Comparison of an ELISA and an LC/MS/MS method for measuring tacrolimus concentrations and making dosage decisions in transplant recipients

This study compared an enzyme-linked immunosorbent assay (ELISA) to a liquid chromatography-tandem mass spectrometry (LC/MS/MS) technique for measurement of tacrolimus concentrations in adult kidney and liver transplant recipients, and investigated how assay choice influenced pharmacokinetic parameter estimates and drug dosage decisions. Tacrolimus concentrations measured by both ELISA and LC/MS/MS from 29 kidney (n = 98 samples) and 27 liver (n = 97 samples) transplant recipients were used to evaluate the performance of these methods in the clinical setting. Tacrolimus concentrations measured by the two techniques were compared via regression analysis. Population pharmacokinetic models were developed independently using ELISA and LC/MS/MS data from 76 kidney recipients. Derived kinetic parameters were used to formulate typical dosing regimens for concentration targeting. Dosage recommendations for the two assays were compared. The relation between LC/MS/MS and ELISA measurements was best described by the regression equation ELISA = 1.02 . (LC/MS/MS) + 0.14 in kidney recipients, and ELISA = 1.12 . (LC/MS/MS) - 0.87 in liver recipients. ELISA displayed less accuracy than LC/MS/MS at lower tacrolimus concentrations. Population pharmacokinetic models based on ELISA and LC/MS/MS data were similar with residual random errors of 4.1 ng/mL and 3.7 ng/mL, respectively. Assay choice gave rise to dosage prediction differences ranging from 0% to 30%. ELISA measurements of tacrolimus are not automatically interchangeable with LC/MS/MS values. Assay differences were greatest in adult liver recipients, probably reflecting periods of liver dysfunction and impaired biliary secretion of metabolites. While the majority of data collected in this study suggested assay differences in adult kidney recipients were minimal, findings of ELISA dosage underpredictions of up to 25% in the long term must be investigated further.

Influence of planting date, method of handling and seed size on supersweet sweet corn seed quality

A supersweet sweet corn hybrid, Pacific H5, was planted at weekly intervals (P-1 to P-5) in spring in South-Eastern Queensland. All plantings were harvested at the same time resulting in immature seed for the last planting (P-5). The seed was handled by three methods: manual harvest and processing (M-1), manual harvest and mechanical processing (M-2) and mechanical harvest and processing (M-3), and later graded into three sizes (small, medium and large). After eight months storage at 12-14degreesC, seed was maintained at 30degreesC with bimonthly monitoring of germination for fourteen months and seed damage at the end of this period. Seed quality was greatest for M-1 and was reduced by mechanical processing but not by mechanical harvesting. Large and medium seed had higher germination due to greater storage reserves but also more seed damage during mechanical processing. Immature seed from premature harvest (P-5) had poor quality especially when processed mechanically and reinforced the need for harvested seed to be physiologically mature.

Static and hydrodynamic monitoring of citalopram based on its electro-oxidation behavior at a glassy-carbon surface

The electrooxidative behavior of citalopram (CTL) in aqueous media was studied by cyclic voltammetry (CV) and square-wave voltammetry (SWV) at a glassy-carbon electrode. The electrochemical behaviour of CTL involves two electrons and two protons in the irreversible and diffusion controlled oxidation of the tertiary amine group. The maximum analytical signal was obtained in a phosphate buffer (pH ¼ 8.2). For analytical purposes, an SWV method and a flow-injection analysis (FIA) system with amperometric detection were developed. The optimised SWV method showed a linear range between 1.10 10 5–1.20 10 4 molL 1, with a limit of detection (LOD) of 9.5 10 6 molL 1. Using the FIA method, a linear range between 2.00 10 6–9.00 10 5 molL 1 and an LODof 1.9 10 6 molL 1 were obtained. The validation of both methods revealed good performance characteristics confirming applicability for the quantification of CTL in several pharmaceutical products.

