Towards voltammetric point of care detection of leucovorin.

Current healthcare trends have seen an increased emphasis on the move towards personalised precision medicine to tailor treatments to the individual and their response to diseases and disease therapies. This highlighting a transition from traditional "one size fits all" to a more nuanced approach. Despite advancements in fundamental knowledge to facilitate personalised prevision approaches, lack of resources to implement such plans remains one of the largest hurdles to overcome. Monitoring of drug therapies is one key aspect that could aid in the evolution of precision medicine alongside the development of drugs and targeted treatment systems. This contribution illustrates the potential of square wave voltammetry (SWV) as a proof-of-concept for monitoring of circulating blood concentrations of treatment therapies within artificial urine, using leucovorin calcium (LV) as a model cancer therapy drug. A low cost, easy-to-use and portable sensor has been developed and successfully employed for the detection of LV over the linear range 0.5-30 µM which represents the therapeutically relevant concentrations for LV within artificial urine without any prior sample preparation required with a limit of detection of 2.63 µM and initial investigations into saliva and serum as biological matrices. The developed sensor describe herein exhibits a proof-of-concept for the engagement of such electrochemical sensors as point-of-care devices, where the sensors ease of use and removal of time-consuming and complex sample preparation methods will ultimately increase its usability by physicians, widening the avenues where electrochemical sensors could be employed.

Evolution of nanomaterial Electrochemiluminescence luminophores towards biocompatible materials.

Electrochemiluminescence (ECL) is a powerful electrochemical technique for the detection and quantification of molecules, both synthetic and biological in origin. Traditional ECL luminophores are based on organic or organometallic compounds, however nanoparticle-based materials offer the benefits of tuneable wavelengths and narrow emission profiles. Materials based on cadmium have been the most extensively studied for ECL nanoparticles to date. Cadmium based nanoparticles exhibit high levels of toxicity thereby impacting their suitability for mass produced sensing applications. As such, alternative materials with reduced toxicities are required. This review focuses on the innovations and applications of low toxicity semi-conductor quantum dots (SCQDs) utilised as ECL luminophores within biosensors. These materials include silver, copper, zinc, tin, silicon and germanium. This contribution presents an evaluative overview of these materials for use as ECL luminophores in terms of toxicity, tunability of emission, potential for amplification, and water dispersibility. Capacity for functionalisation and multiplexing potential are also explored.

Electrocatalytic enhancement of [Ru(bpy)3]2+ electrochemiluminescence for gemcitabine detection toward precision measurement via gold nanoparticle addition.

With a rise in the development and subsequent employment of precision medicine, their lies an immediate necessity for the development of technology to enable the implementation of such treatment plans into the healthcare environment. Electrochemistry stands to offer one of the most viable techniques for such technologies given its success within current medical devices. One electrochemical technique, electrochemiluminescence (ECL), warrants investigation. Previously we have determined the inability to reliably detect cancer therapy gemcitabine via traditional ruthenium based ECL. Here we demonstrate how the addition of gold nanoparticles into the ECL film can promote GMB detection via enhanced electrocatalytic oxidation, generating the required ECL radicals. Via this approach we have been able to improve the ECL signal intensity 60-fold and achieve detection down to 6.25 µM across a linear range of 6.25-50 µM. Which lies within the therapeutically relevant range. This approach has successfully addressed the prior limitations encountered for the employment of traditional ruthenium based ECL for substance identification, where species exhibit limited electro-activity and suffer from electrochemically induced side reactions.

Electrochemiluminescence within veterinary Science: A review.

