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At least 757 records · Page 42Linked to original sources

Optimization of immunofluorescence methods by quantitative image analysis.

There is a growing trend towards the objective quantification of immunohistochemical staining. However, quantification has not been used previously to optimize the original published immunohistochemical methods. We present a quantitative method for analyzing immunofluorescence staining employing the Applied Imaging MAGISCAN image analysis system, which has then been used to optimize major aspects of the standard immunofluorescent staining protocols. The optimization process resulted in a method that increased specific staining up to fivefold over typical published protocols, with no increase in nonspecific staining. The method is extremely reproducible. For slides stained by a single experimenter in one batch on one day, the coefficient of variation between replicate means is 1.2%. The image analysis protocol gave a linear response with increasing antigen concentration, as determined by using purified antigen dried onto slides. The revisions to the standard protocol presented here can also be applied to nonquantitative staining. It will help users of immunofluorescence to maximize their staining and may enable the detection of previously undetected antigens.

Actins↗

Modification of the mean-square error principle to double the convergence speed of a special case of Hopfield neural network used to segment pathological liver color images.

BACKGROUND: This paper analyzes the effect of the mean-square error principle on the optimization process using a Special Case of Hopfield Neural Network (SCHNN). METHODS: The segmentation of multidimensional medical and colour images can be formulated as an energy function composed of two terms: the sum of squared errors, and a noise term used to avoid the network to be stacked in early local minimum points of the energy landscape. RESULTS: Here, we show that the sum of weighted error, higher than simple squared error, leads the SCHNN classifier to reach faster a local minimum closer to the global minimum with the assurance of acceptable segmentation results. CONCLUSIONS: The proposed segmentation method is used to segment 20 pathological liver colour images, and is shown to be efficient and very effective to be implemented for use in clinics.

Algorithms↗

Optimization of polylactic-co-glycolic acid nanoparticles containing itraconazole using 2(3) factorial design.

This study investigated the utility of a 2(3) factorial design and optimization process for polylactic-co-glycolic acid (PLGA) nanoparticles containing itraconazole with 5 replicates at the center of the design. Nanoparticles were prepared by solvent displacement technique with PLGA X1 (10, 100 mg/mL), benzyl benzoate X2 (5, 20 microg/mL), and itraconazole X3 (200, 1800 microg/mL). Particle size (Y1), the amount of itraconazole entrapped in the nanoparticles (Y2), and encapsulation efficiency (Y3) were used as responses. A validated statistical model having significant coefficient figures (P < .001) for the particle size (Y1), the amount of itraconazole entrapped in the nanoparticles (Y2), and encapsulation efficiency (Y3) as function of the PLGA (X1), benzyl benzoate (X2), and itraconazole (X3) were developed: Y1 = 373.75 + 66.54X1+ 52.09X2 + 105.06X3 - 4.73X1X2 + 46.30X1X3; Y2 = 472.93 + 73.45X1 + 169.06X2 + 333.03X3 + 62.40X1X3 + 141.49X2X3; Y3 = 57.36 + 6.53X1 + 15.52X2 - 12.59X3 + 1.01X1X3 + 1.73X2X3. X1, X2, and X3 had a significant effect (P < .001) on Y1, Y2, and Y3. The particle size, the amount of itraconazole entrapped in the nanoparticles, and the encapsulation efficiency of the 4 formulas were in agreement with the predictions obtained from the models (P < .05). An overlay plot for the 3 responses shows the boundary in which Y1 shows the boundary in which a number of combinations of concentration of PLGA, benzyl benzoate, and itraconazole will result in a satisfactory process. Using the desirability approach with the same constraints, the solution composition having the highest overall desirability (D = 0.769) was 10 mg/mL of PLGA, 16.94 microg/mL of benzyl benzoate, and 1001.01 microg/mL of itraconazole. This approach allowed the selection of the optimum formulation ingredients for PLGA nanoparticles containing itraconazole of 500 microg/mL.

Antifungal Agents↗

Current bioremediation practice and perspective.

