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PubMed2005

Extended-range glucose biosensor via layer-by-layer assembly incorporating gold nanoparticles.

We report on a glucose oxidase (GOx)/polyelectrolyte (PE)/gold nanoparticle (AuNP) multilayer films that can be utilized as efficient glucose biosensors by layer-by-layer self-assembly method. Electrochemical impedance spectroscopy (EIS) and UV-visible spectroscopy were adopted to monitor the regular growth of the multilayer films. The role of gold nanoparticles integrated in the multilayer films not only increase the amount and activity of GOx, but also significantly improve the electron-transfer characteristics of the films. The performance of the multilayer films for sensing glucose could be tailored by controlling the gold nanoparticles loading in the film and the number of layers. A biosensor constructed by four poly(dimethyldiallylammonium chloride) (PDDA)/AuNP/PDDA/GOx multilayer films exhibited a wide linear calibration range from 0 to 60.0 mM with the detection limit of 3.0 microM for the detection of glucose. The dynamic range can be extended up to 120 mM. The biosensor has good stability and reproducibility.

Absorption↗
PubMed1995

New noncompetitive immunoassays of small analytes.

We developed a novel noncompetitive immunoassay format for monoepitopic analytes and describe here a model assay for triiodothyronine (T3), performed on Ciba Corning's ACS:180 analyzer. Acridinium ester (AE)-labeled bivalent anti-T3 was incubated with the sample, producing AE-anti-T3/T3 complexes and unreacted AE-anti-T3. Controlled-pore glass particles (CPG) with immobilized diiodothyronine (T2) were then added in excess, to bind AE-anti-T3 possessing two unoccupied binding sites but not AE-anti-T3 bound to one or two T3 molecules. Paramagnetic particles (PMP) with immobilized anti-AE were then added to the same cuvette to capture AE-anti-T3/T3 complexes; AE-anti-T3 bound to the surface of CPG, however, was not captured, because of steric hindrance. After the incubation, the PMP was magnetically separated to remove the liquid phase and the suspended CPG from the cuvette. The chemiluminescence associate with the PMP remaining in the cuvette was then measured. This noncompetitive T3 assay exhibited a 10-fold lower detection limit than the equivalent competitive T3 assay, i.e., 0.3 vs pg/test. Imprecision (CV) in the clinically significant range was 6% or less. The assay also displayed two- to sevenfold lower cross-reactivities and a wider dynamic range.

Acridines↗
PubMed1993

[Immunoassay].

Since the advent of radioimmunoassay in 1959, the immunoassay technology has developed enormously as a very sensitive assay system suitable for wide range of analytes of clinical interest. Current popularity of the noncompetitive principle is attributable to its higher sensitivity, wide dynamic range and shorter incubation time. Recent innovation includes time-resolved fluorescent immunoassay designed to eliminate non-specific fluorescence, amplified enzyme-linked immunoassay and enhanced chemiluminescent immunoassay for ultra-sensitivity, particle counting immunoassay with a high resolution power, simultaneous measurement of multiple analytes, and easy-to-perform multilayer film immunoassays. Despite these advances in the technology, there remain various assay problems such as lack of standardization in reagents, a large interlaboratory variations despite the use of same assay kits, non-linear results in diluted samples and so-called prozone phenomenon.

Immunoassay↗
PubMed1998

Encoding of acoustic directional information by saccular afferents of the sleeper goby, Dormitator latifrons.

This paper reports on directional response properties of saccular afferents of the sleeper goby, Dormitator latifrons, to 100-Hz acoustic particle motions with a focus on testing the hypothesis that the response directionality of a fish's auditory afferents derives from the morphological polarity of sensory hair cells in the otolithic organs. Spontaneous rates (SR) and best sensitivities (BS) of saccular afferents ranged from 0 to 162 spikes/sec and from 0.2-to 100-nm RMS displacement. SR did not vary with BS. Most saccular afferents were phase-locked to sinusoidal stimulation and had sustained temporal response patterns with some adaptation. All saccular afferents were directionally sensitive to the stimulus, and the sharpness of directional response curves was determined by a directionality index (DI). The DI ranged from 0.64 to 1.50 (mean = 1.02, SE = 0.02, n = 100) and gradually decreased with stimulus level throughout afferents' response dynamic range. Many afferents had approximately symmetric directional response curves relative to their best response axes (BRA). BRA of most afferents remained constant with stimulus level. The BRA distribution had a peak along an axis that correlates closely with the morphological polarity of saccular hair cells. Therefore, our results strongly support the hypothesis.

