Search

The monitoring of marine dinophysistoxin okadaic acid in seawater can serve as an early alert system for preventing the potential negative effects this toxin can have on the food industry and human health in general. A disposable sensor system for electrochemical detection of this toxin has been developed using screen-printed electrodes. The method described is based on the inhibition of protein phosphatase type 2A by okadaic acid, evaluating the enzyme activity. p-Nitrophenyl phosphate, 4-methylumbelliferyl phosphate and phenyl phosphate have been tested as substrates achieving good limits of detection of 2.7·10–12 M of okadaic acid. The proposed method has been successfully applied to okadaic acid determination in seawater samples. A study of divalent cations present in seawater that can interfere with the measurement has been carried out. The electrode systems were printed on both rigid and textile backing materials to observe the influence of those materials on the final performance of the sensor., Ministerio de Ciencia e Innovación (MICINN) (TEC2013-405661-P) Ministerio de Economía y Competitividad (MINECO) and European Regional Development Fund (FEDER) (MUSSEL RTC-2015-4077-2).

This work describes the development of a disposable and wearable screen-printed electrochemical sensor for the determination of chloride ions. The sensor includes three screen-printed electrodes: a working, an auxiliary (both carbon paste electrodes), and an Ag/AgCl pseudo-reference electrode. The sensor works by measuring the Nernstian shift of the voltammetric peak potential of ferrocenemethanol in the presence of chloride ions, due to the use of a pseudo-reference electrode in the electrodic system. The measurements are easily taken by dropping the sample onto the electrodic surface. Different flexible materials, including textiles, were studied as substrates of the electrodic system. A study of ink adherence and conductivity in these materials was carried out. The use of Gore-Tex® displayed good ink adherence, acceptable conductivity and waterproof properties. These features make it an excellent support for wearable sensors. The reproducibility and repeatability, in terms of relative standard deviation values of the developed Gore-Tex sensor, were below 3% (n = 5) and the detection limit was 0.2 mM (α = β = 0.05). The sensor was successfully applied to determine the chloride content in sea water.

Combining the advantages of the biosensor field with the problem of detecting Human Cytomegalovirus (HCMV) in human samples, an inexpensive, simple and disposable electrochemical immunosensor for glycoprotein B detection in urine is proposed. Glycoprotein B has been chosen once is the dominant antigen of HCMV. The approach is based on a sandwich-type immunoassay, with the HCMV glycoprotein B sandwiched between the Anti-HCMV antibody adsorbed onto the working electrode, and the Anti-HCMV labeled with gold nanoparticles. Glycoprotein B detection was carried out through electrochemical stripping analysis of silver nanoparticles quantitatively deposited on the immunosensor through catalysis by the nanogold labels. The influence of different steps in the immunosensor construction, namely the silver deposition time, silver enhancer concentration, antibody concentration, BSA concentration and glycoprotein B incubation time, were examined and optimized. The method showed a linear dependence between glycoprotein B concentration and the corresponding anodic stripping peak current, resulting in detection limits of 3.3±1.7ng/mL for samples prepared in buffer and 3.2±0.2ng/mL for urine samples, suggesting that the biological matrix does not interfere with the immunosensor detection capability. Given its mode of preparation, by physical adsorption of the capture antibody in the working electrode, the immunosensor also exhibited an acceptable reproducibility, with a residual standard deviation of 13.5% for samples prepared in buffer and 11.2% for urine samples, thereby presenting a promising development potential for clinical applications.

This work reports a simple voltammetric method for the determination of chloride, bromide, and iodide ions using screen-printed carbon electrodes modified with silver nanoparticles electrochemically deposited on the working electrode surface. UV/Vis absorption spectroelectrochemistry was used to study the electrodeposition of silver nanoparticles on the working carbon electrode on PET or Gore-Tex® supports, and their subsequent oxidation in the presence of halide ions. The main figures of merit of the developed sensors, such as reproducibility and detection limit, have been calculated. Reproducibility values of 2.22%, 2.83% and 3.23% were obtained for chloride, bromide and iodide determinations, respectively. Additionally, the lowest detected amount of chloride, bromide and iodide ions were 3.0·10−6 M, 5.0·10−6 M and 5.0·10−6 M, respectively. Taking into account the relevance of the determination of chloride ion concentration in sweat, the voltammetric method for the determination of halides has been successfully transferred to a Gore-Tex® support to build a first approach to a wearable sensor that facilitates the quantification of this ion in sweat samples. The Gore-Tex® sensor provides a good reproducibility (RSD = 1.61%)., Spanish Ministry of Science and Innovation (MICINN), Ministry of Economy and Competitiveness (MINECO) and the European Regional Development Fund (TEC20013-40561-P and MUSSEL RTC-2015-4077-2).