Spatio-temporal land use/land cover changes analysis and monitoring in the Valencia Municipality, Spain

Dissertation submitted in partial fulfilment of the requirements for the Degree of Master of Science in Geospatial Technologies

Development of a high-throughput monitoring technique of bacteria photodynamic inactivation

Summary form only given. Bacterial infections and the fight against them have been one of the major concerns of mankind since the dawn of time. During the `golden years' of antibiotic discovery, during the 1940-90s, it was thought that the war against infectious diseases had been won. However currently, due to the drug resistance increase, associated with the inefficiency of discovering new antibiotic classes, infectious diseases are again a major public health concern. A potential alternative to antibiotic treatments may be the antimicrobial photodynamic inactivation (PDI) therapy. To date no indication of antimicrobial PDI resistance development has been reported. However the PDI protocol depends on the bacteria species [1], and in some cases on the bacteria strains, for instance Staphylococcus aureus [2]. Therefore the development of PDI monitoring techniques for diverse bacteria strains is critical in pursuing further understanding of such promising alternative therapy. The present works aims to evaluate Fourier-Transformed-Infra-Red (FT-IR) spectroscopy to monitor the PDI of two model bacteria, a gram-negative (Escherichia coli) and a gram-positive (S. aureus) bacteria. For that a high-throughput FTIR spectroscopic method was implemented as generally described in Scholz et al. [3], using short incubation periods and microliter quantities of the incubation mixture containing the bacteria and the PDI-drug model the known bactericidal tetracationic porphyrin 5,10,15,20-tetrakis (4-N, N, Ntrimethylammoniumphenyl)-porphyrin p-tosylate (TTAP4+). In both bacteria models it was possible to detect, by FTIR-spectroscopy, the drugs effect on the cellular composition either directly on the spectra or on score plots of principal component analysis. Furthermore the technique enabled to infer the effect of PDI on the major cellular biomolecules and metabolic status, for example the turn-over metabolism. In summary bacteria PDI was monitored in an economic, rapid (in minutes- , high-throughput (using microplates with 96 wells) and highly sensitive mode resourcing to FTIR spectroscopy, which could serve has a technological basis for the evaluation of antimicrobial PDI therapies efficiency.

Distributed Forest Fire Monitoring Using Wireless Sensor Networks

Disaster management is one of the most relevant application fields of wireless sensor networks. In this application, the role of the sensor network usually consists of obtaining a representation or a model of a physical phenomenon spreading through the affected area. In this work we focus on forest firefighting operations, proposing three fully distributed ways for approximating the actual shape of the fire. In the simplest approach, a circular burnt area is assumed around each node that has detected the fire and the union of these circles gives the overall fire’s shape. However, as this approach makes an intensive use of the wireless sensor network resources, we have proposed to incorporate two in-network aggregation techniques, which do not require considering the complete set of fire detections. The first technique models the fire by means of a complex shape composed of multiple convex hulls representing different burning areas, while the second technique uses a set of arbitrary polygons. Performance evaluation of realistic fire models on computer simulations reveals that the method based on arbitrary polygons obtains an improvement of 20% in terms of accuracy of the fire shape approximation, reducing the overhead in-network resources to 10% in the best case.

Rapid automated method for on-site determination of sulfadiazine in fish farming: a stainless steel veterinary syringe coated with a selective membrane of PVC serving as a potentiometric detector in a flow-injection-analysis system

Sulfadiazine is an antibiotic of the sulfonamide group and is used as a veterinary drug in fish farming. Monitoring it in the tanks is fundamental to control the applied doses and avoid environmental dissemination. Pursuing this goal, we included a novel potentiometric design in a flow-injection assembly. The electrode body was a stainless steel needle veterinary syringe of 0.8-mm inner diameter. A selective membrane of PVC acted as a sensory surface. Its composition, the length of the electrode, and other flow variables were optimized. The best performance was obtained for sensors of 1.5-cm length and a membrane composition of 33% PVC, 66% onitrophenyloctyl ether, 1% ion exchanger, and a small amount of a cationic additive. It exhibited Nernstian slopes of 61.0 mV decade-1 down to 1.0×10-5 mol L-1, with a limit of detection of 3.1×10-6 mol L-1 in flowing media. All necessary pH/ionic strength adjustments were performed online by merging the sample plug with a buffer carrier of 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid, pH 4.9. The sensor exhibited the advantages of a fast response time (less than 15 s), long operational lifetime (60 days), and good selectivity for chloride, nitrite, acetate, tartrate, citrate, and ascorbate. The flow setup was successfully applied to the analysis of aquaculture waters. The analytical results were validated against those obtained with liquid chromatography–tandem mass spectrometry procedures. The sampling rate was about 84 samples per hour and recoveries ranged from 95.9 to 106.9%.

