An investigative study into an oscillatory reaction in acoustically levitated droplets.

For the first time we have studied an oscillatory chemical reaction (the well-known Belousov-Zhabotinsky (BZ) reaction) in acoustically levitated droplets. Acoustically levitated droplets allow wall-less reaction studies, reduce consumption of sample/reagents, offer high throughput measurements, and enable environmentally friendly chemistry by significantly reducing plastic waste. In this work, microdroplets of the BZ reactants were mixed at the central axis of a low-cost acoustic levitator. The chemical reaction observed in acoustically levitated droplets proceeded in the same way as that in both stirred and unstirred vials where the volume of droplets was 750-fold lower than the solutions in vials. The observed oscillation frequency in droplets was lower than that observed in vials, possibly as a result of evaporative cooling of the droplets. This work has shown that oscillatory reactions can be successfully carried out in acoustically levitated droplets, which allows the application of this technique to areas such as analysis, synthesis and actuation of smart materials and studies of the origins of life.

A study of diffraction-based chitosan leaky waveguide (LW) biosensors.

The waveguide layer of diffraction-based leaky waveguides (LWs) must be made of materials that have low refractive index, are permeable to analytes, can be deposited by spin coating, and can be functionalised and crosslinked. These requirements are fulfilled by thin films of chitosan hydrogels. In this work, we studied the reproducibility of diffraction-based LWs with chitosan waveguides. The average refractive index sensitivity (RIS) and RI limit of detection (LOD) of the eight devices investigated herein were 125.5 ± 3.8 deg RIU-1 and 1.9 × 10-6 ± 1.3 × 10-6 RIU, respectively. While several challenges associated with the realisation of reproducible chitosan LWs have been addressed, reducing the variations in RI LOD requires improving the adhesion of chitosan films to glass substrates, minimising bubbles trapped in microfluidic channels, and using pumps with minimal pulsations. We showed that the buffer baseline of LWs with unmodified chitosan before and after introducing 750 µM bovine serum albumin (BSA), which is equal to the physiological levels of serum albumin, was different by 3.6%. Nevertheless, using biotin, anti-biotin antibody and BSA as exemplar recognition element, analyte and interferent, respectively, we demonstrated that diffraction-based chitosan LWs were suitable for monitoring analyte-RE binding in the presence of 750 µM BSA.

Hydrogel gratings with patterned analyte responsive dyes for spectroscopic sensing.

This is an unprecedented report of hydrogel gratings with an analyte responsive dye immobilised in alternating strips where the patterned dye is its own dispersive element to perform spectroscopy. At each wavelength, the diffraction efficiency of hydrogel gratings is a function of dye absorbance, which in turn is dependent on the concentration of analytes in samples. Thus, changes in intensity of diffracted light of hydrogel gratings were measured for sensing of analytes. Equally, the ratio of diffracted intensities at two wavelengths was used for quantification of analytes to reduce errors caused by variations in intensity of light sources and photobleaching of dyes. 15.27 µm pitch gratings were fabricated by exposing 175 µm thick films of photofunctionalisable poly(acrylamide) hydrogel in a laser interferometric lithography setup, generating an array of alternating lines with and without free functional groups. The freed functional groups were reacted with pH sensitive fluorescein isothiocyanate to create gratings for measurement of pH. The ratio of intensity of diffracted light of hydrogel gratings at 430 and 475 nm was shown to be linear over 4 pH units, which compares favourably with ∼2 pH units for conventional absorption spectroscopy. This increased dynamic range was a result of cancellation of the opposite non-linearities in the pH response of the analyte responsive dye and the diffraction efficiency as a function of dye absorbance.

A Self-Referenced Diffraction-Based Optical Leaky Waveguide Biosensor Using Photofunctionalised Hydrogels.

We report a novel self-referenced diffraction-based leaky waveguide (LW) comprising a thin (~2 µm) film of a photofunctionalisable hydrogel created by covalent attachment of a biotinylated photocleavable linker to chitosan. Streptavidin attached to the chitosan via the photocleavable linker was selectively removed by shining 365 nm light through a photomask to create an array of strips with high and low loading of the protein, which served as sensor and reference regions respectively. The differential measurements between sensor and reference regions were used for measuring analytes (i.e., biotin protein A and IgG) while reducing environmental and non-specific effects. These include changes in temperature and sample composition caused by non-adsorbing and adsorbing species, leading to reduction in effects by ~98%, ~99%, and ~97% respectively compared to the absolute measurements. The novelty of this work lies in combining photofunctionalisable hydrogels with diffraction-based LWs for referencing. This is needed to realise the full potential of label-free optical biosensors to measure analyte concentrations in real samples that are complex mixtures, and to allow for sample analysis outside of laboratories where drifts and fluctuations in temperature are observed.