This paper presents a chronoamperometric method to determine tungsten in water using screen-printed carbon electrodes modified with gold nanoparticles and cross linked alkaline phosphatase immobilized in the working electrode. Enzymatic activity over 2-phospho-L-ascorbic acid trisodium salt, used as substrate, was affected by tungsten ions, which resulted in a decrease of chronoamperometric current, when a potential of 200 mV was applied on 10 mM of substrate in a Tris HCl buffer pH 8.00 and 0.36 M of KCl. Calibration curves for the electrochemical method validation, give a reproducibility of 5.2% (n = 3), a repeatability of 9.4% (n = 3) and a detection limit of 0.29 ± 0.01 μM. Enriched tap water, purified laboratory water and bottled drinking water, with a certified tungsten reference solution traceable to NIST, gave a recovery of 97.1%, 99.1% and 99.1% respectively (n = 4 in each case) and a dynamic range from 0.6 to 30 μM. This study was performed by means of a Lineweaver–Burk plot, showing a mixed kinetic inhibition., Vicerrectorìa de Investigación de la Universidad de Costa Rica (Project 804-B2-297), via Junta de Castilla y León (BU212A12-2), Ministerio de Ciencia e Innovación (MICINN) (TEC2009-12029) and MICINN and Fondo Europeo de Desarrollo Regional (FEDER) (INNPACTO SERIBIO 2011-2014).

An anodic stripping voltammetric method is reported in this study for the determination of sub-nanomolar Pb concentration using disposable sensors, each consisting of three (counter, working and reference) screen-printed electrodes. Sensor performance was optimized for the determination of Pb through several surface modifications, by using single-walled carbon nanotubes, electro-reduced graphene oxide and gold nanoparticles. A scanning electron microscopy study of the deposition of electrogenerated gold nanoparticles of various sizes on the working electrode surface showed that spherical nanoparticles of around 100 nm provided the best results. The modification of working electrodes with graphene and gold nanoparticles permitted the determination of Pb2+ in seawater (Detection Limit: 3.21·10−10 M) without modifying the pH of the sample. The electrode systems were printed on both rigid and textile backing materials, to observe the influence of those materials on the final performance of the sensor.

Putrescine oxidase (PUO) from Micrococcus rubens has been characterized, as a biocatalytic component for the development of electrochemical biosensors for the determination of putrescine. PUO was immobilized by cross-linking on screen-printed carbon electrodes (SPCEs) containing tetrathiafulvalene (TTF) as mediator (TTF/SPCEs). Isothermal titration calorimetry was used for investigation of kinetic properties of immobilized PUO. These PUO/TTF/SPCEs showed a linear response range from 10 a 74 µM, a capability of detection of 10.1±0.6 µM (n=4, α=β=0.05) and a precision of 6.7 %. Octopus and zucchini samples have been analysed using HPLC as a reference technique.

We report on the amperometric determination of sulfite using screen-printed carbon electrodes (SPCEs) modified with gold and silver nanoparticles that were deposited on the electrode to improve the capabilities of detection. The electrode is fairly selective and responds to sulfite with an oxidation current (at 300 mV and pH 6) in the 9.80 to 83.33 μM concentration range. The precision in terms of repeatability and reproducibility is 14.4 % and 10.7 % in the case of SPCEs modified by gold nanoparticles. The method was applied to the determination of sulfite in drinking water, pickle juice and vinegar. Recoveries ranged from 96 % to 104 %.

We have developed screen-printed carbon electrodes for the determination of putrescine (Put) via the enzyme monoamine oxidase that was immobilized on the surface of the electrode by cross-linking it with bovine serum albumin using glutaraldehyde. A mixture of 5 % of tetrathiofulvalene (TTF) and carbon ink was used for the fabrication of the screen-printed working electrode. Put was amperometrically detected by measurement of the current due to the oxidation of the mediator TTF. The use of TTF lowers the working potential to +250 mV (vs. a screen-printed Ag/AgCl reference electrode). Response is linear in the range from 16 to 101 μM, and the detection limit is 17.2 ± 4.6 μM, with a reproducibility of 9.6 % (n = 4) in terms of relative standard deviation. The effects of potentially interfering biogenic amines such as cadaverine, histamine, spermine, spermidine and tryptamine were also evaluated. The biosensor was successfully applied to the determination of Put in zucchini and anchovies.

A chronoamperometric assay for vanadium ions, based on the inhibition of the enzyme alkaline phosphatase, is reported. Screen-printed carbon electrodes modified with gold nanoparticles were used as transducers for the immobilization of the enzyme. The enzymatic activity over riboflavin-5-monophosphate sodium salt is affected by vanadium ions, which results in a decrease in the chronoamperometric current registered. The assay has a detection limit of 3.9 ± 0.4 μM, a repeatability of 7.6% ( n = 4) and a reproducibility of 11.6% ( n = 4). A study of the possible interferences shows that the presence of Mo (VI) may affect vanadium determination. The method was successfully applied to the determination of vanadium in tap water.