A disposable glass-based immunosensor for monitoring the cancer biomarker CEA in urine

NanoPT 2014 International Conference, in Portugal, on February 12-14. Poster presentation based on topic Nanobio/Nanomedicine

Electrochemical sensor for simultaneous determination of herbicide MCPA and its metabolite 4-chloro-2-methylphenol. Application to photodegradation environmental monitoring

The development and application of a polyaniline/carbon nanotube (CNT) cyclodextrin matrix (PANI-β-CD/MWCNT)-based electrochemical sensor for the quantitative determination of the herbicide 4-chloro-2-methylphenoxyacetic acid (MCPA) and its main transformation product 4-chloro-2-methylphenol in natural waters are described. A simple cyclic voltammetry-based electrochemical methodology, in phosphate buffer solution at pH 6.0, was used to develop a method to determine both MCPA and 4-chloro-2-methylphenol, without any previous extraction or derivatization steps. A linear concentration range (10 to 50 μmol L−1) and detection limits of 1.1 and 1.9 μmol L−1, respectively, were achieved using optimized cyclic voltammetric parameters. The proposed method was successfully applied to the determination of MCPA and 4-chloro-2-methylphenol in natural water samples with satisfactory recoveries (94 to 107 %) and in good agreement with the results obtained by an established high-performance liquid chromatography technique, no significant differences being found between the methods. Interferences from ionic species and other herbicides used for broad-leaf weed control were shown to be small. The newly developed methodology was also successfully applied to MCPA photodegradation environmental studies.

ezGo: A voice operated wheelchair with biosignal monitoring for home environments

In this paper we present ezGo, an electric powered wheelchair with a speech based interface and biosignal monitoring instrumentation. The user can use the voice, a natural communication method, for controlling the chair movement and obtain information about his health. Additionally a set of semi-autonomous modes with macro recording enable the execution of navigation tasks with little effort and improved precision. The main purpose of the system is to provide severely disabled persons with an assistive device that can improve their confidence and daily independence. The obtained results on usability tests showed that users consider ezGo a valuable help on their daily tasks and a very desirable addition to standard wheelchairs.

Monitoring Abacavir Bioactivation in Humans: Screening for an Aldehyde Metabolite

The anti-HIV drug abacavir is associated with idiosyncratic hypersensitivity reactions and cardiotoxicity. Although the mechanism underlying abacavir-toxicity is not fully understood, drug bioactivation to reactive metabolites may be involved. This work was aimed at identifying abacavir-protein adducts in the hemoglobin of HIV patients as biomarkers of abacavir bioactivation and protein modification. The protocol received prior approval from the Hospital Ethics Committee, patients gave their written informed consent and adherence was controlled through a questionnaire. Abacavir-derived Edman adducts with the N-terminal valine of hemoglobin were analyzed by an established liquid chromatography-electrospray ionization-tandem mass spectrometry method. Abacavir-valine adducts were detected in three out of ten patients. This work represents the first evidence of abacavir-protein adduct formation in humans. The data confirm the ability of abacavir to modify self-proteins and suggest that the molecular mechanism(s) of some abacavir-induced adverse reactions may require bioactivation.