Optimization Synthesis and Biosensing Performance of an Acrylate-Based Hydrogel as an Optical Waveguiding Sensing Film.

The metal-clad leaky waveguide (MCLW) is an optical biosensor consisting of a metal layer and a low index waveguide layer on a glass substrate. This label-free sensor measures refractive index (RI) changes within the waveguide layer. This work shows the development and optimization of acrylate based-hydrogel as the waveguide layer formed from PEG diacrylate (PEGDA, Mn 700), PEG methyl ether acrylate (PEGMEA, Mn 480), and acrylate-PEG2000-NHS fabricated on a substrate coated with 9.5 nm of titanium. The acrylate-based hydrogel is a synthetic polymer, so properties such as optical transparency, porosity, and hydrogel functionalization by a well-controlled reactive group can be tailored for immobilization of the bioreceptor within the hydrogel matrix. The waveguide sensor demonstrated an equal response to solutions of identical RI containing small (glycerol) and large (bovine serum albumin; BSA) analyte molecules, indicating that the hydrogel waveguide film is highly porous to both sizes of molecule, thus potentially allowing penetration of a range of analytes within the porous matrix. The final optimized MCLW chip was formed from a total hydrogel concentration of 40% v/v of PEGMEA-PEGDA (Mn 700), functionalized with 2.5% v/v of acrylate-PEG2000-NHS. The sensor generated a single-moded waveguide signal with a RI sensitivity of 128.61 ± 0.15° RIU-1 and limit of detection obtained at 2.2 × 10-6 RIU with excellent signal-to-noise ratio for the glycerol detection. The sensor demonstrated RI detection by monitoring changes in the out-coupled angle resulting from successful binding of d-biotin to streptavidin immobilized on functionalized acrylate hydrogel, generating a binding signal of (12.379 ± 0.452) × 10-3°.

Disposable Injection Molded Conductive Electrodes Modified with Antimony Film for the Electrochemical Determination of Trace Pb(II) and Cd(II).

This work describes a novel electrochemical sensor fabricated by an injection molding process. This device features a conductive polymer electrode encased in a plastic holder and electroplated in situ with a thin antimony film. The antimony film sensor was applied to the determination of Pb(II) and Cd(II) by anodic stripping voltammetry (ASV). The deposition of Sb on the sensor was studied by cyclic voltammetry (CV) and microscopy. The experimental variables (concentration of the antimony plating solution, deposition potential and time, stripping waveform) were investigated, and the potential interferences were studied and addressed. The limits of detection were 0.95 µg L-1 for Pb(II) and 1.3 for Cd(II) (at 240 s of preconcentration) and the within-sensor percentage relative standard deviations were 4.2% and 4.9%, respectively, at the 25 µg L-1 level (n = 8). Finally, the sensor was applied to the determination of Pb(II) and Cd(II) in a phosphorite sample and a lake water sample.

A feasibility study of a leaky waveguide aptasensor for thrombin.

This proof-of-principle study demonstrates the feasibility of a leaky waveguide (LW) aptasensor, where aptamers were immobilised in a mesoporous chitosan waveguiding film for the detection of thrombin. This work has demonstrated that aptamers immobilised in hydrogels retain their affinity and selectivity towards their target and thus can be used as bioreceptors. The use of antibodies as bioreceptors for sensing thrombin is not viable because it is a serine protease, which will cleave the antibodies. Currently used assays based on clotting time and chromogenic/fluorogenic substrates have limited potential for thrombin measurement in whole blood. Using the initial binding rate over the first 5 min, the limit of detection of our LW aptasensor for thrombin was ∼22 nM. The sensor was tested with spiked serum samples, giving a reading of 46.1 ± 4.6 nM for a sample containing 50 nM thrombin. Our proposed sensor combines the robustness and low cost of aptamers as molecular recognition elements with the simple fabrication process of the chitosan-based leaky waveguide, making LW aptasensors highly attractive for applications in point-of-care diagnostics and healthcare monitoring.

Method for Determining Average Iron Content of Ferritin by Measuring its Optical Dispersion.