The work describes an amperometric bienzymatic biosensor for gluconic acid based on the coimmobilization of gluconate kinase and 6-phospho- d -gluconate dehydrogenase by crosslinking with glutaraldehyde and bovine serum albumin onto screen-printed carbon electrodes. Experimental design methodology has been used to optimize immobilization and operational conditions. This method shows a reproducibility of 2.94% (n = 8) related to the slopes of calibration curves performed in the concentration range from 7.46 to 71.43 μM, as well as an average capability of detection of 7.46 for a probability of false positive and negative of 0.05. Finally, the developed biosensors have been successfully applied to gluconic acid determination in red wines. This approach represents an interesting alternative to be routinely used at a winery.

A novel amperometric biosensor for the determination of Al(III) based on the inhibition of the enzyme superoxide dismutase has been developed. The oxidation signal of epinephrine substrate was affected by the presence of Al(III) ions leading to a decrease in its amperometric current. The immobilization of the enzyme was performed with glutaraldehyde on screen-printed carbon electrodes modifiedwith tetrathiofulvalene (TTF) and different types ofnanoparticles. Nanoparticles of gold, platinum, rhodium and palladium were deposited on screen printed carbon electrodes by means of two electrochemical procedures. Nanoparticles were characterized trough scanning electronic microscopy, X-rays fluorescence, and atomic force microscopy. Palladium nanoparticles showed lower atomic force microscopy parameters and higher slope of aluminum calibration curves and were selected to perform sensor validation. The developed biosensor has a detection limit of 2.0 0.2 Mfor Al(III), with a reproducibility of 7.9% (n = 5). Recovery of standard reference material spiked to buffer solution was 103.8% with a relative standard deviation of 4.8% (n = 5). Recovery of tap water spiked with the standard reference material was 100.5 with a relative standard deviation of 3.4% (n = 3). The study of interfering ions has also been carried out., Vice-presidency for Research at the University of Costa Rica (Project 804-B5-117), Ministerio de Ciencia e Innovación (MICINN Spain) and Fondo Europeo de Desarrollo Regional (FEDER) (Projects :TEC-TEC20013-40561-P and MUSSEL RTC-2015-4077-2).

We have developed screen–printed carbon electrodes for the determination of tyramine (Tyr) via plasma amine oxidase. The enzyme was immobilized on the carbon working electrode by cross–linking it with bovine serum albumin using glutaraldehyde. The employment of the mediator hydroxymethylferrocene lowers the working potential to +260 mV (vs. a screen–printed Ag/AgCl reference electrode). The effects of pH, potential and mediator concentration were optimized and resulted in reproducibility and repeatability values of 8.6 % and 8.7 %, respectively. Response is linear in the range from 2 to 164 μM, and the limit of detection is 2.0 ± 0.18 μM. The effects of potentially interfering biogenic amines such as putrescine, cadaverine, histamine, spermine, spermidine and tryptamine were also evaluated. The biosensor was successfully applied to the determination of Tyr in cheese.

We report on an amperometric assay for Al(III) ions that is based on the inhibition of the enzyme α-chymotrypsin. Screen-printed carbon electrodes modified with gold nanoparticles were used as solid supports for the immobilization of the enzyme. The amperometric response of the synthetic enzyme substrate substrate N-benzoyl-L-tyrosine ethyl ester is affected by Al(III) ions, and this leads to a decrease in the amperometric oxidation current. The assay has a detection limit of 3.3 μM of Al(III). The repeatability and reproducibility of the method are 6.9% (n = 3) and 6.4% (n = 5), respectively. Main interferents include Mo(VI), W(VI) and Fe(III) ions. The method was successfully applied to the determination of Al(III) in tap water.

An enzymatic amperometric procedure for the direct measurement of As(V) in the presence of As(III) was developed. The method is based on the inhibitive action of this species on acid phosphatase enzyme (AcP) activity. Screen-printed carbon electrodes (SPCEs) were used as support for the cross-linking immobilization of the enzyme AcP. 2-Phospho-l-ascorbic acid was used as a novel substrate, in arsenic determination, which amperometric response decreased by the presence of As(V) ions. The optimum working conditions were found using experimental design methodology. Under these conditions, repeatability and reproducibility of the constructed biosensors were determined, reaching values below 8% in terms of residual standard deviation. The capability of detection obtained for As(V) was 0.11 μM for AcP/SPCE biosensors. Analysis of the possible effect of the presence of foreign ions in the solution was performed. The method was successfully applied to the determination of the As(V) content in a ground water sample.

