'Why didn't they see my scars?' Critical thematic analysis of simulated participants' perceived tensions surrounding objective structured clinical examinations.

BACKGROUND: Simulated participants (SPs) play an important role in simulated assessments of clinical encounters between medical students and patients, most notably in objective structured clinical examinations (OSCEs). SP contributions to OSCEs are invaluable, taking the role of a patient or carer. While SPs in some settings/contexts may rate students, their role has been problematized in the literature for their lack of agency within a standardised format of OSCEs that promotes reliability, objectivity and accountability. In this study, we explored SP experiences for tensions that result from simulated assessments and their potential implications for education. METHODS: Semi-structured interviews were conducted with seven SPs who were also tasked with providing a global mark for students. They were purposively selected to include women and men of different ages, occupation, education and experience as an SP. The interviews were analysed using a critical thematic analysis using a phenomenological approach. RESULTS: SP experiences directly addressed tensions and contradictions around OSCEs. SP participants described their experiences under four themes: industrialising, reducing, performativity and patient safety. OSCEs were compared to an industrial process that promoted efficiency but which bore no resemblance to real-life doctor-patient encounters. They were perceived to have a power and agency that reduced SPs to verbalising scripts to ensure that students were exposed to a standardised simulated experience that also underlined the performative role of SPs as props. These performative and reductionist experiences extended to students, for whom the mark sheet acted as a checklist, promoting standardised responses that lacked genuineness. All of this created a tension for SPs in promoting patient safety by ensuring that those medical students who passed were clinically competent. CONCLUSIONS: OSCEs often form part of high-stakes exams. As such, they are governed by processes of industrialisation, accountability and standardisation. OSCEs possess a power and agency that can have unintended negative consequences. These include 'conditioning' students to adopt behaviours that are not suited to real-life clinical encounters and are not person-centred.

Determination of Zn, Cu and Fe in human patients' serum using micro-sampling ICP-MS and sample dilution.

A high-throughput, sensitive and rapid method was developed for the determination of Zn, Cu and Fe in small volumes (30 µL) of human serum using inductively coupled plasma mass spectrometry (ICP-MS). The sample preparation procedure employed simple 100-fold dilution of the serum samples with 1.0% butanol, 0.5% v/v ammonia, 0.02% v/v Triton X-100 and 0.01% v/v HNO3. The reliability of the method was evaluated using serum UTAK certified reference material, and the results matched well with the certified values. The method was applied to determine Zn, Cu and Fe in 81 human serum samples from participants in Alzheimer disease (AD) and age-related macular degeneration (AMD) studies. No significant differences were found in Zn and Cu levels between age matched controls, AD and AMD patients. Whilst iron levels appeared marginally higher in the AMD group, compared with the AD group, iron showed larger overall variability than the other two elements.

Tricine-SDS-PAGE.

Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (tricine-SDS-PAGE) is an efficient way of separating low molecular mass proteins. However, the standard system is quite complicated and specifically may not be useful when the separated proteins are to be recovered from the gel for quantitative analysis. Here, we describe a simplified system whereby these smaller proteins can be resolved in comparatively low percentage gels which have high compatibility with modern detectors such as UV and inductively coupled plasma-mass spectrometry (ICP-MS).

Content and discontent: a qualitative exploration of obstacles to elearning engagement in medical students.

BACKGROUND: Elearning is ubiquitous in healthcare professions education. Its equivalence to 'traditional' educational delivery methods is well established. There is a research imperative to clarify when and how to use elearning most effectively to mitigate the potential of it becoming merely a 'disruptive technology.' Research has begun to broadly identify challenges encountered by elearning users. In this study, we explore in depth the perceived obstacles to elearning engagement amongst medical students. Sensitising concepts of achievement emotions and the cognitive demands of multi-tasking highlight why students' deeply emotional responses to elearning may be so important in their learning. METHODS: This study used focus groups as a data collection tool. A purposeful sample of 31 participated. Iterative data gathering and analysis phases employed a constant comparative approach to generate themes firmly grounded in participant experience. RESULTS: Key themes that emerged from the data included a sense of injustice, passivity and a feeling of being 'lost at sea'. The actual content of the elearning resource provided important context. CONCLUSIONS: The identified themes have strong emotional foundations. These responses, interpreted through the lens of achievement emotions, have not previously been described. Appreciation of their importance is of benefit to educators involved in curriculum development or delivery.

