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Electrochemical analysis of acetaminophen using a boron-doped diamond thin film electrode applied to flow injection system.

The electrochemistry of acetaminophen in phosphate buffer solution (pH 8) was studied at a boron-doped diamond (BDD) thin film electrode using cyclic voltammetry, hydrodynamic voltammetry, and flow injection with amperometric detection. Cyclic voltammetry was used to study the reaction as a function of concentration of analyte. Comparison experiments were performed using a polished glassy carbon (GC) electrode. Acetaminophen undergoes quasi-reversible reaction at both of these two electrodes. The BDD and GC electrodes provided well-resolved cyclic voltammograms but the voltammetric signal-to-background ratios obtained from the diamond electrode were higher than those obtained from the GC electrode. The diamond electrode provided a linear dynamic range from 0.1 to 8 mM and a detection of 10 microM (S/B approximately 3) for voltammetric measurement. The flow injection analysis results at the diamond electrode indicated a linear dynamic range from 0.5 to 50 microM and a detection limit of 10 nM (S/N approximately 4). Acetaminophen in syrup samples has also been investigated. The results obtained in the recovery study (24.68+/-0.26 mg/ml) were comparable to those labeled (24 mg/ml).

Acetaminophen↗

A psychophysical evaluation of spectral enhancement.

Listeners with sensorineural hearing loss have well-documented elevated hearing thresholds; reduced auditory dynamic ranges; and reduced spectral (or frequency) resolution that may reduce speech intelligibility, especially in the presence of competing sounds. Amplification and amplitude compression partially compensate for elevated thresholds and reduced dynamic ranges but do not remediate the loss in spectral resolution. Spectral-enhancement processing algorithms have been developed that putatively compensate for decreased spectral resolution by increasing the spectral contrast, or the peak-to-trough ratio, of the speech spectrum. Several implementations have been proposed, with mixed success. It is unclear whether the lack of strong success was due to specific implementation parameters or whether the concept of spectral enhancement is fundamentally flawed. The goal of this study was to resolve this ambiguity by testing the effects of spectral enhancement on detection and discrimination of simple, well-defined signals. To that end, groups of normal-hearing (NH) and hearing-impaired (HI) participants listened in 2 psychophysical experiments, including detection and frequency discrimination of narrowband noise signals in the presence of broadband noise. The NH and HI listeners showed an improved ability to detect and discriminate narrowband increments when there were spectral decrements (notches) surrounding the narrowband signals. Spectral enhancements restored increment detection thresholds to within the normal range when both energy and spectral-profile cues were available to listeners. When only spectral-profile cues were available for frequency discrimination tasks, performance improved for HI listeners, but not all HI listeners reached normal levels of discrimination. These results suggest that listeners are able to take advantage of the local improvement in signal-to-noise ratio provided by the spectral decrements.

Acoustic Stimulation↗

Patient performance with the Cochlear Corporation "20 + 2" implant: bipolar versus monopolar activation.

A within-subjects comparison of monopolar versus bipolar stimulation was performed using a modified version of the Nucleus mini-22 cochlear implant, the Nucleus "20 + 2" implant. Six subjects underwent implantation with this device, which is identical to the Nucleus 22 cochlear implant with the addition of two extracochlear indifferent electrodes. These electrodes provide two monopolar modes of stimulation in addition to the standard bipolar modes used with the Nucleus 22 device. One of the indifferent electrodes is a ball placed under the temporalis muscle (MP1), whereas the second electrode is mounted on the lateral aspect of the receiver-stimulator (MP2). After a pre-experimental phase, subjects used each of three stimulation modes (BP + 1, MP1, and MP2) for a total of 4 weeks each. Variables tested with each mode included electric thresholds, comfort levels, dynamic ranges, and speech recognition. Both the MP1 and MP2 modes of stimulation required significantly less current than the bipolar mode (BP + 1) to reach threshold and comfort level. Analysis of dynamic range data also indicated a significant stimulation mode effect. However, stimulation in the various modes did not significantly affect speech recognition scores, although two subjects demonstrated significantly improved speech recognition scores when programmed in a monopolar mode.