Acoustic Stimulation↗
PubMed1989

Development of a microsphere-based fluorescent immunoassay and its comparison to an enzyme immunoassay for the detection of antibodies to three antigen preparations from Candida albicans.

A sensitive assay for the simultaneous detection of multiple serum antibodies by flow cytometry was developed. Polystyrene microspheres of 5, 7 and 9.3 micron in diameter were used as solid supports for the attachment of three different antigen preparations from Candida albicans. These antigens were a whole cell extract; a cytoplasmic protein extract and a cell wall polysaccharide. Microsphere-associated fluorescence was quantitated by flow cytometry, with the different sized microspheres analyzed separately using electronic volume gating. This procedure allowed for different antigen-coated microspheres with discrete sizes to be analyzed independently for immunofluorescence. The assay detected antibody levels in human serum at dilutions up to 10(-6) and provided complete discrimination, using all three antigen preparations, between antibody levels seen in healthy subjects and those seen in patients suspected of having a systemic Candida infection. A standard enzyme immunoassay (EIA) failed to provide complete discrimination between healthy subjects and patient samples: at least 17% of patient values fell within the healthy subject range using all three antigen preparations. The microsphere assay which allowed for the simultaneous detection of multiple antibodies, has increased dynamic range over EIA and provides for better discrimination of patients from healthy subjects in comparison to EIA. Precise quantitation of antibodies is possible and the rapid analysis of thousands of microspheres markedly enhances the statistical accuracy of the assay. We suggest this assay is likely to have many other important applications in immunologic testing.

Antibodies, Fungal↗
PubMed2005

Comparison of the produced and perceived voice range profiles in untrained and trained classical singers.

Frequency and intensity ranges (in true decibel sound pressure level, 20 microPa at 1 m) of voice production in trained and untrained vocalists were compared with the perceived dynamic range (phons) and units of loudness (sones) of the ear. Results were reported in terms of standard voice range profiles (VRPs), perceived VRPs (as predicted by accepted measures of auditory sensitivities), and a new metric labeled as an overall perceptual level construct. Trained classical singers made use of the most sensitive part of the hearing range (around 3-4 kHz) through the use of the singer's formant. When mapped onto the contours of equal loudness (depicting nonuniform spectral and dynamic sensitivities of the auditory system), the formant is perceived at an even higher sound level, as measured in phons, than a flat or A-weighted spectrum would indicate. The contributions of effects like the singer's formant and the sensitivities of the auditory system helped the trained singers produce 20% to 40% more units of loudness, as measured in sones, than the untrained singers. Trained male vocalists had a maximum overall perceptual level construct that was 40% higher than the untrained male vocalists. Although the A-weighted spectrum (commonly used in VRP measurement) is a reasonable first-order approximation of auditory sensitivities, it misrepresents the most salient part of the sensitivities (where the singer's formant is found) by nearly 10 dB.

Adult↗
PubMed2005

Estimating the clinical usefulness of optic disc biometry for detecting glaucomatous change over time.

PURPOSE: First, to determine the absolute measurement precision of scanning laser ophthalmoscopy (SLO) parameters, by expressing them as 95% limits of agreement (LA(95%)). Second, to propose a method for mathematically estimating the clinical ability of a parameter to monitor disease progression, expressed as the Discriminating Capacity Index (DCI). METHODS: We measured the optic disc of 14 healthy volunteers and 14 glaucoma patients. LA(95%)-values were calculated from the average standard deviation of three measurements on the same day for repeatability, and three measurements on separate days within a 6-week period for reproducibility. We then calculated the DCI by dividing the measurement range by its LA(95%) in healthy subjects and glaucoma patients separately. Thus, the DCI takes into account both the dynamic range of disease progression and the extent of measurement variance, providing an index of the possible clinical usefulness of a parameter. As the DCI is dimensionless it allows comparison across various parameters and across technologies. RESULTS: In the glaucoma group, the SLO parameters with the highest DCIs were 'volume below' (DCI, 9.38) and 'mean contour depth' (DCI, 8.02). In the healthy group, 'Neuroretinal rim area' had the highest index (DCI, 2.15). CONCLUSION: SLO optic disc biometry is uniformly reproducible and may prove a clinically useful method for glaucoma follow-up, due to the high DCI found for several parameters. The capacity to detect conversion from health to glaucoma is less pronounced, possibly due to a larger biological variability found in healthy volunteers.