The use of microbes to clean up polluted environments, bioremediation, is a rapidly changing and expanding area of environmental biotechnology. Although bioremediation is a promising approach to improve environmental conditions, our limited understanding of biological contribution to the effect of bioremediation and its impact on the ecosystem has been an obstacle to make the technology more reliable and safer. Providing fundamental data to resolve these issues, i.e., the behavior of the target bacteria directly related to the degradation of contaminants and the changes in microbial communities during bioremediation, has been a challenge for microbiologists since many environmental bacteria cannot yet be cultivated by conventional laboratory techniques. The application of culture-independent molecular biological techniques offers new opportunities to better understand the dynamics of microbial communities. Fluorescence in situ hybridization (FISH), in situ PCR, and quantitative PCR are expected to be powerful tools for bioremediation to detect and enumerate the target bacteria that are directly related to the degradation of contaminants. Nucleic acid based molecular techniques for fingerprinting the 16S ribosomal DNA (rDNA) of bacterial cells, i.e., denaturing gradient gel electrophoresis (DGGE) and terminal restriction fragment length polymorphism (T-RFLP), enable us to monitor the changes in bacterial community in detail. Such advanced molecular microbiological techniques will provide new insights into bioremediation in terms of process optimization, validation, and the impact on the ecosystem, which are indispensable data to make the technology reliable and safe.

Journal Article↗

Separation of gamma-globulins from porcine plasma by reversed micellar extraction.

Factors affecting the separation of gamma-globulins from porcine plasma using reversed micelles were screened based on a fractional factorial design. The optimal processing conditions for obtaining the maximum yield of gamma-globulins under various constraints of product purity were determined using response surface methodology (RSM) and nonlinear programming. Results showed that the pH and sodium chloride concentration of the aqueous phase, and the concentration of surfactant (bis-(2-ethylhexyl) sulfosuccinate sodium salt, AOT) of the organic phase were the most important factors affecting the extraction performance. An eighty-five percent product purity and ninety-seven percent yield were obtained under the extraction conditions of 400 mM NaCl, 350 mM AOT, and pH 7.0. The extract exhibited immunological reactivity against anti-pig IgG.

Journal Article↗

Statistical principle and methodology in the NISAN system.

The NISAN system is a new interactive statistical analysis program package constructed by an organization of Japanese statisticans. The package is widely available for both statistical situations, confirmatory analysis and exploratory analysis, and is planned to obtain statistical wisdom and to choose optimal process of statistical analysis for senior statisticians.

Computers↗

Meeting report: Structural determination of environmentally responsive proteins.

The three-dimensional structure of gene products continues to be a missing lynchpin between linear genome sequences and our understanding of the normal and abnormal function of proteins and pathways. Enhanced activity in this area is likely to lead to better understanding of how discrete changes in molecular patterns and conformation underlie functional changes in protein complexes and, with it, sensitivity of an individual to an exposure. The National Institute of Environmental Health Sciences convened a workshop of experts in structural determination and environmental health to solicit advice for future research in structural resolution relative to environmentally responsive proteins and pathways. The highest priorities recommended by the workshop were to support studies of structure, analysis, control, and design of conformational and functional states at molecular resolution for environmentally responsive molecules and complexes; promote understanding of dynamics, kinetics, and ligand responses; investigate the mechanisms and steps in posttranslational modifications, protein partnering, impact of genetic polymorphisms on structure/function, and ligand interactions; and encourage integrated experimental and computational approaches. The workshop participants also saw value in improving the throughput and purity of protein samples and macromolecular assemblies; developing optimal processes for design, production, and assembly of macromolecular complexes; encouraging studies on protein-protein and macromolecular interactions; and examining assemblies of individual proteins and their functions in pathways of interest for environmental health.

Environmental Health↗

Iterative reverse optimization procedure for calibration of aspheric wave-front measurements on a nonnull interferometer.

The accuracy of interferometric measurements made with a nonnull configuration is degraded by test-dependent aberrations. Calibration of the data can be done with reverse optimization methods. An iterative reverse optimization process that improves efficiency and eliminates sensitive merit function weighting issues is described. The process is shown to calibrate a nonnull interferometric measurement of a wave front with more than 200 waves of departure to an accuracy of 0.16 waves peak to valley and 0.02 waves rms.

Journal Article↗

Phase demodulation method from a single fringe pattern based on correlation with a polynomial form.

The method presented extracts the demodulated phase from only one fringe pattern. Locally, this method approaches the fringe pattern morphology with the help of a mathematical model. The degree of similarity between the mathematical model and the real fringe is estimated by minimizing a correlation function. To use an optimization process, we have chosen a polynomial form such as a mathematical model. However, the use of a polynomial form induces an identification procedure with the purpose of retrieving the demodulated phase. This method, polynomial modulated phase correlation, is tested on several examples. Its performance, in terms of speed and precision, is presented on very noised fringe patterns.