We report a method where the refractive index increments of an iron storage protein, ferritin, and apoferritin (ferritin minus iron) were measured over the wavelength range of 450-678 nm to determine the average iron content of the protein. The protein used in this study had ∼3375 iron atoms per molecule. The measurement of optical dispersion over the broad wavelength range was enabled by the use of mesoporous leaky waveguides (LWs) made of chitosan. We present a facile approach for fabricating mesoporous chitosan waveguides for improving the measurement sensitivity of macromolecules such as ferritin. Mesoporous materials allow macromolecules to diffuse into the waveguide, maximizing their interaction with the optical mode and thus increasing sensitivity by a factor of ∼9 in comparison to nonporous waveguides. The sensitivity was further improved and selectivity toward ferritin was achieved by the incorporation of antibodies in the waveguide. The method presented in this work is a significant advance over the state of the art method, the enzyme linked immunosorbent assay (ELISA) used in clinics, because it allows determining the average content of ferritin in a single step. The average iron content of ferritin is an important marker for conditions such as injury, inflammation, and infection. Thus, the approach presented here of measuring optical dispersion to determine the average iron content of ferritin has a significant potential to improve the point of care analysis of the protein for disease diagnosis and screening.

Over two decades of orthopaedic surgery in patients with inhibitors-Quantifying the complication of bleeding.

Patients with haemophilia who have developed inhibitors against factor VIII (FVIII) or factor IX present a significant concern to those surgeons who operate on them. The evidence base for bypassing agents such as recombinant factor VIIa and activated prothrombin complex concentrate has amassed over several decades. The literature is open to positive interpretation on the successful use of these agents in the treatment of inhibitor-positive patients. However, there are equally persistent concerns amongst surgeons, in particular orthopaedic surgeons, regarding the high complication rate of bleeding. To explore and quantify this concern, we present a literature review spanning two decades of publications on haemophilia patients with inhibitors undergoing orthopaedic surgery. Irrespective of the progress made with haemostatic protocols, trepidation on embarking on surgery is valid. The high risk of bleeding is a function of the inherent complexity of the disease and rightfully translates into difficulties in its management. Combined with the prospect of orthopaedic surgery, those involved in the care of such patients are justified in their continued anxiety and diligence when considering the benefits in quality of life against the prevalent complications.

Microfluidic channel-assisted screening of hematopoietic malignancies.

A simple microfluidic fluorescence in situ hybridization (FISH) device allowing accurate analysis of interphase nuclei in 1 hr in narrow channels is presented. Photolithography and fluorosilicic acid etching were used to fabricate microfluidic channels (referred to as FISHing lines) that allowed analysis of 10 samples on a glass microscope slide 0.2 µl of sample volume was used to fill a micro-channel, which resembled a 250-fold reduction compared to conventional FISH. FISH signals were comparable to conventional FISH, with 50-fold less probe consumption and 10-fold less time. Cells were immobilized in single file in channels just exceeding the diameter of the cells, and were used for minimal residual disease (MRD) analysis. To test the micro-channels for application in FISH, MRD was simulated by mixing K562 cells (an established chronic myeloid leukemia cell line) carrying the BCR/ABL fusion gene across 1:1 to 1:1,000 Jurkat cells (an established acute lymphoblastic leukemia cell line). The limit of detection was seen to be 1:100 cells and 1:1,000 cells for FISHing lines and conventional FISH, respectively; however, the conventional method seemed to over-score the presence of K562 cells. This may in part be attributed to FISHing lines practically eliminating the chance of duplicate screening of cells and hastened the time of screening, enhancing scoring of all cells within the channels. This was compared to 1 in 500 cells on the slide being analyzed with the conventional FISH.

Miniaturised free flow isotachophoresis of bacteria using an injection moulded separation device.

A new design of miniaturised free flow electrophoresis device has been produced. The design contains a separation chamber that is 45 mm long by 31.7 mm wide with a depth of 50 µm and has nine inlet and nine outlet holes to allow for fraction collection. The devices were formed of polystyrene with carbon fibre loaded polystyrene drive electrodes and produced using injection moulding. This means that the devices are low cost and can potentially be mass produced. The devices were used for free flow isotachophoresis (FFITP), a technique that can be used for focussing and concentrating analytes contained within complex sample matrices. The operation of the devices was demonstrated by performing separations of dyes and bacterial samples. Analysis of the output from FFITP separations of samples containing the bacterium Erwinia herbicola, a biological pathogen, by cell culturing and counting showed that fractionation of the output was achieved.