High-Speed, Integrated Ablation Cell and Dual Concentric Injector Plasma Torch for Laser Ablation-Inductively Coupled Plasma Mass Spectrometry.

In recent years, laser ablation-inductively coupled plasma mass spectrometry (LA-ICPMS) has gained increasing importance for biological analysis, where ultratrace imaging at micrometer resolution is required. However, while undoubtedly a valuable research tool, the washout times and sensitivity of current technology have restricted its routine and clinical application. Long periods between sampling points are required to maintain adequate spatial resolution. Additionally, temporal signal dispersion reduces the signal-to-noise ratio, which is a particular concern when analyzing discrete samples, such as individual particles or cells. This paper describes a novel, two-volume laser ablation cell and integrated ICP torch designed to minimize aerosol dispersion for fast, efficient sample transport. The holistic design utilizes a short, continuous diameter fused silica conduit, which extends from the point of ablation, through the ICP torch, and into the base of the plasma. This arrangement removes the requirement for a dispersive component for argon addition, and helps to keep the sample on axis with the ICP cone orifice. Hence, deposition of sample on the cones is theoretically reduced with a resulting improvement in the absolute sensitivity (counts per unit mole). The system described here achieved washouts of 1.5, 3.2, and 4.9 ms for NIST 612 glass, at full width half, 10%, and 1% maximum, respectively, with an 8-14-fold improvement in absolute sensitivity, compared to a single volume ablation cell. To illustrate the benefits of this performance, the system was applied to a contemporary bioanalytical challenge, specifically the analysis of individual biological cells, demonstrating similar improvements in performance.

A feasibility study of the use of saliva as an alternative to leukocytes as a source of DNA for the study of Pt-DNA adducts in cancer patients receiving platinum-based chemotherapy.

This note presents a comparison of the use of saliva versus leukocytes for the determination of Pt-DNA adducts obtained from patients undergoing platinum-based chemotherapy. Samples of both blood and saliva were taken pre- and post-treatment and were analysed via sector-field inductively coupled plasma mass spectrometry (SF-ICP-MS) to determine the level of Pt-DNA adducts formed. As expected, significant inter-patient variability was seen; however, a lack of correlation between the levels of adducts observed in saliva and blood samples was also observed (Pearson correlation coefficient r = -0.2598). A high yield of DNA was obtained from saliva samples, but significant difficulties were experienced in obtaining patient adherence to the saliva sampling procedure. In both leukocyte and saliva samples, not only was Pt from previous chemotherapy cycles detected, but the rapid appearance of Pt in the DNA was noted in both sample types 1 h after treatment.

Quantitative analysis of gold nanoparticles in single cells by laser ablation inductively coupled plasma-mass spectrometry.

Single cell analysis has become an important field of research in recent years reflecting the heterogeneity of cellular responses in biological systems. Here, we demonstrate a new method, based on laser ablation inductively coupled plasma mass spectrometry (LA-ICPMS), which can quantify in situ gold nanoparticles (Au NPs) in single cells. Dried residues of picoliter droplets ejected by a commercial inkjet printer were used to simulate matrix-matched calibration standards. The gold mass in single cells exposed to 100 nM NIST Au NPs (Reference material 8012, 30 nm) for 4 h showed a log-normal distribution, ranging from 1.7 to 72 fg Au per cell, which approximately corresponds to 9 to 370 Au NPs per cell. The average result from 70 single cells (15 ± 13 fg Au per cell) was in good agreement with the result from an aqua regia digest solution of 1.2 × 10(6) cells (18 ± 1 fg Au per cell). The limit of quantification was 1.7 fg Au. This paper demonstrates the great potential of LA-ICPMS for single cell analysis and the beneficial study of biological responses to metal drugs or NPs at the single cell level.

Laser ablation-inductively coupled plasma mass spectrometry: an emerging technology for detecting rare cells in tissue sections.