Veterinary science or veterinary medicine is a diverse and significant field. Concerned not only with the diagnosis and treatment of domestic animals and livestock, but it also places focus upon zoonotic diseases, the development and effectiveness of potential vaccines and the possibility of transmission of veterinary medication or viruses into animal food products. Electrochemiluminescence (ECL) is a powerful analytical technique, which despite its significant intrinsic benefits has not seen enormous adoption into the wider analytical chemical community. In contrast, the veterinary science sector has reaped the merit of ECL as far back as the late 90's and continue to benefit from development of the technique a further three decades later. ECL offers the superb sensitivity, low running costs, rapid results and high reliability required within the veterinary science sector, as such its employment in this area shouldn't be surprising. To this end this article aims to summarise the standing of ECL within the veterinary science field, in an attempt increase the awareness of its successful employment within this area to the electro-analytical and wider analytical chemistry communities. Where it is hope veterinary science will gain recognition as possible end user targets for academic and industrial electrochemical researchers.

Development of an Electrochemical CCL17/TARC Biosensor toward Rapid Triage and Monitoring of Classic Hodgkin Lymphoma.

A point-of-care blood test for the detection of an emerging biomarker, CCL17/TARC, could prove transformative for the clinical management of classic Hodgkin lymphoma (cHL). Primary care diagnosis is challenging due to nonspecific clinical presentation and lack of a diagnostic test, leading to significant diagnostic delays. Treatment monitoring encounters false-positive and negative results, leading to avoidable chemotherapy toxicity, or undertreatment, impacting patient morbidity and mortality. Here, we present an amperometric CCL17/TARC immunosensor, based on the utilization of a thiolated heterobifunctional cross-linker and sandwich antibody assay, to facilitate novel primary care triage and chemotherapy monitoring strategies for cHL. The immunosensor shows excellent analytical performance for clinical testing; linearity (R2 = 0.986), detection limit (194 pg/mL), and lower and upper limits of quantitation (387-50 000 pg/mL). The biosensor differentiated all 42 newly diagnosed cHL patients from healthy volunteers, based on serum CCL17/TARC concentration, using blood samples collected prior to treatment intervention. The immunosensor also discriminated between paired blood samples of all seven cHL patients, respectively, collected prior to treatment and during chemotherapy, attributed to the decrease in serum CCL17/TARC concentration following chemotherapy response. Overall, we have shown, for the first time, the potential of an electrochemical CCL17/TARC biosensor for primary care triage and chemotherapy monitoring for cHL, which would have positive clinical and psychosocial implications for patients, while streamlining current healthcare pathways.

Emission from the working and counter electrodes under co-reactant electrochemiluminescence conditions.

We present a new approach to explore the potential-dependent multi-colour co-reactant electrochemiluminescence (ECL) from multiple luminophores. The potentials at both the working and counter electrodes, the current between these electrodes, and the emission over cyclic voltammetric scans were simultaneously measured for the ECL reaction of Ir(ppy)3 and either [Ru(bpy)3]2+ or [Ir(df-ppy)2(ptb)]+, with tri-n-propylamine as the co-reactant. The counter electrode potential was monitored by adding a differential electrometer module to the potentiostat. Plotting the data against the applied working electrode potential and against time provided complementary depictions of their relationships. Photographs of the ECL at the surface of the two electrodes were taken to confirm the source of the emissions. This provided a new understanding of these multifaceted ECL systems, including the nature of the counter electrode potential and the possibility of eliciting ECL at this electrode, a mechanism-based rationalisation of the interactions of different metal-complex luminophores, and a previously unknown ECL pathway for the Ir(ppy)3 complex at negative potentials that was observed even in the absence of the co-reactant.

Electrochemiluminescent screening for methamphetamine metabolites.