Dupuytren's Disease: Review of the Current Literature.

Dupuytren's disease is one of the most common condition seen by hand surgeons. It is not only prevalent but can also be a most debilitating condition resulting in significant loss of function of the fingers involved. The cause of this disease, however still remains largely unknown although some recent evidence suggests a stem cell etiology. This review article summarizes the current known knowledge of Dupuytren's as well as the clinical findings, investigations and treatments available.

Thiocyanate and nitrite analysis using miniaturised isotachophoresis on a planar polymer chip.

A new method has been developed to improve the determination of thiocyanate using isotachophoresis. This method uses complexation with copper(II) as a mechanism for improving the separation of thiocyanate from chlorate and perchlorate. By using a pH of 3.25 the method can also be used to analyse nitrite. Separations were carried out using a miniaturised poly(methyl methacrylate) (PMMA) separation device. Linearity was observed from 1.25 to 75 mg dm(-3) with a correlation coefficient of 0.998 for both thiocyanate and nitrite. Limits of detection for these two species were calculated to be 0.8 mg dm(-3) and 0.9 mg dm(-3) respectively. The method was successfully applied to the analysis of these anions in a range of samples including explosive residues.

Strength of interactions between immobilized dye molecules and sol-gel matrices.

In this paper we present a new theory to re-examine the immobilization technique of dye doped sol-gel films, define the strength and types of possible bonds between the immobilized molecule and sol-gel glass, and show that the immobilized molecule is not free inside the pores as was previously thought. Immobilizing three different pH sensitive dyes with different size and functional groups inside the same sol-gel films revealed important information about the nature of the interaction between the doped molecule and the sol-gel matrix. The samples were characterized by means of ultraviolet-visible spectrophotometer (UV-VIS), thermal gravimetric analysis (TGA), mercury porosimetry (MP), nuclear magnetic resonance spectroscopy ((29)Si NMR) and field-emission environmental scanning electron microscopy (ESEM-FEG). It was found that the doped molecule itself has a great effect on the strength and types of the bonds. A number of factors were identified, such as number and types of the functional groups, overall charge, size, pK(a) and number of the silanol groups which surround the immobilized molecule. These results were confirmed by the successful immobilization of bromocresol green (BCG) after a completely polymerized sol-gel was made. The sol-gel consisted of 50% tetraethoxysilane (TEOS) and 50% methyltriethoxysilane (MTEOS) (w/w). Moreover, the effect of the immobilized molecule on the structure of the sol-gel was studied by means of a leaky waveguide (LW) mode for doped films made before and after polymerization of the sol-gel.

Determination of the potassium content of explosive residues using miniaturised isotachophoresis.

A new method has been developed to allow the determination of potassium in post-explosion residues to be made using miniaturised isotachophoresis. The method is based on the use of a caesium leading ion with 4.5 mM 18-crown-6 ether added to retard the potassium to allow reliable determinations to be made. With the conditions selected no interference was noted from other small inorganic cations, such as ammonium, barium, calcium, magnesium, sodium or strontium. The method was successfully applied to the analysis of seven samples containing explosive residues obtained from the unconfined burning of several flash powders. The procedure was found to offer good linearity for potassium determinations over the concentration range of 1.25-150 µg/mL with a coefficient of determination of 0.999 achieved.

Optimisation of secondary electrospray ionisation (SESI) for the trace determination of gas-phase volatile organic compounds.