Monitoring antibiotics in the environment. Study of Quinoxaline derivatives bioactivity

Dissertação para obtenção do Grau de Doutor em Química Sustentável

In-house ELISA method to analyze anti-Trypanosoma cruzi IgG reactivity for differential diagnosis and evaluation of Chagas disease morbidity

INTRODUCTION: The goal was to develop an in-house serological method with high specificity and sensitivity for diagnosis and monitoring of Chagas disease morbidity. METHODS: With this purpose, the reactivities of anti-T. cruzi IgG and subclasses were tested in successive serum dilutions of patients from Berilo municipality, Jequitinhonha Valley, Minas Gerais, Brazil. The performance of the in-house ELISA was also evaluated in samples from other relevant infectious diseases, including HIV, hepatitis C (HCV), syphilis (SYP), visceral leishmaniasis (VL), and American tegumentary leishmaniasis (ATL), and noninfected controls (NI). Further analysis was performed to evaluate the applicability of this in-house methodology for monitoring Chagas disease morbidity into three groups of patients: indeterminate (IND), cardiac (CARD), and digestive/mixed (DIG/Mix), based on their clinical status. RESULTS: The analysis of total IgG reactivity at serum dilution 1:40 was an excellent approach to Chagas disease diagnosis (100% sensitivity and specificity). The analysis of IgG subclasses showed cross-reactivity, mainly with NI, VL, and ATL, at all selected serum dilutions. Based on the data analysis, the IND group displayed higher IgG3 levels and the DIG/Mix group presented higher levels of total IgG as compared with the IND and CARD groups. CONCLUSIONS: These findings demonstrated that methodology presents promising applicability in the analysis of anti-T. cruzi IgG reactivity for the differential diagnosis and evaluation of Chagas disease morbidity.

Monitoring Trypanosoma cruzi infection in triatomines using PCR in Mato Grosso do Sul, Brazil

Introduction The aim of the present study was to assess the polymerase chain reaction (PCR) as a method for detecting Trypanosoma cruzi infection in triatomines that had been previously determined by microscopic examination in the State of Mato Grosso do Sul, Brazil. Methods In total, 515 specimens were collected. Material from the digestive tract of each triatomine was analyzed for the presence of T. cruzi by microscopic examination and PCR using the 121/122 primer set. Results Among the 515 specimens tested, 58 (11.3%) were positive by microscopy and 101 (19.61%) were positive by PCR and there was an association between the results of the techniques (χ2 = 53.354, p = 0.001). The main species of triatomine identified was T. sordida (95.5%) Conclusions The use of PCR in entomological surveillance may contribute to a better assessment of the occurrence of T. cruzi in triatomine populations.

Monitoring the treatment of sepsis with vancomycin in term newborn infants

A prospective study was conducted to determine if standardized vancomycin doses could produce adequate serum concentrations in 25 term newborn infants with sepsis. Purpose: The therapeutic response of neonatal sepsis by Staphylococcus sp. treated with vancomycin was evaluated through serum concentrations of vancomycin, serum bactericidal titers (SBT), and minimum inhibitory concentration (MIC). METHOD: Vancomycin serum concentrations were determined by the fluorescence polarization immunoassay technique , SBT by the macro-broth dilution method, and MIC by diffusion test in agar . RESULTS: Thirteen newborn infants (59.1%) had adequate peak vancomycin serum concentrations (20--40 mg/mL) and one had peak concentration with potential ototoxicity risk (>40 µg/mL). Only 48% had adequate trough concentrations (5--10 mg/mL), and seven (28%) had a potential nephrotoxicity risk (>10 µg/mL). There was no significant agreement regarding normality for peak and trough vancomycin method (McNemar test : p = 0.7905). Peak serum vancomycin concentrations were compared with the clinical evaluation (good or bad clinical evolution) of the infants, with no significant difference found (U=51.5; p=0.1947). There was also no significant difference between the patients' trough concentrations and good or bad clinical evolution (U = 77.0; p=0.1710). All Staphylococcus isolates were sensitive to vancomycin according to the MIC. Half of the patients with adequate trough SBT (1/8), also had adequate trough vancomycin concentrations and satisfactory clinical evolution. CONCLUSIONS: Recommended vancomycin schedules for term newborn infants with neonatal sepsis should be based on the weight and postconceptual age only to start antimicrobial therapy. There is no ideal pattern of vancomycin dosing; vancomycin dosages must be individualized. SBT interpretation should be made in conjunction with the patient's clinical presentation and vancomycin serum concentrations. Those laboratory and clinical data favor elucidation of the probable cause of patient's bad evolution, which would facilitate drug adjustment and reduce the risk of toxicity or failing to achieve therapeutic doses.