Biometry↗
PubMed1994

Second-derivative spectrophotometric determination of naproxen in the presence of its metabolite in human plasma.

A second-derivative spectrophotometric method for the determination of naproxen in the absence or presence of its 6-desmethyl metabolite in human plasma is described. The method consists of direct extraction of the non-ionized form of the drug with pure diethyl ether and determination of the naproxen by measuring the peak amplitude (mm) in the second-order derivative spectrum at a wavelength of 328.2 nm. The efficiency of the extraction procedure expressed by the absolute recovery was 94.6 +/- 0.7% (mean +/- s) for the concentration range tested, and the limit of quantification attained according to the IUPAC definition was 2.42 mg l-1. The linear dynamic range for naproxen was 5.0-100.0 mg l-1, the correlation coefficient for the calibration graphs was excellent, r = 0.99993 (n = 6), the precision (Sr) was better than 4.58% and the accuracy was satisfactory (Er < 2.32%). The results obtained by the proposed method were in good agreement with those found by an HPLC method.

Biotransformation↗
PubMed2005

A new type of gradiometer for the receiving circuit of magnetic induction tomography (MIT).

Magnetic induction tomography (MIT) is a low-resolution imaging modality which aims at the three-dimensional (3D) reconstruction of the electrical conductivity in objects from alternating magnetic fields. In MIT systems the magnetic field perturbations to be detected are very small when compared to the excitation field (ppm range). The voltage which is induced by the excitation field in the receiver coils must be suppressed for providing sufficient dynamic range. In the past, two very efficient strategies were proposed: adjusted planar gradiometers (PGRAD) and the orientation of a receiver coil with respect to the excitation coil such that the net magnetic flow is zero (zero flow coil, ZFC). In contrast to the PGRAD no voltage is induced in the ZFC by the main field. This is advantageous because two comparatively high voltages in the two gradiometer coils can never be subtracted perfectly, thus leaving a residual voltage which is prone to drift. However, a disadvantage of the ZFC is the higher susceptibility to interferences from far RF sources. In contrast, in the gradiometer such interferences are cancelled to a high degree. We developed a new type of gradiometer (zero flow gradiometer, ZFGRAD) which combines the advantages of ZFC and PGRAD. All three systems were compared with respect to sensitivity and perturbation to signal ratio (PSR) defined as the ratio of the signal change due to a magnetic perturbation field at the carrier frequency and the signal change due to shifting a metallic sphere between two test points. The spatial sensitivity of the three systems was found to be very similar. The PSR of the ZFGRAD was more than 12 times lower than that of the ZFC. Finally, the feasibility of image reconstruction with two arrays of eight excitation coils and eight ZFGRAD, respectively, was shown with a single-step Gauss-Newton reconstructor and simulated measurement data generated for a cylindrical tank with a spherical perturbation. The resulting images show a clear, bright feature at the correct position of the sphere and are comparable to those with PGRAD arrays.

Animals↗
PubMed1995

The use of exogenous fluorescent probes for temperature measurements in single living cells.