Administering immunoregulatory cells to patients as medicinal agents is a potentially revolutionary approach to the treatment of immunologically mediated diseases. Presently, there are no satisfactory, clinically applicable methods of tracking human cells in patients with adequate spatial resolution and target cell specificity over a sufficient period of time. Laser ablation-inductively coupled plasma mass spectrometry (LA-ICP-MS) represents a potential solution to the problem of detecting very rare cells in tissues. In this article, this exquisitely sensitive technique is applied to the tracking of gold-labeled human regulatory macrophages (Mregs) in immunodeficient mice. Optimal conditions for labeling Mregs with 50-nm gold particles were investigated by exposing Mregs in culture to variable concentrations of label: Mregs incubated with 3.5 × 10(9) particles/ml for 1 h incorporated an average of 3.39 × 10(8) Au atoms/cell without loss of cell viability. Analysis of single, gold-labeled Mregs by LA-ICP-MS registered an average of 1.9 × 10(5) counts/cell. Under these conditions, 100% labeling efficiency was achieved, and label was retained by Mregs for ≥36 h. Gold-labeled Mregs adhered to glass surfaces; after 24 h of culture, it was possible to colabel these cells with human-specific (154)Sm-tagged anti-HLA-DR or (174)Yb-tagged anti-CD45 mAbs. Following injection into immunodeficient mice, signals from gold-labeled human Mregs could be detected in mouse lung, liver, and spleen for at least 7 d by solution-based inductively coupled plasma mass spectrometry and LA-ICP-MS. These promising results indicate that LA-ICP-MS tissue imaging has great potential as an analytical technique in immunology.

Safe moving and handling of patients: an interprofessional approach.

A gap in the medical undergraduate curriculum on safe moving and handling of patients was identified, and a project to enhance moving and handling education for undergraduates in various healthcare disciplines was undertaken. A team of nurses, doctors, physiotherapists and e-learning professionals developed a cross-discipline e-learning resource, piloted with medical and nursing students at Queen's University Belfast. One outcome of the project was the development of a deeper recognition of the common curriculum across healthcare disciplines.

Single cell tracking of gadolinium labeled CD4+ T cells by laser ablation inductively coupled plasma mass spectrometry.

Cellular therapy is emerging as a promising alternative to conventional immunosuppression in the fields of hematopoietic stem cell (HSC) transplantation, autoimmune disease, and solid organ transplantation. Determining the persistence of cell-based therapies in vivo is crucial to understanding their regulatory function and requires the combination of an extremely sensitive detection technique and a stable, long-lifetime cell labeling agent. This paper reports the first application of laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) to perform single cell detection of T cell populations relevant to cellular immunotherapy. Purified human CD4(+) T cells were labeled with commercially available Gd-based magnetic resonance imaging (MRI) contrast agents, Omniscan and Dotarem, which enabled passive loading of up to 10(8) Gd atoms per cell. In mixed preparations of labeled and unlabeled cells, LA-ICP-MS was capable of enumerating labeled cells at close to the predicted ratio. More importantly, LA-ICP-MS single cell analysis demonstrated that the cells retained a sufficient label to remain detectable for up to 10 days post-labeling both in vitro and in vivo in an immunodeficient mouse model.

Oxaliplatin complexes with carnosine and its derivatives: in vitro cytotoxicity, mass spectrometric and computational studies with a focus on complex fragmentation.

The complexation of the Pt-based anti-cancer drug oxaliplatin (OxPt) with biological ligands other than DNA is believed to be a major cellular sink for the drug reducing its therapeutic potential and acting as a potential cause of toxicity. In this paper, the very first hypothesis driven investigation of the role of the naturally abundant cytoplasmic dipeptide ligand ß-alanyl-l-histidine dipeptide (carnosine) in OxPt detoxification is presented. In vitro studies on hepatocellular carcinoma HepG2 cells suggest that carnosine may inhibit the cytotoxic action of OxPt most likely through the formation of complexes that are less cytotoxic than OxPt alone. Evidence is provided to suggest that pre-exposure of HepG2 cells to elevated levels of carnosine appears to have a lasting effect on reducing the cytotoxicity of OxPt even after the removal of the externally added carnosine. This effect, however, is likely under kinetic control as its magnitude was shown not to vary significantly with the level of carnosine exposure within the concentration range used in this study. Various mass spectrometry techniques employing electrospray ionization and chip nanospray were employed to study the interaction of oxaliplatin with carnosine as well as two of its derivatives ß-alanyl-N-methylhistidine (anserine) and N-acetylcarnosine (NAC). Evidence of complexation between OxPt and each of the three ligands examined is presented. Most species observed were unambiguously assigned and compared to their theoretical isotopic patterns. Common fragmentation products due to the collisionally-activated protonated complexes of each of the ligands examined with OxPt, [M + OxPt + H](+), where M = carnosine, anserine or NAC, were reported. Density functional calculations at the B3LYP/LANL2DZ level were used to obtain structural information and relative free energies of different isomers of the observed precursor [Carnosine + OxPt + H](+) both in the gas phase and in solution as well as to probe its fragmentation, highlighting plausible fragmentation mechanisms that account for all the experimental results. Data are presented to show several binding modes between electron rich sites such as N and O centers of carnosine and the Pt metal of OxPt. Calculations were also employed to obtain proton affinities and free energies of key reactions. The proton affinities of carnosine, anserine and NAC at 298 K were calculated to be 254.4, 255.9 and 250.2 kcal mol(-1) respectively. To the best of our knowledge the proton affinities of anserine and N-acetyl-carnosine are the first reported values in the literature.