Adult↗

Quantitative proteome analysis in benign thyroid nodular disease using the fluorescent ruthenium II tris(bathophenanthroline disulfonate) stain.

Thyroid tumorigenesis involves qualitative and quantitative changes in protein expression, which can be comprehensively studied by proteome analysis. However, one of the technical bottlenecks of proteomics remains a reliable, sensitive and inexpensive method for quantification of differentially expressed proteins. This is due to the limited linear range of most available protein stains, i.e. silver and Coomassie blue, and high costs of commercially available fluorescent stains. In this paper we describe our experience with a lab-made ruthenium based fluorescent stain (ruthenium II tris(bathophenanthroline disulfonate) (RuBPs)) to perform proteome analysis of nodular thyroid disease. We first compared the properties of RuBPs with two highly sensitive protein stains: (1) silver staining and (2) the commercially available fluorescent dye Sypro Ruby. We show that in addition to its highly sensitive staining capabilities similar to Sypro Ruby and silver (2 ng), RuBPs offers several advantages such as a broad dynamic range (similar to Sypro Ruby and 500 times broader than the dynamic range of silver stain), low costs ( 0.03 per gel) and excellent compatibility with mass spectrometry. We then applied the inexpensive RuBPs stain to 2D gels (pH 4-7) of four benign thyroid nodules and normal thyroid tissue. We were able to detect approximately 1800 protein spots/gel in our thyroid samples. Quantitative changes in protein expression levels of at least 20-42 proteins were noted in the benign nodules compared with the normal thyroid tissue of the same patient. Differentially expressed spots were further characterised by nano-LC-FTICR and MALDI-TOF mass spectrometry. In summary we demonstrate, that the novel fluorescent ruthenium II tris(bathophenanthroline disulfonate) stain is a highly sensitive, reliable and inexpensive tool for quantitative proteome analysis in thyroid nodular disease.

Dextrans↗

Optimal pressure conditions for unbiased external ion accumulation in a two-dimensional radio-frequency quadrupole for Fourier transform ion cyclotron resonance mass spectrometry.

When combined with on-line separations (e.g., capillary liquid chromatography (LC)), Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) provides a powerful tool for biological applications, and particularly proteomic studies. The sensitivity, dynamic range, and duty cycle provided by FTICR-MS have been shown to be increased by ion trapping and accumulation in a two-dimensional (2D) radio-frequency (rf)-only multipole positioned externally to an FTICR cell. However, it is important that ions be detected across the desired m/z range without a significant bias. In this work we found that pressure inside the accumulation rf-quadrupole plays an important role in obtaining "unbiased" ion accumulation. Pressure optimization was performed in both pulsed and continuous modes. It was found that unbiased accumulation in a 2D rf-only quadrupole could be achieved in the pressure range of 5 x 10(-4) to 5 x 10(-3) Torr. External ion accumulation performed at the optimal pressure resulted in an increase in both the spectrum acquisition rates and dynamic range.

Algorithms↗

High-performance liquid chromatographic-fluorescence determination of traces of selenium in biological materials.

An improved method for the determination of selenium in biological materials has been developed. This work both extends and validates the procedure of Vézina and Bleau (J. Chromatog. 426, 385-391, 1988) which is based on high-performance liquid chromatographic determination of the fluorophore formed by reaction of Se(IV) with 2,3-diaminonaphthalene. The mass detection limit is 48 pg selenium (3 sigma) and the concentration detection limits are 48 parts per trillion in biological fluids and 120 to 480 parts per trillion in dried biological materials. The linear dynamic range of the method has been extended up to approximately 800 ng. Relative standard deviations of 9.4 to 2.7% were observed in repeated analyses of standards in the range of 0.5 to 500 ng. The proposed method was validated with respect to 23 biological reference materials spanning an 1800-fold range of selenium concentrations and was found to be free of significant constant or proportional biases despite greatly different matrix compositions. This method offers an unsurpassed combination of sensitivity, accuracy, linear dynamic range, and freedom from matrix interferences and may be considered a reference method for the reliable determination of selenium in biological materials.

