ synchrotron powder X-ray diffraction (PXRD) enables us to monitor the evolution of this system. UBA has two polymorphs of almost equivalent free-energy and so is typically obtained as a polymorphic mixture. In situ PXRD analysis uncovered a progression of polymorphs from UBA III to the thermodynamic polymorph UBA I, where different positions along the length of the tubular flow crystallizer correspond to different reaction times. Addition of UBA I seed crystals modified this pathway such that only UBA I was observed throughout, while transformation from UBA III into UBA I still occurred in the presence of UBA III seeds. Information regarding the mixing-dependent kinetics of the CBZ form II to III transformation was also uncovered in a series of seeded and unseeded flow crystallization runs, despite atypical habit expression. These results illustrate the importance of coupling controlled reaction environments with in situ XRD to study the phase relationships in polymorphic materials

Wake up, wake up! It's me! It's my life! patient narratives on person-centeredness in the integrated care context: a qualitative study

Person-centered care emphasizes a holistic, humanistic approach that puts patients first, at the center of medical care. Person-centeredness is also considered a core element of integrated care. Yet typologies of integrated care mainly describe how patients fit within integrated services, rather than how services fit into the patient's world. Patient-centeredness has been commonly defined through physician's behaviors aimed at delivering patient-centered care. Yet, it is unclear how 'person-centeredness' is realized in integrated care through the patient voice. We aimed to explore patient narratives of person-centeredness in the integrated care context

Interactions between genes involved in the antioxidant defence system and breast cancer risk

The aim of the study is to examine the association between multilocus genotypes across 10 genes encoding proteins in the antioxidant defence system and breast cancer. The 10 genes are SOD1, SOD2, GPX1, GPX4, GSR, CAT, TXN, TXN2, TXNRD1 and TXNRD2. In all, 2271 cases and 2280 controls were used to examine gene–gene interactions between 52 single nucleotide polymorphisms (SNPs) that are hypothesised to tag all common variants in the 10 genes. The statistical analysis is based on three methods: unconditional logistic regression, multifactor dimensionality reduction and hierarchical cluster analysis. We examined all two- and three-way combinations with unconditional logistic regression and multifactor dimensionality reduction, and used a global approach with all SNPs in the hierarchical cluster analysis. Single-locus studies of an association of genetic variants in the antioxidant defence genes and breast cancer have been contradictory and inconclusive. It is the first time, to our knowledge, the association between multilocus genotypes across genes coding for antioxidant defence enzymes and breast cancer is investigated. We found no evidence of an association with breast cancer with our multilocus approach. The search for two-way interactions gave experiment-wise significance levels of P=0.24 (TXN [t2715c] and TXNRD2 [g23524a]) and P=0.58 (GSR [c39396t] and TXNRD2 [a442g]), for the unconditional logistic regression and multifactor dimensionality reduction, respectively. The experiment-wise significance levels for the three-way interactions were P=0.94 (GPX4 [t2572c], TXN [t2715c] and TXNRD2 [g23524a]) and P=0.29 (GSR [c39396t], TXN [t2715c] and TXNRD2 [a442g]) for the unconditional logistic regression and multifactor dimensionality reduction, respectively. In the hierarchical cluster analysis neither the average across four rounds with replacement of missing values at random (P=0.12) nor a fifth round with more balanced proportion of missing values between cases and controls (P=0.17) was significant

Explanation and relations. How do general practitioners deal with patients with persistent medically unexplained symptoms: a focus group study

Contains fulltext : 80758.pdf (publisher's version ) (Open Access)BACKGROUND: Persistent presentation of medically unexplained symptoms (MUS) is troublesome for general practitioners (GPs) and causes pressure on the doctor-patient relationship. As a consequence, GPs face the problem of establishing an ongoing, preferably effective relationship with these patients. This study aims at exploring GPs' perceptions about explaining MUS to patients and about how relationships with these patients evolve over time in daily practice. METHODS: A qualitative approach, interviewing a purposive sample of twenty-two Dutch GPs within five focus groups. Data were analyzed according to the principles of constant comparative analysis. RESULTS: GPs recognise the importance of an adequate explanation of the diagnosis of MUS but often feel incapable of being able to explain it clearly to their patients. GPs therefore indicate that they try to reassure patients in non-specific ways, for example by telling patients that there is no disease, by using metaphors and by normalizing the symptoms. When patients keep returning with MUS, GPs report the importance of maintaining the doctor-patient relationship. GPs describe three different models to do this; mutual alliance characterized by ritual care (e.g. regular physical examination, regular doctor visits) with approval of the patient and the doctor, ambivalent alliance characterized by ritual care without approval of the doctor and non-alliance characterized by cutting off all reasons for encounter in which symptoms are not of somatic origin. CONCLUSION: GPs feel difficulties in explaining the symptoms. GPs report that, when patients keep presenting with MUS, they focus on maintaining the doctor-patient relationship by using ritual care. In this care they meticulously balance between maintaining a good doctor-patient relationship and the prevention of unintended consequences of unnecessary interventions

Do physician outcome judgments and judgment biases contribute to inappropriate use of treatments? Study protocol

<p>Abstract</p> <p>Background</p> <p>There are many examples of physicians using treatments inappropriately, despite clear evidence about the circumstances under which the benefits of such treatments outweigh their harms. When such over- or under- use of treatments occurs for common diseases, the burden to the healthcare system and risks to patients can be substantial. We propose that a major contributor to inappropriate treatment may be how clinicians judge the likelihood of important treatment outcomes, and how these judgments influence their treatment decisions. The current study will examine the role of judged outcome probabilities and other cognitive factors in the context of two clinical treatment decisions: 1) prescription of antibiotics for sore throat, where we hypothesize overestimation of benefit and underestimation of harm leads to over-prescription of antibiotics; and 2) initiation of anticoagulation for patients with atrial fibrillation (AF), where we hypothesize that underestimation of benefit and overestimation of harm leads to under-prescription of warfarin.</p> <p>Methods</p> <p>For each of the two conditions, we will administer surveys of two types (Type 1 and Type 2) to different samples of Canadian physicians. The primary goal of the Type 1 survey is to assess physicians' perceived outcome probabilities (both good and bad outcomes) for the target treatment. Type 1 surveys will assess judged outcome probabilities in the context of a representative patient, and include questions about how physicians currently treat such cases, the recollection of rare or vivid outcomes, as well as practice and demographic details. The primary goal of the Type 2 surveys is to measure the specific factors that drive individual clinical judgments and treatment decisions, using a 'clinical judgment analysis' or 'lens modeling' approach. This survey will manipulate eight clinical variables across a series of sixteen realistic case vignettes. Based on the survey responses, we will be able to identify which variables have the greatest effect on physician judgments, and whether judgments are affected by inappropriate cues or incorrect weighting of appropriate cues. We will send antibiotics surveys to family physicians (300 per survey), and warfarin surveys to both family physicians and internal medicine specialists (300 per group per survey), for a total of 1,800 physicians. Each Type 1 survey will be two to four pages in length and take about fifteen minutes to complete, while each Type 2 survey will be eight to ten pages in length and take about thirty minutes to complete.</p> <p>Discussion</p> <p>This work will provide insight into the extent to which clinicians' judgments about the likelihood of important treatment outcomes explain inappropriate treatment decisions. This work will also provide information necessary for the development of an individualized feedback tool designed to improve treatment decisions. The techniques developed here have the potential to be applicable to a wide range of clinical areas where inappropriate utilization stems from biased judgments.</p