Journal Article↗

Frequency-domain optical imaging of absorption and scattering distributions by a Born iterative method.

We presents a Born; iterative method, for reconstructing optical properties of turbid media by means of frequency-domain data. The approach is based on iterative solution of a linear perturbation equation, which is derived from the integral from of the Helmholtz wave equation for photon-density waves in each iteration the total field and the associated weight matrix are recalculated based on the previous reconstructed image. We then obtain a new estimate by solving the updated perturbation equation. The forward solution, also based on a Helmholtz equation, is obtained by a multigrid finite difference method. The inversion is carried out through a Tikhonov regularized optimization process by the conjugate gradient descent method. Using this method, we first reconstruct the distribution of the complex wave numbers in a test medium, from which the absorption and the scattering distributions are then derived. Simulation results with two-dimensional test media have shown that this method can yield quantitatively (in terms of coefficient valued) as well as qualitatively (in terms of object location and shape) accurate reconstructions of absorption and scattering distributions in cases in which the first-order Born approximation cannot work well. Both full-angle and limited-angle measurement schemes have been simulated to examine the effect of the location of detectors and sources. The robustness of the algorithm to noise has also been evaluated.

Absorption↗

Intensity control with a free-form lens.

A family of free-form lenses for intensity control is designed. The lens can shape an incident collimated beam with a given intensity distribution I1 into a new collimated beam with intensity distribution I2. No symmetry is assumed for the two intensity profiles. The key idea is that the lens design problem can be formulated and solved in terms of an optimization process involving a specific action functional. It is further shown that the free-form lens can be manufactured by a surfacing process using a convex tool.

Journal Article↗

Differential toolbox to shape dispersion behavior in photonic crystal fibers.

We present an analytical procedure to compute the first derivatives of the propagation constants with respect to several structural parameters in photonic crystal fibers (PCFs). From them we can easily evaluate the same derivatives of other directly related magnitudes. The above derivatives provide the trend of the magnitude at issue, which allows us to take advantage of a gradient-based algorithm to shape the properties of the guiding structure. In this way we implement an optimization process to carry out real inverse design in PCFs. We focus our attention on designing PCFs with a specific chromatic dispersion behavior. Likewise, the same approach makes it possible to analyze their fabrication tolerances.

Journal Article↗

Drug characterization in low dosage pharmaceutical tablets using Raman microscopic mapping.

Raman micro-spectroscopic mapping is utilized to analyze pharmaceutical tablets containing a low concentration (0.5% w/w) of active pharmaceutical ingredient (API). The domain sizes and spatial distributions of the API and the major excipients are obtained. Domain size of the API is found to be dependent upon the particle size distribution of the ingoing API material, making the Raman maps good indicators of the source of API used in tablet manufacturing. Multivariate classification was performed to simultaneously check for the presence of two undesired API polymorphs within tablets. Raman mapping was demonstrated capable of detecting in the tablet matrix as little as 10% form conversion of the low-dosage (0.5% w/w) API, which is equivalent to detection of a 0.05% w/w polymorphic impurity. Overall, the information provided by Raman micro-spectroscopic mapping was found to have potential utility for manufacturing process optimization or predictive stability assessments.

Drug Industry↗

Evaluation of optimization techniques for an extractive alcoholic fermentation process.

The mathematical optimization of a continuous alcoholic fermentation process combined with a flash column under vacuum was studied. The objective was to maximize % yield and productivity in the fermentor. The results using surface response analysis combined with modeling and simulation were compared withy those obtained when the problem was written as a nonlinear programming problem and was solved with a successive quadratic programming (SQP) technique. Two process models were evaluated when the process was optimized using the SQP technique. The first one is a deterministic model, whose kinetic parameters were experimentally determined as functions of the temperature, and the second is a statistical model obtained using the factorial design technique combined with simulation. Although the best result was the one obtained using the rigorous model, the values for productivity and % yield obtained using the simplified model are acceptable, and these models can be used when the development of a rigorous model is excessively difficult, slow, or expensive.

Alcohols↗

Effects of hemicellulose and lignin on enzymatic hydrolysis of cellulose from dairy manure.