Tricine-SDS-PAGE.

Tricine-sodium dodecyl sulphate-polyacrylamide gel electrophoresis (tricine-SDS-PAGE) is an efficient way of separating low-molecular-mass proteins. However, the standard system is quite complicated and specifically may not be useful when the separated proteins require to be recovered from the gel for quantitative analysis. Here, we describe a simplified system whereby these smaller proteins can be resolved in comparatively low-percentage gels which have high compatibility with modern detectors such as UV and inductively coupled plasma mass spectrometry (ICP-MS).

Analysis of mono-phosphate nucleotides as a potential method for quantification of DNA using high performance liquid chromatography-inductively coupled plasma-mass spectrometry.

The determination of total deoxyribonucleic acid (DNA) concentration is of great importance in many biological and bio-medical analyses. The quantification of DNA is traditionally performed by UV spectroscopy; however the results can be affected greatly by the sample matrix. The proposed method quantifies phosphorus in digested calf thymus DNA and human DNA by high performance liquid chromatography (HPLC) combined with inductively coupled plasma mass spectrometry (ICP-MS). The method presented showed excellent baseline separation between all four DNA mono-nucleotides and 5'UMP. The ability of LC-ICP-MS to provide an internal check that only DNA derived phosphorus was counted in the assay was demonstrated by establishing a mass balance between the total phosphorous signal from undigested DNA and that from the speciated DNA. Column recoveries ranging from 95% to 99% for phosphorus resulted in a mass balance of 95% ± 0.5% for standard nucleotides, determined by LC-ICP-MS, compared to total DNA determined by flow injection coupled to ICP-MS (FI-ICP-MS). The method for quantification was validated by analysis of NIST SRM 2,372; a total speciated DNA recovery of 52.1 ng/µL, compared with an expected value of 53.6 ng/µL, was determined by external calibration. From repeat measurements, a mass balance of 97% ± 0.5% for NIST DNA was achieved. The method limits of detection for individual nucleotides were determined between 0.8 and 1.7 µg L(-1) ((31)P) for individual nucleotides by LC-ICP-MS, and 360 ng L(-1) for 5'AMP by direct nebulisation.

A comparison of Tris-glycine and Tris-tricine buffers for the electrophoretic separation of major serum proteins.

This paper compares different buffer systems for the electrophoretic separation of the five most abundant serum proteins on native-PAGE gel and cellulose membranes. A modified Tris-tricine system was shown to be superior for the separation of these serum proteins in a 7% m/v native-PAGE gel as compared with the traditionally used Tris-glycine and Tris-tricine methods. This modified Tris-tricine buffer system was also employed for the separation of serum proteins using a cellulose acetate membrane and very effective separation was observed as compared with the traditionally used Tris-barbital and Tris-glycine buffer systems.

Speciation of oxaliplatin adducts with DNA nucleotides.

This paper describes a set of fast and selective high performance liquid chromatography (HPLC) methods coupled to electro-spray ionisation linear ion trap mass spectrometry (ESI-MS), sector-field inductively coupled plasma mass spectrometry (SF-ICP-MS) and UV detection for in vitro studies of the bifunctional adducts of oxaliplatin with mono-nucleotides, di-nucleotides and cellular DNA. The stationary phases and the optimised conditions used for each separation are discussed. Interaction of oxaliplatin with A and G mono-nucleotides resulted in the formation of five bifunctional platinum diaminocyclohexane (DACHPt) adducts. These were two isomers of the A-DACHPt-A and A-DACHPt-G adducts, and one G-DACHPt-G adduct, as confirmed by MS/MS spectra obtained by collision induced dissociation. These adducts were also characterised by UV absorption data and SF-ICP-MS elemental (195)Pt and (31)P signals. Further, interaction of oxaliplatin with AG and GG di-nucleotides resulted in the formation of three adducts: DACHPt-GG and two isomers of the DACHPt-AG adduct, as confirmed by ESI-MS and the complementary data obtained by UV and SF-ICP-MS. Finally, a very sensitive LC-ICP-MS method for the quantification of oxaliplatin GG intra-strand adducts (DACHPt-GG) was developed and used for monitoring the in vitro formation and repair of these adducts in human colorectal cancer cells. The method detection limit was 0.14 ppb Pt which was equivalent to 0.22 Pt adduct per 10(6) nucleotides based on a 10 µg DNA sample. This detection limit makes this method suitable for in vivo assessment of DACHPt-GG adducts in patients undergoing oxaliplatin chemotherapy.