This work describes the development of a miniaturized potentiometric system comprising a miniaturized quasi-reference electrode (QRE) coupled to a solid-state ion-selective electrode (ISE) for the monitoring of pH. We describe the optimization of materials and fabrication processes including screen-printing (SP), electrode treatments (thermal and electrochemical) and the formulation and deposition of an ion-selective membrane (ISM), to obtain a system compliant with biomedical specifications. We developed a potentiometric system composed of an Ag/AgCl QRE and a pH-electrode (ISM deposited on a graphite electrode) that can be used continuously for a period of not less than 7 days in aqueous solutions. Curing the Ag/AgCl pastes during 20 minutes at 120 °C after printing allowed the QREs to display excellent potential stability, as demonstrated by an open-circuit-potential standard deviation of ±1.2 mV over a period of 7 days (n=3 samples). Promoting adhesion of the pH membrane over graphite electrodes improved the pH-electrode performance. This was achieved through a combination of thermal treatment and electrochemical activation of the electrodes by cyclic voltammetry (CV). The final device integrated both the QRE and the pH-electrode, and displayed an average pH sensitivity of −60.8±1.7 mV per pH unit, over a pH range of 7.00 to 7.63.

Introduction: Multiple Myeloma (MM) is an incurable disease. In young patients, autologous bone marrow transplantation (ABMT) remains a cornerstone treatment after induction therapy. Induction therapy has varied during time, from alkylating polychemotherapy (VBAD,VCMP) or VAD chemotherapy (AVAD) to Velcade-Dexametasone based regimens (VD). We present results of follow-up of a large cohort of patients treated with ABMT. We described overall survival (OS; from transplant to death by any cause) and progression free survival (PFS; from transplant to death by any case or progressive disease defined by reappearance by inmunofixation, or duplication of monoclonal peak after ABMT) , and the impact of induction therapy regiments. Patients: 183 patients transplanted from 2002 to 2017. The median age of the patients was 59 years (33-72). Before 2008 all the patients were treated in alkylating based chemotherapy (42 patients). After 2008 patients were treated with VD based regimens (141patients). Only 12 patients received maintenance therapy based in PETHEMA trials 2005 and 2012. No one patient received a planed second transplant; only 32 patients received a second transplant after relapse as consolidation therapy. Results: Median follow-up of patients still alive is 3.65 years (0.15-14.77). Median OS of all patients was 9.12 years (95% confidence interval (CI): 6.28-NR); Median PFS was 3.02 years(95% CI: 2.46-3.76). At 13 years only 2% of patients remains progression free (CI: 0.00-17%). There were significant differences between patients treated before and after VD regimens. The median OS of patients treated with APVAD was significantly shorter compared to VD (6.22 years, CI[3.39-12] vs. NR, CI[6.28-NR], p=0.025) (HR=0.49, p=0.01). Conclusions: VD schemes of induction before ABMT have improved remarkably OS inpatients with Myeloma; nonetheless, plateau is not observed in EFS. Further analysis must address if EFS could represent a strong indicator of OS, mainly due to novel effective salvage therapies after relapse/refractoriness could be a confounding factor. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

Disposable screen-printed biosensors have been successfully employed in the development of analytical methods that respond to the growing need to perform rapid “in situ” analyses. Thus, the early detection of microorganisms, which plays an important role in the prevention of human health problems, animals and plants epidemics, has been carried out using this kind of devices. Moreover, microorganisms have been used as biological sensing elements in the development of sensitive microbial biosensors for the analysis of different analytes of interest. This review presents the electrochemical application of disposable screen-printed biosensors in the microbiology field.

Enzymatic amperometric procedures for measuring arsenic, based on the inhibitive action of this metal on acetylcholinesterase enzyme activity, have been developed. Screen-printed carbon electrodes (SPCEs) were used with acetylcholinesterase covalently bonded directly to its surface. The amperometric response of acetylcholinesterase was affected by the presence of arsenic ions, which caused a decrease in the current intensity. The experimental optimum working conditions of pH, substrate concentration and potential applied, were established. Under these conditions, repeatability and reproducibility of biosensors were determined, reaching values below 4% in terms of relative standard deviation. The detection limit obtained for arsenic was 1.1 × 10−8 M for Ach/SPCE biosensor. Analysis of the possible effect of the presence of foreign ions in the solution was performed. The method was applied to determine levels of arsenic in spiked tap water samples., Junta de Castilla y León (BU022A07, Q0968272E) and the Ministerio de Ciencia e Innovación (TEC2008-01367/TEC) is gratefully acknowledged.

Three-electrode configuration chips containing a Pt, Au and a screen-printed Ag/AgCl as counter, working and reference electrode, respectively, have been developed. Selective determination of Phenobarbital (PB) has been carried out by Cytochrome P450 2B4 (CYP450) immobilization into a polypyrrole matrix onto the gold working electrode. Chronoamperometric experiments show a PB diffusion coefficient of 2.42x10(-6)cm(2)s(-1), a reproducibility and repeatability in terms of residual standard deviation (RSD) of 13% and 5.51%, respectively, and a limit of detection (LOD) of 0.289 micromol dm(-3) (alpha=beta=0.05) for the developed CYP450-biosensor chip. Its performance has been showed by the determination of PB in pharmaceutical drugs. HPLC has been used as reference technique.

