Automated ordering of fingerprinted clones

MOTIVATION: A considerable amount of human intervention is currently required to produce high-quality fingerprint-based physical maps for genomic studies. RESULTS: An algorithm has been developed and implemented to automatically order fingerprinted clones within contigs. The resulting software, named CORAL (Clone ORdering ALgorithm), has been tested on maps that have previously been manually edited and on maps derived from in silico simulations. The fingerprint map and DNA sequence of the human genome has provided an additional test to CORAL. Measurements suggest that CORAL performs significantly better than the software currently used by most laboratories to order fingerprinted clones at throughputs far exceeding those that can be achieved manually.     

Performances of aseptic sampling devices for on-line monitoring of fermentation processes

Two liquid sampling systems, which are set in the bioreactor and can be sterilized in place, are described. They are composed primarily of an inorganic membrane filter which is either stationary or is set in a rotating motion. These systems have been tested during three types of fermentation processes (ethanol, lactic acid, and polysaccharide productions). The rotating system gave better filtration performances than the stationary sampler which can be used only in alcoholic fermentation. The rotating sampler was coupled with a high-pressure liquid chromatograph for the on-line quantitation of the major compounds present in the filtrate. The results are in good agreement with those obtained from off-line analysis on samples taken manually. (c) 1992 John Wiley & Sons, Inc.     

A method for the rapid,automated filtration of protein hydrolyzates

A method for the automated filtration of protein hydrolyzates prior to amino acid analysis is described. Minor modification of a Technicon Sampler II enables it to function simultaneously as a sampler and a filtrate collector. Samples are drawn from cups in the sampler tray and are forced through a Teflon filter (pore size, 0.2 μm) in a Millipore Swinnex filter holder by a variable-speed Technicon proportioning pump. The filtrates are collected in cups in the sampler tray opposite those containing unfiltered hydrolyzates. Using this technique, 12 hydrolyzates can be filtered in 25 min compared to the approximately 2 h of technician time required for their manual filtration. Aliquots from each of 48 samples representing different proteins and hydrolysis conditions are filtered manually and by the automated technique. Analysis of variance of the resulting recoveries of each amino acid indicate little likelihood of effects due to filtration method.     

Direct visualization of fluorescent signals in protein gels using a backlit blue light plate

The visualization of fluorescently labeled protein gels is required for the analysis of electrophoretic results or manually picking protein bands or spots from gels. To accomplish this task, an UV light table is generally utilized, but may be hazardous to operators. The blue light transilluminator is another apparatus that may be used for this purpose, but is sometimes insufficient for revealing weak fluorescent signals. In this study, we invented a new setup utilizing a backlit blue light plate for illuminating fluorescently stained protein gels. This method employs Snell's law and allows for the direct visualization of fluorescent signals in protein gels without the use of a filter or filter glasses. This safe, convenient, economic, and effective setup was found to be an ideal alternative for illuminating fluorescent protein gels in proteomic experiments.     

Co-composting of filter cake and bagasse; by-products from a sugar mill

Thailand has nearly 2 million tonnes of filter cake waste containing 1.8% total N from the sugar cane industry to dispose of annually. Compost studies were conducted to determine how rapidly this material can be converted to a stable product that may be useful in crop production, and to characterize the N transformations. Two kinds of sugar mill by-products were composted, filter cake and filter cake mixed with bagasse, at a 2:1 ratio to reduce the C:N ratio in an attempt to reduce N loss during composting. Materials were mixed manually at 3-5 day intervals during the composting process. Both composts were analyzed at least weekly to measure temperature, pH, NH4+, NO3-, total N content, C loss, and germination index. For both mixtures, the thermophilic stage lasted 15-20 days and was higher than ambient for nearly 80 days. The degradation of organic matter (OM) was rapid in both mixtures to approximately 40 days, after which it began to stabilize. Both mixtures achieved maturity at approximately 90 days as indicated by a stable C/N, low NH4+/NO3-, lack of heat production and a germination index higher than 80%. Mixing filter cake with bagasse helped conserve N during composting. Because N was in excess, approximately 12-15% was lost from the composts. Mixing more bagasse with the filter cake may result in further reduction in N losses. Both composts have potential for use in crop production.     