The fluorescent membrane probes 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) and 6-dodecanoyl-2-dimethylamino-naphthalene (laurdan) have been studied for use as optical thermometers in living cells. The thermal sensitivity of NBD is primarily a consequence of rapid, heat-induced electronic changes, which increase the observed fluorescence decay rate. As a result, fluorescence intensity and lifetime variations of membrane-bound NBD-conjugated phospholipids and fatty acids can be directly correlated with cellular temperature. In contrast, laurdan fluorescence undergoes a dramatic temperature-dependent Stokes shift as the membrane undergoes a gel-to-liquid-crystalline phase transition. This facilitates the use of fluorescence spectra to record the indirect effect of microenvironmental changes, which occur during bilayer heating. Microscope and suspension measurements of cells and phospholipid vesicles are compared for both probes using steady-state and fluorescence lifetime (suspension only) data. Our results show that NBD fluorescence lifetime recordings can provide reasonable temperature resolution (approximately 2 degrees C) over a broad temperature range. Laurdan's microenvironmental sensitivity permits better temperature resolution (0.1-1 degree C) at the expense of a more limited dynamic range that is determined solely by bilayer properties. The temperature sensitivity of NBD is based on rapid intramolecular rotations and vibrations, while laurdan relies on a slower, multistep mechanism involving bilayer rearrangement, water penetration and intermolecular processes. Because of these differences in time scale, NBD appears to be more suitable for monitoring ultrafast phenomena, such as the impact of short-pulse microirradiation on single cells.

2-Naphthylamine↗
PubMed1994

Somatosensory, multisensory, and task-related neurons in cortical area 7b (PF) of unanesthetized monkeys.

1. The goal of this study was to quantitatively characterize the response properties of somatosensory and multisensory neurons in cortical area 7b (or PF) of monkeys that were behaviorally trained to perform an appetitive tolerance-escape task. Particular emphasis was given to characterizing nociceptive thermal responses and correlating such responses to thermal pain tolerance as measured by escape frequency. 2. A total of 244 neurons that responded to somatosensory stimulation alone or to both somatosensory and visual stimulation (multisensory) were isolated and studied in the trigeminal region of cortical area 7b. Thirty neurons responded only to visual stimulation. Thermoreceptive neurons formed approximately 13% (31 of 244) of the neurons that had somatosensory response properties. Thermal nociceptive neurons made up approximately 9% (21 of 244) of the neurons that had somatosensory response properties or approximately 68% (21 of 31) of the neurons that had thermoreceptive response properties. Thermal nociceptive neurons responded either exclusively to noxious thermal stimuli (high-threshold thermoreceptive, HTT) or differentially to nonnoxious and noxious thermal stimuli (wide-range thermoreceptive, WRT). Multimodal HTT neurons had nonnociceptive (low-threshold mechanoreceptive, LTM) and/or nociceptive (nociceptive-specific, wide-dynamic-range) mechanical receptive fields, whereas multimodal WRT neurons had only nonnociceptive (LTM) mechanical receptive fields. Thermal nonnociceptive neurons (low-threshold thermoreceptive, LTT) made up approximately 3% (8 of 244) of the neurons that had somatosensory properties or approximately 26% (8 of 31) of the neurons that were thermoreceptive. The background discharge of two thermoreceptive neurons (6%, 2 of 31) was inhibited by innocuous thermal stimulation. 3. Thermal nociceptive neurons (HTT and WRT) were functionally differentiated by statistical analyses into subpopulations that did encode (HTT-EN, WRT-EN) and did not encode (HTT-NE, WRT-NE) the magnitude of noxious thermal stimulus intensities. The mean slopes and median regression coefficients for the stimulus-response (S-R) functions of HTT-EN and WRT-EN neurons, respectively, were significantly greater than those for the S-R functions of HTT-NE and WRT-NE neurons. In contrast to HTT-NE and WRT-NE neurons, HTT-EN and WRT-EN neurons reliably encoded the magnitude of noxious thermal intensity by grading their mean discharge frequency. 4. The S-R functions of HTT-EN and WRT-EN neurons, unlike those of HTT-NE and WRT-NE neurons, closely approximated stimulus intensity-escape frequency functions.(ABSTRACT TRUNCATED AT 400 WORDS)

Afferent Pathways↗
PubMed1984

Tactile sensory coding during development: signaling capacities of neurons in kitten dorsal column nuclei.