The tumoral clone of Waldenström's macroglobulinemia (WM) shows a wide morphological heterogeneity, which ranges from B lymphocytes (BL) to plasma cells (PC). By means of genome-wide expression profiling we have been able to identify genes exclusively deregulated in BL and PC from WM, but with a similar expression pattern in their corresponding cell counterparts from chronic lymphocytic leukemia (CLL) and multiple myeloma (MM), as well as normal individuals. The differentially expressed genes have important functions in B-cell differentiation and oncogenesis. Thus, two of the genes downregulated in WM-BL were IL4R, which plays a relevant role in CLL B-cell survival, and BACH2, which participates in the development of class-switched PC. Interestingly, one of the upregulated genes in WM-BL was IL6. A set of four genes was able to discriminate clonal BL from WM and CLL: LEF1 (WNT/beta-catenin pathway), MARCKS, ATXN1 and FMOD. We also found deregulation of genes involved in plasma cell differentiation such as PAX5, which was overexpressed in WM-PC, and IRF4 and BLIMP1, which were underexpressed. In addition, three of the target genes activated by PAX5 - CD79, BLNK and SYK - were upregulated in WM-PC. In summary, these results indicate that both PC and BL from WM are genetically different from the MM and CLL cell counterpart.

This work presents the simultaneous determination of cadaverine, histamine, putrescine and tyramine by square wave voltammetry using a boron-doped diamond electrode. A multivariate calibration method based on partial least square regressions has allowed the resolution of the very high overlapped voltammetric signals obtained for the analyzed biogenic amines. Prediction errors lower than 9% have been obtained when concentration of quaternary mixtures were calculated. The developed procedure has been applied in the analysis of ham samples, which results are in good agreement with those obtained using the standard HPLC method.

A disposable electrodic system consisting of two working electrodes connected in array mode has been developed for the simultaneous determination of histamine (His) and putrescine (Put). Histamine deshydrogenase and putrescine oxidase enzymes were respectively immobilized by crosslinking on each working screen-printed electrode, both modified with tetrathiafulvalene. The dual system allowed the simultaneous amperometric determination of both species by measuring the oxidation current of the mediator in each working electrode. The effect of other potentially interfering biogenic amines was also evaluated. The capability of detection was of 8.1 ± 0.7 for His and 10 ± 0.6 μM for Put. The precision in terms of relative standard deviation was of 3.5% and 6.7% for His and Put, respectively. The developed biosensor was successfully applied to the determination of His and Put in different food samples.

An enzymatic amperometric procedure for measurement of chromium(III), based on the inhibitive action of this metal on tyrosinase enzyme activity, was developed for this purpose. A glassy carbon electrode was used as a support for the electropolymerisation of a polypyrrole (PPy) film, in which tyrosinase was immobilised. The parameters were optimised using experimental design methodology. To determine the optimum conditions and the influence of the factors in the experiment, a canonical analysis of the response surface was carried out. Under these conditions, the repeatability (R.S.D.

Pyrrole was electropolymerized onto a Pt electrode in the presence of LiClO(4) and horseradish peroxidase (HRP). This HRP-based biosensor has been used for the amperometric detection of rifampicin (RIF) in the presence of a constant concentration of H(2)O(2). The C(H(2)O(2)) as well as the applied potential (E(ap)) and the pH of the phosphate buffer have simultaneously been optimized through a central composite design. Under these conditions, repeatability, reproducibility, and stability of the modified electrode have been analyzed. The detection limit for RIF has been calculated taking into account the probability of false-positive (alpha) and -negative (beta), reaching a value of 5.06x10(-6) mol dm(-3). The biosensor was applied to the determination of RIF in pharmaceutical preparations and biological samples.

A novel amperometric biosensor for the determination of Al(III) based on the inhibition of the enzyme acetylcholinesterase has been developed. The immobilization of the enzyme was performed on screen-printed carbon electrodes modified with gold nanoparticles. The oxidation signal of acetylthiocholine iodide enzyme substrate was affected by the presence of Al(III) ions leading to a decrease in the amperometric current. The developed system has a detection limit of 2.1 ± 0.1 μM for Al(III). The reproducibility of the method is 8.1% (n = 4). Main interferences include Mo(VI), W(VI) and Hg(II) ions. The developed method was successfully applied to the determination of Al(III) in spiked tap water . The analysis of a certified standard reference material was also carried out. Both results agree with the certified values considering the respective associated uncertainties., Research Vicerrectory of Costa Rica University (Project 804-B0-197), Junta de Castilla y León (BU212A12-2), Ministerio de Ciencia e Innovación (MICINN) and Fondo Europeo de Desarrollo Regional (FEDER) (INNPACTO SERIBIO 2011–2014) and TEC-2009/12029