This study focused on the effect of hemicellulose and lignin on enzymatic hydrolysis of dairy manure and hydrolysis process optimization to improve sugar yield. It was found that hemicellulose and lignin in dairy manure, similar to their role in other lignocellulosic material, were major resistive factors to enzymatic hydrolysis and that the removal of either of them, or for best performance, both of them, improved the enzymatic hydrolysis of manure cellulose. This result combined with scanning electron microscope (SEM) pictures further proved that the accessibility of cellulose to cellulase was the most important feature to the hydrolysis. Quantitatively, fed-batch enzymatic hydrolysis of fiber without lignin and hemicellulose had a high glucose yield of 52% with respect to the glucose concentration of 17 g/L at a total enzyme loading of 1300 FPU/L and reaction time of 160 h, which was better than corresponding batch enzymatic hydrolysis.

Carbohydrates↗

Application of design-of-experiments procedures to optimize efficiently pretreatment of lipase for use in a nonaqueous reaction.

A variety of different pretreatments can improve the performance of enzymes in nonpolar reaction media. These pretreatments have primarily been studied in isolation; however, interactions between some pairs of pretreatments are known to exist. The presence of these interactions complicates the design of an optimum multifactor pretreatment. Modern design-of-experiments techniques allow the simultaneous optimization of two or more variables. To improve the performance of lipase in a model reaction, we used a technique called the method of steepest ascent to optimize three variables simultaneously: pretreatment pH and sodium phosphate concentration, and the concentration of acetic acid (one of the reactants) in the reaction mixture. In only 26 experimental runs, this optimization process determined a combination of variable settings that yielded a reaction product approx 180 times faster than achieved with untreated enzyme. Evidence is presented to demonstrate that locating this optimum with single-factor experiments would be inefficient. This article demonstrates the efficiency of the method of steepest ascent particularly for evaluation of enzymatic reaction conditions exhibiting significant interactions.

Acetic Acid↗

Application of factorial design to study of heavy metals biosorption by waste biomass from beverage distillery.

A full factorial design leading to 20 sets of sorption runs was conducted to study the influence of four variables (bleaching earth and biomass concentrations, pH, and sorption time) on the iron, nickel, and chromium removal from stainless steel effluent using waste biomass from a beverage industry. Similar factor effects and interactions were found for each metal involved in this biosorption study, and the main factors were pH (positive effect) and biomass concentration (negative effect). Response surface methodology was adopted and an empirical linear polynomial model constructed on the basis of the specific uptake (mg of metal/g of biomass as dryweight) for each metal species. Under optimized process conditions (pH 4.0, biomass concentration of 2.0 g/L, absence of Celite), uptake values of 155 mg of Fe/g, 38 mg of Cr/g, and 0.4 mg of Ni/g were achieved after 3 h. This corresponded to a reduction in heavy metals concentration of approx 94% for Cr, 57% for Fe, and 25% for Ni.

Adsorption↗

Cerebrospinal fluid tau and Abeta42 concentrations in healthy subjects: delineation of reference intervals and their limitations.

Many limitations and conflicting results have cast serious doubts on the validity of cerebrospinal fluid tau and Abeta42 levels for the biological diagnosis of Alzheimer's disease, particularly extreme variations of the reference limits found by unrelated groups as a consequence of different reference populations used. In this study, we addressed the issue of defining reference limits for cerebrospinal fluid tau and Abeta42 in healthy adult individuals. One hundred and five neurologically intact subjects were enrolled according to strict inclusion criteria, 10 of them with autopsy confirmation of brain integrity. All cerebrospinal fluid samples were similarly and optimally processed as were the dosage methods used and the statistical analyses performed. A robust correlation with age was demonstrated for Abeta42 but not for tau. For tau, we found that an upper cut-off value of 443 ng/l allowed 95% of the subjects to be correctly classified as normal. For Abeta42, a lower cut-off value of 90 ng/l allowed a correct classification of 90% of the subjects. However, a large variance of the reference values, partly explained by the potential contamination of the reference population with presymptomatic dementia patients, may limit the use of reference limits based on living subjects. We propose that the issue of defining reference limits for both cerebrospinal fluid tau and Abeta42 may ultimately be settled by studying large numbers of autopsy-proven neurologically intact individuals only.

Adult↗