The functional maturation of cuneate neurons was studied using reproducible tactile stimulation procedures and quantitative assessment of coding capacities in anesthetized (N2O/O2 plus barbiturate infusion) or decerebrate cats from six different age groups; neonatal (1-5 days), 10-15, 25-30, 55-63, and 83-90 days, and an adult group. Cuneate neurons were distinguished from input fibers to the nucleus on criteria of spike configuration and time course and on response profiles. Extracellular spike durations underwent a progressive shortening with age, reaching the adult range at approximately 3 mo when background activity levels also reached maturity. Despite an increase in conduction-path length, response latencies decreased to adult values by 25-30 days of age, presumably reflecting the increased conduction velocity in the input fibers. In each age group three functional classes of neurons responsive to tactile stimulation of the footpads were identifiable. One consisted of slowly adapting neurons whose stimulus-response relations resembled those of the adult in responsiveness and dynamic range by 25-30 days postnatal age. The remaining neurons were purely dynamically sensitive, but among them two classes could be distinguished by their differential sensitivity to cutaneous vibration, one receiving rapidly adapting (RA) fiber input the other pacinian corpuscle (PC) input. The combined bandwidth of vibration sensitivity in dynamically sensitive cuneate neurons expands from approximately 5-300 Hz in the neonate to the mature range of 5-1,000 Hz by 1 mo of age. The PC class of cuneate neurons showed an upward shift in peak sensitivity from 30-200 Hz in the neonate to 100-300 Hz at 10-15 days, reaching adult values of 200-600 Hz by 25-30 days. Over this period absolute thresholds dropped by an order of magnitude at 200-500 Hz reflecting the threshold trends in primary PC fibers. During the first 1-2 postnatal months, the capacity of cuneate PC neurons to signal information in a pattern code about vibration frequencies around 300 Hz is restricted because of poor phase locking and low responsiveness, which preclude an impulse periodicity reflecting the vibratory frequency.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1995

Development and validation of an automated particle-enhanced nephelometric immunoassay method for the measurement of human plasma C1q.

We have developed a sensitive immunoassay based on latex particle agglutination for measuring C1q concentrations in human plasma. In this simple and fast particle-enhanced immunoassay, we used carboxylated latex particles (diameter 210 nm) covalently coated with F(ab')2 fragments of anti-C1q antibodies. These particles are incubated with diluted sample (400-fold) for 6 min at room temperature, with the resulting agglutination quantified by measuring the change of light-scatter produced. The assay has been automated on the Behring nephelometer analyzer with a sampling rate of 150 samples/hr. This assay generates a standard curve in the range of 24-775 mg/L, showing intraassay and interassay precision of < 8% and < 10%, respectively. Dilution linearity was validated throughout the dynamic range of the assay. There were no interferences from bilirubin, Intralipid, haemoglobin, and rheumatoid factor. Results obtained in 45 clinical samples correlated well with those obtained by a commercial radial immunodiffusion method (r = 0.936), and with those obtained by the Behring immunoprecipitation nephelometric test (r = 0.950). The mean concentration in plasma from healthy subjects was 180 mg/L and the reference interval was from 128 to 237 mg/L. This latex nephelometric procedure is a convenient method and an interesting alternative to other immunoassays for routine measurement of human C1q.

Age Distribution↗
PubMed2001

Balloon-borne near-infrared diode laser spectroscopy for in situ measurements of atmospheric CH4 and H2O.

Absorption spectroscopy with near-infrared telecommunication laser diodes is a very convenient technique to measure in situ methane and water vapor in both the troposphere and the lower stratosphere (LS) and thereby to address many topics in the science of the atmosphere. This technique offers a high temporal resolution that ranges from 10 ms to 1 s, a precision error in the concentration retrieval of within a few percents and a dynamic range for the measurements of four orders of magnitude. A balloon-borne near-infrared diode laser spectrometer is described that provides simultaneous in situ methane (in the 1.65-microm region) and water vapor (in the 1.39-microm region) measurements at 1 s intervals. Tropospheric and stratospheric vertical concentration profiles of methane and water vapor are reported.

Atmosphere↗
PubMed1994

Comprehensive two-dimensional gas chromatography for the fast separation and determination of pesticides extracted from human serum.