A simple and rapid method for the joint polarographic determination of rifampicin (RIF), isoniazid (INH) and pyrazinamide (PZA) by differential pulse polarography (DPP) is described. In this paper, a partial least square (PLS) regression was used for the resolution of the overlapped polarographic signals from mixtures of the three drugs. A genetic algorithm (GA) was used to select some of the predictor variables (potentials of the polarogram). The PLS model constructed with these selected potentials also leads to satisfactory results. The procedure was successfully applied to pharmaceutical preparations and biological fluids.

A chronoamperometric method for vanadium ion determination, based on the inhibition of the enzyme alkaline phosphatase, is reported. Screen-printed carbon electrodes modified with gold nanoparticles were used as transducers for the immobilization of the enzyme. The enzymatic activity over 4-nitrophenyl phosphate sodium salt is affected by vanadium ions, which results in a decrease in the chronoamperometric current registered. The developed method has a detection limit of 0.39 ± 0.06 µM, a repeatability of 7.7% (n = 4) and a reproducibility of 8% (n = 3). A study of the possible interferences shows that the presence of Mo(VI), Cr(III), Ca(II) and W(VI), may affect vanadium determination at concentration higher than 1.0 mM. The method was successfully applied to the determination of vanadium in spiked tap water, Research Vicerrectory of Costa Rica University (Project 804-B0-058) and Spanish Ministry of Science and Innovation (TEC-2009/12029). This work was supported by the Spanish Ministry of Science and Innovation (MICINN) 410 and the European Regional Development Fund (FEDER) (INNPACTO SERIBIO 2011-411 2014) and TEC2009-12029, as well as through Junta de Castilla y León (BU212A12-2).

The following paper describes the development of a screen-printed voltammetric pH-sensor based on graphite electrodes incorporating both internal indicator (i.e., phenanthraquinone) and reference species (i.e., dimethylferrocene). The key advantages of this type of system stem from its simplicity, low cost and ease of fabrication. More importantly, as opposed to conventional voltammetric systems where the height of the voltammetric peaks is taken into account to quantify the amount of a species of interest, here, the difference between the peak potential of the indicator species and the peak potential of the reference species is used. Thus, this measurement principle makes the electrochemical system presented here less dependent on the potential of the reference electrode (RE), as is often the case in other electrochemical systems. The developed system displays very promising performances, with a reproducible Super Nernstian response to pH changes and a lifetime of at least nine days.

Enzymatic amperometric procedures for measurement of Hg (II), based on the inhibitive action of this metal on urease enzyme activity, were developed. Screen-printed carbon electrodes (SPCEs) and gold nanoparticles modified screen-printed carbon electrodes (AuNPs/SPCEs) were used as supports for the cross-linking inmobilization of the enzyme urease. The amperometric response of urea was affected by the presence of Hg (II) ions which caused a decreasing in the current intensity. The optimum working conditions were found using experimental design methodology. Under these conditions, repeatability and reproducibility for both types of biosensors were determined, reaching values below 6% in terms of residual standard deviation. The detection limit obtained for Hg (II) was 4.2x10(-6)M for urease/SPCE biosensor and 5.6x10(-8)M for urease/AuNPs/SPCE biosensor. Analysis of the possible effect of the presence of foreign ions in the solution was performed. The method was applied to determine levels of Hg (II) in spiked human plasma samples.

An easy covalent immobilization method used to develop enzyme biosensors based on carbon and gold screen printed electrodes (SPCEs and gold SPEs) is described. The linkage of biomolecules through 4-nitrobenzenediazonium tetrafluoroborate, mercaptopropionic acid and thioctic acid monolayers has been attempted using bare SPCEs and gold SPEs, as well as gold nanoparticles (AuNPs) modified SPCEs and gold SPEs. Direct covalent attachment of Cytochrome P450 2B4 (CYP450 2B4) to the transducer has been carried out by carbodiimide and hydroxysuccinimide. Experimental variables in the immobilization process and in the chronoamperometric determination of Phenobarbital (PB) have been optimized by the experimental design methodology. Reproducibility of the different biosensors has been checked under the optimum conditions, yielding values lower than 6%. Their performances have been shown by the determination of PB in pharmaceutical drugs.

This paper describes a procedure that has been optimized for the determination of lamotrigine by Differential Pulse Adsorptive Stripping Voltammetry (DPAdSV) using carbon screen-printed electrodes (CSPE) and mercury coated carbon screen-printed electrodes. Selection of the experimental parameters was made using experimental design methodology. The detection limit found was 5.0 x 10-6 M and 2.0 x 10-6 M for the non modified and Hg modified CSPE, respectively. In terms of reproducibility, the precision of the above mentioned methods was calculated in %RSD values at 9.83% for CSPE and 2.73% for Hg-CSPE. The Hg-coated CSPEs developed in this work were successfully applied in the determination of lamotrigine in pharmaceutical preparations.