The abuse of methamphetamine (MA) is to date detected and subsequently verified through the monitoring of MA and its metabolites within biological specimens. Current approaches require complex sample purification strategies alongside significant analysis time. Given the high prevalence of MA within the global drug market, there remains a need for rapid, portable and alternative screening approaches appropriate for direct detection within biological matrices for employment across the forensic and clinical environments. This contribution illustrates the use of an electrochemiluminescence (ECL) strategy for the screening of MA, amphetamine (AMP) and para hydroxy-methamphetamine (pOH-MA) for such applications. The sensing system showed ideal analytical performance with linear ranges at forensically relevant concentrations of 0.1 µM to 0.5 mM for MA, 10 µM to 1 mM AMP and 10 µM to 5 mM for pOH-MA, and superb detection limits of 74.6 nM, 6 µM and 82. µM for MA, AMP and pOH-MA respectively. Furthermore, the sensor was successful in the detection of MA, AMP and pOH-AMP within human pooled serum, artificial urine and saliva, without any prior purification strategies. Here a portable ECL sensor is detailed for the successful employment of the direct screening of these amphetamine type substances and their corresponding metabolites at clinically and forensically relevant concentrations within a range of biological matrices. This approach successfully represents a strong proof-of-concept, for a novel, simple and rapid screening method with significant potential for high-throughput screening of biological samples for drug metabolites, widening the avenues where ECL sensors could be employed.

Electrochemical monitoring of alcohol in sweat.

Rapid, periodic monitoring and detection of ethanol (EtOH) after consumption via a non-invasive measurement has been an area of increased research in recent years. Current point-of-care or on-site detection strategies rely on single use sensors which are inadequate for monitoring during a longer period. A low cost, portable and novel approach is developed here for real-time monitoring over several days utilising electrochemical techniques. The sensor shows oxidation of the ethanol in phosphate buffer and artificial sweat using the amperometric response from the application of +0.9 V to the polyaniline modified screen printed electrode using 1 mM EtOH as the averaged amount of EtOH eliminated in sweat after the consumption of one alcoholic beverage. Our enzyme based electrochemical sensor exhibits a qualitative assessment of the presence of EtOH in small volumes (≤40 µL) of 0.1 M sodium bicarbonate and subsequently artificial sweat, with 50 measurements taken daily over 11 days. While quantitative information is not obtained, the sensor system exhibits excellent stability after 3 months' dried storage in this complex biological matrix in an oxygen free cabinet. This addresses one of the key challenges for enzyme based electrochemical sensors, namely, the ability for real-time monitoring in complex biological matrices. The qualitative response illustrates the potential for this sensor to be exploited by non-experts which suggests the promise for their wider application in next-generation wearable electronics necessary for alcohol monitoring.

Datura quids at Pinwheel Cave, California, provide unambiguous confirmation of the ingestion of hallucinogens at a rock art site.

While debates have raged over the relationship between trance and rock art, unambiguous evidence of the consumption of hallucinogens has not been reported from any rock art site in the world. A painting possibly representing the flowers of Datura on the ceiling of a Californian rock art site called Pinwheel Cave was discovered alongside fibrous quids in the same ceiling. Even though Native Californians are historically documented to have used Datura to enter trance states, little evidence exists to associate it with rock art. A multianalytical approach to the rock art, the quids, and the archaeological context of this site was undertaken. Liquid chromatography-mass spectrometry (LC-MS) results found hallucinogenic alkaloids scopolamine and atropine in the quids, while scanning electron microscope analysis confirms most to be Datura wrightii Three-dimensional (3D) analyses of the quids indicate the quids were likely masticated and thus consumed in the cave under the paintings. Archaeological evidence and chronological dating shows the site was well utilized as a temporary residence for a range of activities from Late Prehistory through Colonial Periods. This indicates that Datura was ingested in the cave and that the rock painting represents the plant itself, serving to codify communal rituals involving this powerful entheogen. These results confirm the use of hallucinogens at a rock art site while calling into question previous assumptions concerning trance and rock art imagery.

Structural refinement and electrochemical properties of one dimensional (ZnO NRs)1-x(CNs)x functional hybrids for serotonin sensing studies.