Chromatography, High Pressure Liquid↗

Highly sensitive enzyme-linked immunosorbent assay for chlorpyrifos. Application to olive oil analysis.

Highly sensitive enzyme-linked immunoassays for chlorpyrifos, one of the most applied insecticides, are presented. Several haptens were synthesized for immunoreagent production and ELISA development. The best immunoassays obtained are based on an indirect coated-plate immunoassay format. Two assays were optimized; one shows a limit of detection of 0.3 ng L(-1), an I50 of 271 ng L(-1), and a dynamic range between 4 and 16 474 ng L(-1). The other one has a limit of detection of 0.07 ng L(-1), an I50 of 7 ng L(-1), and a dynamic range between 0.4 and 302 ng L(-1). The assays were used to quantify chlorpyrifos in olive oil using a simple and rapid sample extraction procedure. The good recoveries achieved in both assays (109% mean value) and the agreement with values given by the GC reference method (110% mean value) indicate their potential for either screening or laboratory quantification.

Chlorpyrifos↗

Intensity perception XI. Experimental results on the relation of intensity resolution to loudness matching.

According to a recent extension of our theory of intensity perception [Lim et al., J. Acoust. Soc. Am 62, 1256-1267 (1977)], two stimuli are matched in loudness if and only if their intensities divide the respective dynamic ranges proportionally in terms of just noticeable differences. This study reports results of intensity discrimination and loudness matching experiments designed to test this prediction. Data were obtained over most of the dynamic range for three different types of sounds: a 1000-Hz tone in quiet, a 1000-Hz tone partially masked by a 2-octave band of noise, and spectrally flat wide-band noise. Of the five subjects tested, three produced results that had sufficient internal consistency to be useful for testing the predictions. For these subjects, the data and the theory were found to be reasonably consistent. Comparison with other studies, however, reveals that loudness matching results vary with matching paradigm by an amount that is significant with respect to the task of evaluating the theory. Hence, a rigorous test of the theory requires an improved understanding of the effects of matching paradigm.

Humans↗

Place-pitch sensitivity and its relation to consonant recognition by cochlear implant listeners using the MPEAK and SPEAK speech processing strategies.

Two related studies investigated the relationship between place-pitch sensitivity and consonant recognition in cochlear implant listeners using the Nucleus MPEAK and SPEAK speech processing strategies. Average place-pitch sensitivity across the electrode array was evaluated as a function of electrode separation, using a psychophysical electrode pitch-ranking task. Consonant recognition was assessed by analyzing error matrices obtained with a standard consonant confusion procedure to obtain relative transmitted information (RTI) measures for three features: stimulus (RTI stim), envelope (RTI env[plc]), and place-of-articulation (RTI plc[env]). The first experiment evaluated consonant recognition performance with MPEAK and SPEAK in the same subjects. Subjects were experienced users of the MPEAK strategy who used the SPEAK strategy on a daily basis for one month and were tested with both processors. It was hypothesized that subjects with good place-pitch sensitivity would demonstrate better consonant place-cue perception with SPEAK than with MPEAK, by virtue of their ability to make use of SPEAK's enhanced representation of spectral speech cues. Surprisingly, all but one subject demonstrated poor consonant place-cue performance with both MPEAK and SPEAK even though most subjects demonstrated good or excellent place-pitch sensitivity. Consistent with this, no systematic relationship between place-pitch sensitivity and consonant place-cue performance was observed. Subjects' poor place-cue perception with SPEAK was subsequently attributed to the relatively short period of experience that they were given with the SPEAK strategy. The second study reexamined the relationship between place-pitch sensitivity and consonant recognition in a group of experienced SPEAK users. For these subjects, a positive relationship was observed between place-pitch sensitivity and consonant place-cue performance, supporting the hypothesis that good place-pitch sensitivity facilitates subjects' use of spectral cues to consonant identity. A strong, linear relationship was also observed between measures of envelope- and place-cue extraction, with place-cue performance increasing as a constant proportion (approximately 0.8) of envelope-cue performance. To the extent that the envelope-cue measure reflects subjects' abilities to resolve amplitude fluctuations in the speech envelope, this finding suggests that both envelope- and place-cue perception depend strongly on subjects' envelope-processing abilities. Related to this, the data suggest that good place-cue perception depends both on envelope-processing abilities and place-pitch sensitivity, and that either factor may limit place-cue perception in a given cochlear implant listener. Data from both experiments indicate that subjects with small electric dynamic ranges (< 8 dB for 125-Hz, 205-microsecond/ph pulse trains) are more likely to demonstrate poor electrode pitch-ranking skills and poor consonant recognition performance than subjects with larger electric dynamic ranges.