An electrospray ionisation triple quadrupole mass spectrometer (Varian 1200 L) was modified to accept nitrogen samples containing low concentrations of volatile organic compounds. Six candidate probe compounds, methyl decanoate, octan-3-one, 2-ethylhexanoic acid, 1,4-diaminobutane, dimethyl methylphosphonate, and 2,3-butanediol, at concentrations below 50 ppb(v) were generated with permeation tubes in a test atmosphere generator. The concept of using a set of molecular probes to evaluate gas-phase electrospray ionisation of volatile analytes was assessed and the feasibility of adopting a unified ionisation approach for gas and liquid contamination of exobiotic environments established. 450 experiments were run in a five-replicate, fifteen-level, three-factor, central-composite-design with exponential dilution for each of the six probe compounds studied. The three factors studied were ionisation voltage, drying-gas flow and nebulising-gas flow. Parametric modelling by regression analysis enabled the differences in the ionisation behaviours of the probe compounds to be described by the optimisation models. Regression coefficients were in the range 0.91 to 0.99, indicating satisfactory levels of precision in the optimisation models. A wide range in ionisation efficiency was observed, with different optimised conditions required for the probe compounds. It was evident that no one factor appeared to dominate the response and the different factors produced different effects on the responses for the different molecules. 1,4-Butanediamine and dimethyl methylphosphonate required significantly lower ionisation voltages (1.2 kV) than the other four, which achieved optimised sensitivity towards the maximum voltage used in this design (5 to 6 kV). Drying-gas flow rates were found to be more important than nebulising-gas flow rates. However, variations in the constant term B(0) in the optimisation models indicated that other factors, not included in this study, were also likely to be involved in the ionisation process. Electrolyte-flow rate and ionisation temperature were proposed for follow up studies. Exponential dilution data indicated sensitive and analytically useful responses in the target range of 5 to 50 ppb(v) for all six compounds. Significantly, responses were seen at concentrations significantly below 5 ppb(v), with sub ppt(v) responses observed for 1,4-butanediamine, 2-ethylhexanoic acid, dimethylmethylphosphonate, and 1,3-butanediol. Responses in the ppt(v) to ppb(v) range were observed for the remaining two compounds. The observations from this study demonstrated the utility of adopting a set of probe compounds to evaluate electrospray ionisation performance for volatile organic compound based assays; indicated the existence of multiple ionisation mechanisms; and revealed potential sensitivity at the parts per quadrillion level ppq(v).

Is there Light at the End of the Tunnel? Controversies in the Diagnosis and Management of Carpal Tunnel Syndrome.

Carpal tunnel syndrome is a common disorder responsible for considerable patient suffering and cost to health services. Despite extensive research, controversies still exist with regards to best practice in diagnosis, treatment, and service provision. Current best practise would support the use of history, examination and electro-diagnostic studies. The role for ultrasound scanning in diagnosis of carpal tunnel syndrome is yet to be proven. It appears magnetic resonance image scanning has a role where a rare cause for carpal tunnel syndrome may be suspected and also in the detailed reconstruction of the anatomy to aid endoscopic procedures. Treatment options can be surgical or non-surgical and patient choice will dictate the decision. For non-surgical interventions many options have been trialled but until now only steroid use, acupuncture, and splinting have shown discernable benefits. Open surgical decompression of the carpal tunnel appears to be more simple and cost-effective than minimally invasive interventions. For those patients who reject surgery, splinting, acupuncture, and steroid injection can play a role. Recent work looking at different service delivery options has shown some positive results in terms of decreasing patient waiting time for definitive treatment. However, no formal cost-effectiveness analysis has been published and concerns exist about the impact of a stream-lined service on surgical training. In this review, we look at the different diagnostic and treatment options for managing carpal tunnel syndrome. We then consider the different service delivery options and finally the cost-effectiveness evidence.

A miniaturised isotachophoresis method for magnesium determination.

The use of malonic acid as a complexing agent has enabled a new method to be devised to allow the determination of magnesium to be made using miniaturised isotachophoresis. Using a leading electrolyte of 10 mmol L(-1) caesium hydroxide and 2 mmol L(-1) malonic acid at pH 5.1 gave the method a high specificity towards magnesium. Investigations using a poly(methyl methacrylate) chip device with an integrated conductivity detector showed that no interference from calcium, strontium, barium and sodium should occur. The method was found to be linear over the range of magnesium concentrations from 0.625 to 75 mg L(-1) and the limit of detection was calculated to be 0.45 mg L(-1). Separations were demonstrated with water samples but the procedure should also be applicable to more complex sample matrices such as inorganic explosive residues, blood or urine.

Label-free detection with the resonant mirror biosensor.

The resonant mirror (RM) biosensor is a leaky waveguide-based instrument that uses the evanescent field to probe changes in the refractive index at the sensing surface.The RM can therefore be used to monitor in real-time and label-free the interaction between an analyte in solution and its biospecific partner immobilized on the waveguide surface.The RM has been used in studying the interaction of a variety of moieties including proteins, carbohydrates, cells, nucleic acids and receptors, leading to applications in areas such as clinical diagnostics, homeland security, and pharmaceutical and biomolecular interactions. This chapter will review the principle of this biosensor, and the recent advances in instrumentation, different immobilization chemistries, and kinetic studies, as well as some applications.