Herein, the efficient serotonin (5-HT) sensing studies have been conducted using the (ZnO NRs)1-x(CNs)x nanocomposites (NCs) having appropriate structural and electrochemical properties. Initially, the different compositions of ZnO nanorods (NRs), with varying content of carbon nanostructures (CNs=MWCNTs and RGO), are prepared using simple in-situ wet chemical method and thereafter these NCs have been characterized for physico-chemical properties in correlation to the 5-HT sensing activity. XRD Rietveld refinement studies reveal the hexagonal Wurtzite ZnO NRs oriented in (101) direction with space group 'P63mc' and both orientation as well as phase of ZnO NRs are also retained in the NCs due to the small content of CNs. The interconnectivity between the ZnO NRs with CNs through different functional moieties is also studied using FTIR analysis; while phases of the constituents are confirmed through Raman analysis. FESEM images of the bare/NCs show hexagonal shaped rods with higher aspect ratio (4.87) to that of others. BET analysis and EIS measurements reveal the higher surface area (97.895 m2/g), lower charge transfer resistance (16.2 kΩ) for the ZCNT 0.1 NCs to that of other NCs or bare material. Thereafter, the prepared NCs are deposited on the screen printed carbon electrode (SPCE) using chitosan as cross-linked agent for 5-HT sensing studies; conducted through cyclic voltammetry (CV) and square wave voltammetry (SWV) measurements. Among the various composites, ZCNT0.1 NCs based electrodes exhibit higher sensing activity towards 5-HT in accordance to its higher surface area, lower particle size and lower charge transfer resistance. SWV measurements provide a wide linear response range (7.5-300 µM); lower limit of detection (0.66 µM), excellent limit of quantification (2.19 µM) and good reproducibility to ZCNT 0.1 NCs as compared to others for 5-HT sensing studies.

Electrochemiluminescent sensors as a screening strategy for psychoactive substances within biological matrices.

With the rapid growth and appearance of novel psychoactive substances (NPS) onto the global drug market, the need for alternative screening methodologies for implementation within clinical environments is substantial. The immunoassay methods currently in use are inadequate for this new drug trend with the potential for misdiagnosis and subsequent administration of incorrect patient treatment increased. This contribution illustrates a strong proof-of-concept for the use of electrochemiluminescence (ECL) as a screening methodology for NPS within biological fluids, using the hallucinogen scopolamine as a model compound. A low cost, easy-to-use and portable sensor has been developed and successfully employed for the detection of scopolamine at clinically relevant concentrations within a variety of biological matrices, including human pooled serum, urine, artificial saliva and sweat, without any prior sample preparation required. Moreover, assessment of the sensor's potential as a point-of-care wearable device was performed with sample collection from the surface of skin, demonstrating its capability for the qualitative identification of scopolamine despite collection of only minimal volumes off the skins surface. The developed sensor described herein exhibits a strong proof-of-concept for the employment of such ECL sensors as point-of-care devices, where the sensors ease of use and removal of time-consuming and complex sample preparation methods will ultimately increase its usability by physicians, widening the avenues where ECL sensors could be employed.

Tale of Two Alkaloids: pH-Controlled Electrochemiluminescence for Differentiation of Structurally Similar Compounds.

Electrochemiluminescence (ECL) has increased in popularity as a result of its inherent advantages, including but not limited to portability, simplicity of use, and low reagent consumption. However, its significant advantages are often over shadowed as a result of its limited specificity. ECL emissions are intrinsically broad and lack the definition of other available analytical techniques. Furthermore, species with similar functional groups have almost identical electrochemical behavior and thus typically emit within approximately the same potential region. Within this contribution we have demonstrate the use of pH controlled ECL to prove the presence of two individual species within a mixed sample. Analysis at a single pH would not provide this information. We have illustrated the potential of this methodology to quantify scopolamine alongside sister tropane alkaloid atropine, a known ECL interferent. Previously the two alkaloids could not be distinguished from one another using a single technique which did not involve a separation strategy. pH controlled ECL is a simple approach to improve the specificity of a basic [Ru(bpy)3]2+ film based sensor. By exploiting molecular characteristics, such as pKa, we have been able to fine-tune our methodology to facilitate identification of analytes previously exhibiting indistinguishable ECL emission. Thus, by improving specificity, while maintaining operational simplicity and inexpensive design, we have been able to highlight the potential power of ECL for identification of structurally similar compounds. Further improvements of specificity, such as demonstrated within this contribution, will only further future applications of ECL sensors across a range of different fields.