Adult↗

Making blind robots see: the synergy between fluorescent dyes and imaging devices in automated proteomics.

Proteomics investigations endeavor to provide a global understanding of gene product synthesis rate, degradation rate, functional competence, posttranslational modification, subcellular distribution and physical interactions with other cell components. Protein expression encompasses an enormous dynamic range. Since rare proteins cannot be amplified by any type of PCR method, sensitive detection is critical to proteome projects. Fluorescence methods deliver streamlined detection protocols, superior detection sensitivity, broad linear dynamic range and excellent compatibility with modern microchemical identification methods such as mass spectrometry. Two general approaches to fluorescence detection of proteins are currently practiced: the covalent derivatization of proteins with fluorophores or noncovalent interaction of fluorophores either via the SDS micelle or through direct electrostatic interaction with proteins. One approach for quantifying fluorescence is to use a photomultiplier tube detector combined with a laser light scanner. In addition, fluorescence imaging is performed using a charge-coupled device camera combined with a UV light or xenon arc source. Fluorescent dyes with bimodal excitation spectra may be broadly implemented on a wide range of analytical imaging devices, permitting their widespread application to proteomics studies and incorporation into semiautomated analysis environments.

Animals↗

The effect of bicuculline application on auditory response properties of inferior collicular neurons of mice with or without monaural middle ear destruction in early age.

Previous studies have demonstrated that abnormal auditory stimulation during early postnatal development can be manifested through physiological changes that occur in the inferior colliculus (IC) of mammals. To determine the contribution of the GABAergic transmitter systems to the development of response properties of IC neurons, we examined the effect of application of bicuculline (which is an antagonist for the GABA(A) receptors) on response properties of IC neurons of the laboratory mice, Mus musculus, with or without early monaural middle ear destruction. Monaural middle ear destruction was performed at 12-14 days after birth. At adulthood, the auditory response properties of IC neurons were examined in both experimental conditions. All IC neurons determined before and during bicuculline application can be described as (1) phasic responders which discharged 1-2 impulses; (2) phasic bursters which discharged 3-7 impulse; and (3) tonic responders which discharged impulses throughout the duration of presented sound pulses. Early monaural middle ear destruction only affected the percent distribution but not the type of discharge pattern, rate-intensity function and frequency tuning curve of IC neurons in the control and experimental mice. Neurons in the contralateral IC of experimental mice typically had longer latencies, higher minimum thresholds, broader frequency tuning curves and smaller dynamic ranges than neurons in the ipsilateral IC and in control mice. Bicuculline application produced differential effects in decreasing the latencies and minimum thresholds as well as broadening frequency tuning curves and dynamic ranges of IC neurons in these two groups of mice. All these data suggest that early monaural middle ear destruction did not affect the shaping of auditory response properties of IC neurons by GABAergic transmitter system.

Age Factors↗

Fiber optic-based biosensor for ricin.