INTRODUCTION Anemia is the most frequent cytopenia in lower-risk MDS. Erythropoietic-stimulating agents (ESAs) are commonly used in these patients. The use of ÒclassicalÓ parameters (EPO and ferritin levels) and the revised IPSS (IPSS-R) has been proposed1 (SantiniÕs score) to predict response to ESAs and overall survival (OS) among patients with lower risk MDS by IPSS and a favorable Nordic group score2. OBJECTIVES The main objective of the study was to evaluate overall response rate (ORR) to ESAs and OS according to the proposed SantiniÕs score in an independent and large cohort of anemic lower risk MDS patients receiving treatment with ESAs. METHODS Data from 530 anemic patients with low/int1 risk IPSS de novo MDS (according to FAB and WHO criteria) and sufficient follow-up data available were recorded in Spresas3 (SPanish Registry of Erythropoietic Stimulating Agents Study from GESMD). Two hundred and twenty six patients (42.6% of the patients) were selected according to specific criteria regarding the published SantiniÕs score1: Hb level 200mU/mL(=1), serum ferritin (SF) >350 ng/mL(=1) and IPSS-R very low=0, low=1, intermediate=2 and high=3) yielded a score ranging from 0 to 5. ESAs response rate and overall survival were analysed according to these score. Response to treatment was evaluated according to IWG 2006 response criteria and a multivariate logistic regression analysis was used to identify independent predictors of erythroid response (ER). OS were defined as the time between diagnosis and the corresponding event or last follow up (Feb 2015) and were analyzed using univariable and multivariable Cox proportional hazards regression methods. RESULTS Median age was 77 years (interquartile range [IQR] 25%-75%: 71-83 y), median Hb level at start of treatment was 10 g/dL (IQR25-75: 9-10), median EPO level was 90 (IQR25-75: 27,25-108) and median ferritin level was 338,5 (IQR25-75: 146,5-568,75). Among 139 patients with this data available, 85 patients (61,1%) were RBC transfusion dependent before ESAs treatment. Median time from diagnosis to ESAs treatment was 82 (IQR25-75: 27-353) days. According to the IPSS, 68.6% (N=155) and 31.4% (N=71) were in low and Int-1 risk groups, respectively. Regarding IPSS-R, 23% (N=52), 66.8% (N=151), 9.7% (N=22) and 0.4% (N=1) were in very low, low, intermediate and high risk, respectively. ORR to ESA treatment was 71.2% (N=161), with a median duration of response of 2.06 years. Prognosis factors of ER showed a trend toward to a higher ER among patients in the lower IPSS-R (P>0.05), low IPSS (p=0.039) and lower EPO levels (p According to SantiniÕs score, 11.5%(N=26), 42.9%(N=97), 25.8%(N=81), 8%(N=18) and 1.8%(N=4) of the patients were in the 0, 1, 2 3 and 4 score. Erythroid response was better for patients in the lower scores, with response rates of 73.1%, 82.5%, 65.4%, 50% and 0%, for patients in 0, 1, 2, 3 and 4 score, respectively (p CONCLUSIONS The present study confirms that SantiniÕs score is useful to identify patients with a higher probability of response to ESAs and better OS among lower risk MDS patients with an expected favorable response to ESA according to Nordic group score. Spresas study was partly supported by Janssen 1.-Santini et al, Blood 122(13), 2013. 2.-Hellstršm-Lindberg, Br J Haematol 120(6), 2003. 3.-D'ez Campelo, EHA 2015 meeting, P244. Disclosures Díez Campelo: Novartis: Research Funding, Speakers Bureau; Janssen: Research Funding; Celgene: Research Funding, Speakers Bureau. Off Label Use: Use of erythropoietic stimulating agents for anemia in patients with myelodysplastic syndromes. Ramos:JANSSEN: Honoraria, Membership on an entity's Board of Directors or advisory committees; AMGEN: Consultancy, Honoraria; NOVARTIS: Consultancy, Honoraria; CELGENE: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Falantes:Celgene: Honoraria. Garcia:Celgene: Research Funding. Sanz:JANSSEN CILAG: Honoraria, Research Funding, Speakers Bureau.