Evaluation of the one-step Lumicyano™ used in the visualisation of fingermarks on fabrics.

This study consisted of three parts to evaluate the performance of Lumicyano™ on a variety of fabrics. One part assessed the impact of dye percentage (8%, 9% and 10%) on visualisation of fingermark detail and luminescent brightness in split grab marks. A 9% dye produced the highest quality detail of grab impressions with least interference from background fluorescence. The second part investigated the optimal relative humidity (RH, 75-84%) for certain fabric types using Lumicyano on split, six-series depletion fingermarks. It was concluded that the recommended RH of 80% remained the ideal cyanoacrylate fuming environment. The final and third part of this study determined the impact of sequential addition of Basic Yellow 40 (BY40) on Lumicyano compared to traditional cyanoacrylate (CA) followed with BY40 application. Results from this study demonstrated that Lumicyano on its own developed fingermarks with superior quality to Lumicyano with sequential addition of BY40 or traditional cyanoacrylate followed by BY40. Inclusion of more fabrics, donors and longer ageing periods should be explored in future studies to determine what frameworks are best for certain types of fabrics.

Utilization of an Electrochemiluminescence Sensor for Atropine Determination in Complex Matrices.

A major challenge within forensic science is the development of accurate and robust methodologies that can be utilized on-site for detection at crime scenes and can be used for analyzing multiple sample types. The recent expansion of electrochemical sensors to tackle this hurdle requires sensors that can undergo analysis without any pretreatment. Given the vast array of samples that are submitted for forensic analysis, this can pose a major challenge for all electrochemical sensors, including electrochemiluminescent (ECL)-based sensors. Within this contribution, we demonstrate the capacity for an ECL-based sensor to address this challenge and it is potential to detect and quantify atropine from a wide range of samples directly from herbal material to spiked solutions. This portable platform demonstrates satisfactory analytical parameters with linearity across a concentration range of 0.75 to 100 µM, reproducibility of 3.0%, repeatability of 9.2%, and a detection limit of ∼0.75 µM. The sensor displays good selectivity toward alkaloid species and, in particular, the hallucinogenic tropane alkaloid functionality within complex matrices. This portable sensor provides rapid detection alongside low cost and operational simplicity, thus, providing a basis for the exploitation of ECL-based sensors within the forensic arena.

'Cathodic' electrochemiluminescence of [Ru(bpy)3]2+ and tri-n-propylamine confirmed as emission at the counter electrode.

A new approach to examine electrochemiluminescence (ECL), in which the potentials at both the working and counter electrodes are measured and the emitted light is detected by a photomultiplier tube, camera and then a charge-coupled device (CCD) spectrometer, provides unequivocal evidence that the purported cathodic ECL of [Ru(bpy)3]2+ and tri-n-propylamine actually arises from anodic reactions at the counter electrode.

Cathodic Quantum Dot Facilitated Electrochemiluminescent Detection in Blood.

The expansion of electrochemical sensors to biomedical applications at point of care requires these sensors to undergo analysis without any pretreatment or extraction. This poses a major challenge for all electrochemical sensors including electrochemiluminescent (ECL)-based sensors. ECL offers many advantages for biomedical applications; however, obtaining results from complex matrixes has proven to be a large hurdle for the application of ECL sensors within this field. This work demonstrates the potential of cathodic ECL to detect and quantify homocysteine (Hcy) with a 0.1 nM limit of detection, which is associated with hyperhomocysteinemia, in blood. This near-infrared quantum dot (NIR QD)-based ECL sensor displays good linearity allowing for rapid detection and providing a basis for exploitation of ECL-based sensors for biomedical diagnostics utilizing Hcy as a model cathodic coreactant. This work will lay the foundations for future developments in biosensing and imaging fields and stands as an initial proof of concept for the utilization of cathodic ECL technologies for biomedical applications once the limits of detection within clinically relevant levels has been achieved. This work illustrates the potential of cathodic ECL sensors, using Hcy as a model complex, for the detection of biomolecules.