We report on an evanescent wave fiber-optic biosensor for detecting a potently toxic protein, ricin, in the picograms per milliliter range. A sandwich immunoassay scheme was used to detect ricin. First, an anti-ricin IgG was immobilized onto the surface of an optical fiber in two different ways. In the first method, the antibody was directly coated to the silanized fiber using a crosslinker. Second, avidin-coated fibers were incubated with biotinylated anti-ricin IgG to immobilize the antibody using an avidin-biotin bridge. The assay using the avidin-biotin linked antibody demonstrated higher sensitivity and wider linear dynamic range than the assay using antibody directly conjugated to the surface. The linear dynamic range of detection for ricin in buffer using the avidin-biotin chemistry is 100 pg/ml-250 ng/ml. The limits of detection for ricin in buffer solution and river water are 100 pg/ml and 1 ng/ml, respectively. At higher concentrations of ricin (> 50 ng/ml), we observe a strong interaction of ricin with the avidin coated on the surface of the fibers. We have demonstrated that this interaction is primarily due to the lectin activity of ricin and is significantly reduced using fibers coated with neutravidin or by adding galactose to the ricin samples.

Adsorption↗

Determination of phenylalanine isotope ratio enrichment by liquid chromatography/time- of-flight mass spectrometry.

The application of time-of-flight mass spectrometry to isotope ratio measurements has been limited by the relatively low dynamic range of the time-to-digital converter detectors available on commercial LC/ToF-MS systems. Here we report the measurement of phenylalanine isotope ratio enrichment by using a new LC/ToF-MS system with wide dynamic range. Underivatized phenylalanine was injected onto a C18 column directly with 0.1% formic acid/acetonitrile as the mobile phase. The optimal instrument parameters for the time-of-flight mass spectrometer were determined by tuning the instrument with a phenylalanine standard. The accuracy of the isotope enrichment measurement was determined by the injection of standard solutions with known isotope ratios ranging from 0.02% to 9.2%. A plot of the results against the theoretical values gave a linear curve with R2 of 0.9999. The coefficient of variation for the isotope ratio measurement was below 2%. The method is simple, rapid, and accurate and presents an attractive alternative to traditional GC/MS applications.

Animals↗

Real-time reverse transcription-PCR assay for future management of ERBB2-based clinical applications.

BACKGROUND: Gene amplification/overexpression of ERBB2 (HER2, neu) is a major event in human breast tumorigenesis. ERBB2-based therapeutic agents and ERBB2-specific gene therapy are under development. These new perspectives call for a sensitive and accurate method to screen breast cancer patients for ERBB2 alterations. METHODS: We have developed and validated a real-time quantitative reverse transcription (RT)-PCR assay, based on fluorescent TaqMan methodology, to quantify ERBB2 gene expression at the mRNA level in breast tumors. This recently developed method of nucleic acid quantification in homogeneous solutions has the potential for a wide dynamic range, interlaboratory agreement, and high-throughput capacity without tedious post-PCR processing. The ERBB2 mRNA signal was normalized to the signal for TATA box-binding protein mRNA. RESULTS: The dynamic range was >1000-fold. The relationship between C(t) and log starting concentration was linear (r(2) >/=0.99). The mean (SD) normalized expression of ERBB2 in healthy breast tissue was 0.95 (0.37). Overexpression (>5 SD above mean for healthy breast) of the ERBB2 gene was observed (at 3.2- to 135-fold) in 23 (17%) of 134 breast tumor RNA samples. As expected, ERBB2 overexpression was present in all tumors with ERBB2 gene amplification but was uncommon and at a low ratio (<5) in breast cancers without gene amplification. CONCLUSIONS: This new simple, rapid, semi-automated assay is a major alternative to fluorescence in situ hybridization and immunochemistry for gene alteration analysis in human tumors and may be a powerful tool for large randomized, prospective cooperative group trials and to support future ERBB2-based biological and gene therapy approaches.

Adult↗

Multichannel syllabic compression for severely impaired listeners.