A comprehensive two-dimensional gas chromatograph with flame ionization detection was constructed and evaluated for the fast separation and analysis of pesticides. A two-stage thermal desorption modulator served as an interface between the two capillary GC columns. By controlling the temperature of the modulator chamber, all sample substances covering a wide polarity and volatility range were modulated without sample breakthrough. Orthogonal separations were achieved with a nonpolar first column and a moderately polar second column. The system allows fast separations of complex mixtures. When we used the method to analyze pesticides extracted from human samples, we achieved complete separation of 15 pesticides in less than 4 min. The flame ionization detector gave detection limits for particular pesticides that ranged from 1.8 to 3.8 pg on-column. The relative standard deviations were from 6.2% to 8.8% over the linear dynamic range.

Chromatography, Gas↗
PubMed1998

Receptor clustering as a cellular mechanism to control sensitivity.

Chemotactic bacteria such as Escherichia coli can detect and respond to extremely low concentrations of attractants, concentrations of less than 5 nM in the case of aspartate. They also sense gradients of attractants extending over five orders of magnitude in concentration (up to 1 mM aspartate). Here we consider the possibility that this combination of sensitivity and range of response depends on the clustering of chemotactic receptors on the surface of the bacterium. We examine what will happen if ligand binding changes the activity of a receptor, propagating this change in activity to neighbouring receptors in a cluster. Calculations based on these assumptions show that sensitivity to extracellular ligands increases with the extent of spread of activity through an array of receptors, but that the range of concentrations over which the array works is severely diminished. However, a combination of low threshold of response and wide dynamic range can be attained if the cell has both clusters and single receptors on its surface, particularly if the extent of activity spread can adapt to external conditions. A mechanism of this kind can account quantitatively for the sensitivity and response range of E. coli to aspartate.

Aspartic Acid↗
PubMed2006

Use of new T-cell-based cell lines expressing two luciferase reporters for accurately evaluating susceptibility to anti-human immunodeficiency virus type 1 drugs.

Two new T-cell-based reporter cell lines were established to measure human immunodeficiency virus type 1 (HIV-1) infectivity. One cell line naturally expresses CD4 and CXCR4, making it susceptible to X4-tropic viruses, and the other cell line, in which a CCR5 expression vector was introduced, is susceptible to both X4- and R5-tropic viruses. Reporter cells were constructed by transfecting the human T-cell line HPB-Ma, which demonstrates high susceptibility to HIV-1, with genomes expressing two different luciferase reporters, HIV-1 long terminal repeat-driven firefly luciferase and cytomegalovirus promoter-driven renilla luciferase. Upon HIV infection, the cells expressed firefly luciferase at levels that were highly correlated (r2=0.91 to 0.98) with the production of the capsid antigen p24. The cells also constitutively expressed renilla luciferase, which was used to monitor cell numbers and viability. The reliability of the cell lines for two in vitro applications, drug resistance phenotyping and drug screening, was confirmed. As HIV-1 efficiently replicated in these cells, they could be used for multiple-round replication assays as an alternative method to a single-cycle replication protocol. Coefficients of variation for drug susceptibility evaluated with the cell lines ranged from 17 to 41%. The new cell lines were beneficial for evaluating antiretroviral drug resistance. Firefly luciferase gave a wider dynamic range for evaluating virus infectivity, and the introduction of renilla luciferase improved assay reproducibility. The cell lines were also beneficial for screening new antiretroviral agents, as false inhibition caused by the cytotoxicity of test compounds was easily detected by monitoring renilla luciferase activity.

Anti-HIV Agents↗
PubMed2006

Liquid deformable mirror for high-order wavefront correction.

We propose and demonstrate a novel liquid deformable mirror, based on electrocapillary actuation, for high-order wavefront correction. The device consists of a two-dimensional array of vertically oriented microchannels filled with two immiscible liquids, an aqueous electrolyte, and a viscous dielectric liquid, where the dielectric liquid overfills the top end of the channel and forms a thin layer on top. To remedy the poor reflectivity of pure liquids, a free-floating reflective membrane or a dye-coated liquid can be used. The proposed device offers several advantages for adaptive optics applications. These advantages include a high number of actuators, high stroke dynamic range, low power dissipation, fast response time, an initially flat surface, and low cost. However, the device is mainly suitable for dynamic wavefront correction and is limited by its orientation.

Journal Article↗