Independent validation of body fluid-specific CpG markers and construction of a robust multiplex assay.

Potential forensic use of tissue-specific DNA methylation markers has recently been discussed for the identification of the biological source of a stain. In this study 13 promising markers were evaluated to identify suitable candidate markers for the development of a robust and reliable multiplex assay. The results of this study suggest that a combination of only four highly informative markers will be enough for clear body fluid identification. A multiplex assay was developed for the identification of menstrual blood, saliva, semen, and venous blood. This assay was successfully applied to the identification of these body fluids in mixtures and crime scene stains. The multiplex assay aids in the identification of not only single source body fluids but also of body fluid mixtures. The main advantage of using DNA methylation assays over alternative tests is that it can be applied at a later time point in the investigative process since testing is possible even after DNA analysis.

A proof of principal study on the use of direct PCR of semen and spermatozoa and development of a differential isolation protocol for use in cases of alleged sexual assault.

Sexual assault samples are some of the most common samples encountered in forensic analysis. These samples can require a significant time investment due to differential extraction processes. We report on the first record of successful direct amplification of semen for STR analysis. Neat seminal fluid, dilutions ranging from 1:5 to 1:160 and GEDNAP samples were successfully amplified using a direct method. A mild differential isolation technique to enrich spermatozoa was developed and successfully implemented to separate and directly amplify a mixture of semen and female epithelial cells. Aliquots of samples subjected to the differential isolation protocol were stained with Haemotoxylin and Eosin for sperm scoring. Samples stained after PCR showed a complete lack of intact spermatozoa demonstrating that the cells are lysed during the PCR process. This paper demonstrates the potential to incorporate direct PCR in cases of sexual assault to more rapidly obtain results and achieve a higher sensitivity.

Optimising electrogenerated chemiluminescence of quantum dots via co-reactant selection.

We demonstrate that for quantum dot (QD) based electrochemiluminescence (ECL), the commonly used co-reactant does not perform as effectively as potassium persulfate. By exploiting this small change in co-reactant, ECL intensity can be enhanced dramatically in a cathodic-based ECL system. However, TPA remains the preferential co-reactant-based system for anodic ECL. This phenomenon can be rationalised through the relative energy-level profiles of the QD to the co-reactant in conjunction with the applied potential range. This work highlights the importance of understanding the co-reactant pathway for optimising the application of ECL to bioanalytical analysis, in particular for near-infrared (NIR) QDs which can be utilised for analysis in blood. Graphical Abstract Optimising ECL Production Through Careful Selection of Co-Reactions Based on Energetics Involved.

The EpiTect Methyl qPCR Assay as novel age estimation method in forensic biology.

Human aging is associated with epigenetic modification of the genome. DNA methylation at cytosines appears currently as the best characterised modification that occurs during the mammalian lifetime. Such methylation changes at regulatory region can provide insights to track contributor age for criminal investigation. The EpiTect Methyl II PCR system (QIAGEN) was used to compare methylation levels of CpG islands in the promoter regions of a number of age related genes, of which four successfully showed changes across the lifespan (NPTX2, KCNQ1DN, GRIA2 and TRIM58). This technique is based on the detection of remaining input genome after digestion with a methylation-sensitive restriction enzyme. This study examined DNA specimens from 80 female subjects of various ages (18-91 years) obtained from blood, using primers designed to flank the studied gene loci. The data obtained from DNA methylation quantification showed successful discrimination among volunteered ages. Overall, the difference between predicted and real age was about 11 years and absolute mean differences (AMD) was only 7.2 years error. We suggest the EpiTect system can be used as fast and simple innovative tool in future forensic age estimation.