Two listeners with congenital hearing losses characterized by flat audiograms and dynamic ranges of 18-33 dB were tested with three compression systems and one (reference) linear amplification system. The compression systems placed progressively larger amounts of speech energy within the listener's residual dynamic range, by raising to audibility and compressing 25, 50, and 90 percent of the short-term input amplitude distribution in each of 16 frequency bands. The comparison linear system was defined by adjusting six octave-wide bands of speech to comfortable levels. System performance was evaluated with nonsence CVC syllables presented at a constant input level and spoken by two talkers. Extensive training was provided to ensure stable performance. The results were notably speaker-dependent, with compression consistently providing better performance for one speaker, linear amplification for the other. Averaged over speakers, however, there was no net advantage for any of the compression systems for any listener. The use of high compression ratios and large input ranges tended to degrade perception of initial consonants and vowels. Under some conditions, however, final consonant scores were higher with compression than with linear amplification. Compression generally enhanced the distinction between stops and fricatives, but degraded spectral-concentration and relative-intensity cues required to identify place of articulation.

Adult↗

A microcapillary trap cartridge-microcapillary high-performance liquid chromatography electrospray ionization emitter device capable of peptide tandem mass spectrometry at the attomole level on an ion trap mass spectrometer with automated routine operation.

Reversed-phase microcapillary chromatography (RP-microLC) combined with electrospray ionization tandem mass spectrometry (ESI-MS/MS) is one of two prevailing techniques in proteomic analysis, the other being matrix-assisted laser desorption/ionization (MALDI). Despite the arguably better dynamic range obtainable with ESI, MALDI is increasingly popular due to ease of use, ruggedness and the ability to decouple separation from ionization. By contrast, in order to take advantage of the sensitivity and dynamic range afforded by the concentration-dependent nature of ESI, it is directly coupled to separations that take place in small i.d. RP-microLC columns. This gain in sensitivity often comes at a loss of ruggedness due to clogging of the small i.d. RP-microLC columns, one result of which is limited sample throughput. Here we describe a combined micropre-column-microLC-ESI device that is sensitive, rugged and modular in design allowing facile construction and troubleshooting. Due to low signal-to-noise as little as 1 attomole of a peptide can be selected by data-dependent methods for collision-induced dissociation. Importantly, the resulting tandem mass spectrum is of high enough quality to identify the peptide sequence by a database search against a complex database using SEQUEST. Finally, the device is demonstrated to be rugged as judged by >60 consecutive reversed-phase microLC separations on complex peptide mixtures before chromatographic resolution is degraded.

Angiotensin I↗

Coding of odour intensity in a sensory neuron.

A deterministic biophysical model of an olfactory sensory neuron under constant stimulation is presented with the aim of describing the successive conversion steps, including receptor activation, conductance change, receptor potential and firing frequency, that are involved in the coding of odorant concentration. This model is divided in two parts. The odorant-sensitive part (OSP), consisting of one cylindrical dendrite, is connected to the odorant-insensitive part (OIP), corresponding to passive dendrite, soma and axon. Each part exerts a specific effect on the coding properties of the conversion steps, i.e. their magnitude, sensitivity and dynamic range. The maximum conductance of the OSP affects positively all coding properties whereas the input resistance of the OIP, which depends on its size and shape, affects positively the sensitivity and negatively the dynamic range. These findings are helpful for understanding the input-output properties of many types of neurons.

Action Potentials↗

Two types of electrochemical nitric oxide (NO) sensing systems with heat-denatured Cyt C and radical scavenger PTIO.

To develop an in situ NO sensing system for primarily biological and medical uses, two types of NO sensing materials, which may be coupled with an electrochemical reaction for signal transduction, have been investigated. Heat-denatured Cyt C and a radical scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetra methyl imidazoline-1-oxyl-3-oxide (C-PTIO) were found to be effective and were incorporated into electrochemical sensing systems. Heat-denatured Cyt C deposited on a 4-mercaptopyridine modified gold electrode responded to NO with an increase of cathodic current through electrochemical reduction of Cyt C (Fe3+), when the electrode potential was controlled at 0 mV vs Ag/AgCl. The dynamic range of the sensing system was 0.5-4 microM. The sensing system with C-PTIO exhibited an anodic output in response to NO at 0.7 V vs Ag/AgCl, and showed a wider dynamic range from 0.05 to 100 microM.

Animals↗