Modification of tricine-SDS-PAGE for online and offline analysis of phosphoproteins by ICP-MS.

This study describes a modification to tricine sodium dodecyl sulphate polyacrylamide gel electrophoresis to make it more effective for the separation of low molecular mass proteins and for coupling with inductively coupled plasma mass spectrometry (ICP-MS). The modified method employs low-percentage polyacrylamide gels (7-10%) (w/v) and low reagent concentrations that provide efficient separations, good quantitation and low matrix levels that are compatible with ICP-MS. Using phosphopeptides as a model system, and offline analysis, we obtained recoveries of 73% (w/v) in a 9% gel compared with 55% in a conventional 16% gel. Online coupling was achieved by modification of a standard commercially available gel electroelution apparatus and casting of the gel into a 7.3-cm-long tube. Online separation of a digest of beta-casein was demonstrated with recovery of the mono- and tetraphosphopeptides, which were identified by comparison with peptide standards. A mass balance study with the standards yielded recoveries of 95% for tetraphosphopeptides and 48% for monophosphopeptides. The factors affecting the separations and recoveries are discussed in detail. The detection limits for 10-microL samples of the mono- and tetraphosphopeptides were 0.7 microM (7 pmol) and 0.2 microM (2 pmol) respectively.

Determination of iodine and molybdenum in milk by quadrupole ICP-MS.

A reliable method for the determination of iodine and molybdenum in milk samples, using alkaline digestion with tetramethylammonium hydroxide and hydrogen peroxide, followed by quadrupole ICP-MS analysis, has been developed and tested using certified reference materials. The use of He+O2 (1.0 ml min(-1) and 0.6 ml min(-1)) in the collision-reaction cell of the mass spectrometer to remove (129)Xe+-- initially to enable the determination of low levels of 129I--also resulted in the quantitative conversion of Mo(+) to MoO2+ which enabled the molybdenum in the milk to be determined at similar mass to the iodine with the use of Sb as a common internal standard. In order to separate and pre-concentrate iodine at sub microg l(-1) concentrations, a novel method was developed using a cation-exchange column loaded with Pd2+ and Ca2+ ions to selectively retain iodide followed by elution with a small volume of ammonium thiosulfate. This method showed excellent results for aqueous iodide solutions, although the complex milk digest matrix made the method unsuitable for such samples. An investigation of the iodine species formed during oxidation and extraction of milk sample digests was carried out with a view to controlling the iodine chemistry.

Silver nanoparticles and polymeric medical devices: a new approach to prevention of infection?

OBJECTIVES: Implantable devices are major risk factors for hospital-acquired infection. Biomaterials coated with silver oxide or silver alloy have all been used in attempts to reduce infection, in most cases with controversial or disappointing clinical results. We have developed a completely new approach using supercritical carbon dioxide to impregnate silicone with nanoparticulate silver metal. This study aimed to evaluate the impregnated polymer for antimicrobial activity. METHODS: After impregnation the nature of the impregnation was determined by transmission electron microscopy. Two series of polymer discs were then tested, one washed in deionized water and the other unwashed. In each series, half of the discs were coated with a plasma protein conditioning film. The serial plate transfer test was used as a screen for persisting activity. Bacterial adherence to the polymers and the rate of kill, and effect on planktonic bacteria were measured by chemiluminescence and viable counts. Release rates of silver ions from the polymers in the presence and absence of plasma was measured using inductively coupled plasma mass spectrometry (ICP-MS). RESULTS: Tests for antimicrobial activity under various conditions showed mixed results, explained by the modes and rates of release of silver ions. While washing removed much of the initial activity there was continued release of silver ions. Unexpectedly, this was not blocked by conditioning film. CONCLUSIONS: The methodology allows for the first time silver impregnation (as opposed to coating) of medical polymers and promises to lead to an antimicrobial biomaterial whose activity is not restricted by increasing antibiotic resistance.