Background: In higher-risk patients with myelodysplastic syndromes (MDS), the DNA methyltransferase inhibitors produce responses in 20% to 30% of patients and azacitidine (AZA) has demonstrated a survival advantage when comparedwith conventional therapies. Introduction: AZA is increasingly used in low-risk patients failing to respond to recombinant human erythropoietin (r-EPO) and in acute myeloid leukaemia patients (AML) with low marrow blast count or considered not fit for intensive chemotherapy. Purpose:We report a retrospective review on AZA treatment of MDS and AML patients in the common clinical practice of an Italian region (Liguria). Materials and Methods: Of 67 patients who started therapy only 46 patients received at least 4 courses of therapy, and were therefore considered evaluable for response and included in the study. Median age was 74 years (56-84), male/female ratio was 25/21. Nine patients (19%) had untreated AMLwithmarrow blasts ranging from 25 to 41%. MDS patients had RA or RARS (n.7, 19%), AREB-1 (n. 13, 35%), AREB 2 (n. 15, 40%), other forms (n. 2, 5%). In MDS patients the IPSS score was low/int-1 in 18 pts, int-2/high in 13 pts, not assessed in 7 pts. Twenty-six patients had a transfusion-dependent anaemia and the median number of packed erythrocyte units transfused weekly was 1 (range 1-2). All low and int-1 risk MDS patients had transfusion dependent anaemia and were unresponsive to r-EPO. Results: After a median of 8 courses (range 4-44) 26 patients (63%) achieved a haematological response (CR in 26%, PR in 24%, HI in 6,5%) whereas 20 (43,5%) were unresponsive. According to diagnosis and IPSS score responders were 6 (66,6%) among AML patients (CR 1, PR 4, HI 1), 10 (55,5%) in low/int-1 risk MDS patients and 7 (54%) among int-2/high risk MDS patients (CR 4, PR 3). Response was achieved after a median of 5 (range 3-6), 3 (range 2-12) and 5 (range 1-12) AZA courses in AML, low/int-1 risk and int-2/high risk MDS patients, respectively. Grade 1-2 myelotoxicity was commonly observed but no life threatening infections were reported. Eight-teen patients concomitantly received r-EPO therapy. Response lasted a median of 16 months (range 4-40) and median survival was 6 months in AML patients (4-9) and 23 months (6-48) in MDS patients. Conclusions: These preliminary data confirm efficacy and feasibility of AZA therapy in the common clinical practice, for both AML and all risk MDS patients.

The tumoral clone of Waldenström’s macroglobulinemia (WM) shows a wide morphological heterogeneity which ranges from B lymphocytes (BL) to plasma cells (PC), including a lymphoplasmacytic population that defines the disease. The differences between these cell compartments and their cell-counterpart in other lymphoproliferative disorders have not yet been sufficiently explored. We compared the gene expression profiling (GEP) of BL and PC from patients diagnosed with WM, with clonal BL and PC from patients with chronic lymphocytic leukemia (CLL) and multiple myeloma (MM) respectively. Bone marrow samples from 10 WM and 10 MM patients and peripheral blood from 10 CLL patients were used for the analysis. The isolation of the different cell populations was carried out by multiparameter flow cytometry sorting with the following monoclonal antibodies combination: Kappa or Lambda-FITC, CD10-PE, CD38-PerCP-Cy5.5, CD19-PE-Cy7, CD34-APC and CD45-APC-Cy7. Total RNA (100–500 ng) was amplified and labeled using the “GeneChip Two-Cycle cDNA Synthesis Kit” and hybridized to “Human Genome U133A” microarray (Affymetrix). Processing of genechip data was carried out using the Robust Multi-chip Average (RMA) and the Affymetrix Microarray Suite v.5 (MAS5) gene expression algorithms. Two-way hierarchical cluster analysis showed that GEP was able to classify PC from WM and PC from MM in two different groups. In a similar way, BL from WM and CLL were grouped in different clusters. The Significance Analysis of Microarrays (SAM) algorithm identified gene expression changes in a total of 163 genes (103 up and 60 down-regulated) when MW PC were compared with MM PC. Some of these genes were related to regulation of PC development: PAX5 was overexpressed and IRF4 infraexpressed in WM PC. PC from WM displayed high expression levels of MYB and DEK oncogenes while MM PC showed an elevated expression level of v-MAF oncogene. Regarding gene functional categories, “immune response” and “signal transduction” were the biological process more active in WM PC. Comparison between BL from WM and BL from CLL revealed that 31 genes were differentially expressed: IL4 receptor, LEF1 (WNT/b-catenin pathway), fibromodulin (modulator of TGF-b activity) and FGR oncogene (tyrosine kinase protein) showed very low expression levels in WM BL compared to CLL BL. In contrast, the growth factor IL6 was over-expressed in WM BL. These results indicate that both PC and BL from WM are genetically different from the MM and CLL cell-counterpart. The differentially expressed genes have important functions in the B-cell differentiation and oncogenesis. Supported by Spanish Myeloma Network (G03/136) and “Ministerio de Ciencia y Tecnología” (SAF04/06587) and “Junta de Castilla y León” grants (SA032/04)