A model-based high throughput method for fecundity estimation in fruit fly studies

ABSTRACT

The ability to quantify fecundity is critically important to a wide range of experimental applications, particularly in widely-used model organisms such as Drosophila melanogaster. However, the standard method of manually counting eggs is time consuming and limits the feasibility of large-scale experiments. We develop a predictive model to automate the counting of eggs from images of eggs removed from the media surface and washed onto dark filter paper. Our method uses the simple relationship between the white area in an image and the number of eggs present to create a predictive model that performs well even at high egg densities where clumping can complicate the individual identification of eggs. A cross-validation approach demonstrates our method performs well, with a correlation between predicted and manually counted values of 0.88. We show how this method can be applied to a large data set where egg densities vary widely.     

A New Approach to Segment Both Main and Peripheral Retinal Vessels Based on Gray-Voting and Gaussian Mixture Model

Vessel segmentation in retinal fundus images is a preliminary step to clinical diagnosis for some systemic diseases and some eye diseases. The performances of existing methods for segmenting small vessels which are usually of more importance than the main vessels in a clinical diagnosis are not satisfactory in clinical use. In this paper, we present a method for both main and peripheral vessel segmentation. A local gray-level change enhancement algorithm called gray-voting is used to enhance the small vessels, while a two-dimensional Gabor wavelet is used to extract the main vessels. We fuse the gray-voting results with the 2D-Gabor filter results as pre-processing outcome. A Gaussian mixture model is then used to extract vessel clusters from the pre-processing outcome, while small vessels fragments are obtained using another gray-voting process, which complements the vessel cluster extraction already performed. At the last step, we eliminate the fragments that do not belong to the vessels based on the shape of the fragments. We evaluated the approach with two publicly available DRIVE (Staal et al., 2004) and STARE (Hoover et at., 2000) datasets with manually segmented results. For the STARE dataset, when using the second manually segmented results which include much more small vessels than the first manually segmented results as the “gold standard,” this approach achieved an average sensitivity, accuracy and specificity of 65.0%, 92.1% and 97.0%, respectively. The sensitivities of this approach were much higher than those of the other existing methods, with comparable specificities; these results thus demonstrated that this approach was sensitive to detection of small vessels.     

Investigation of New Microstrip Bandpass Filter Based on Patch Resonator with Geometrical Fractal Slot

A compact dual-mode microstrip bandpass filter using geometrical slot is presented in this paper. The adopted geometrical slot is based on first iteration of Cantor square fractal curve. This filter has the benefits of possessing narrower and sharper frequency responses as compared to microstrip filters that use single mode resonators and traditional dual-mode square patch resonators. The filter has been modeled and demonstrated by Microwave Office EM simulator designed at a resonant frequency of 2 GHz using a substrate of ε_r = 10.8 and thickness of h = 1.27 mm. The output simulated results of the proposed filter exhibit 22 dB return loss, 0.1678 dB insertion loss and 12 MHz bandwidth in the passband region. In addition to the narrow band gained, miniaturization properties as well as weakened spurious frequency responses and blocked second harmonic frequency in out of band regions have been acquired. Filter parameters including insertion loss, return loss, bandwidth, coupling coefficient and external quality factor have been compared with different values of perturbation dimension (d). Also, a full comparative study of this filter as compared with traditional square patch filter has been considered.     

Boolean Combinations of Implicit Functions for Model Clipping in Computer-Assisted Surgical Planning

This paper proposes an interactive method of model clipping for computer-assisted surgical planning. The model is separated by a data filter that is defined by the implicit function of the clipping path. Being interactive to surgeons, the clipping path that is composed of the plane widgets can be manually repositioned along the desirable presurgical path, which means that surgeons can produce any accurate shape of the clipped model. The implicit function is acquired through a recursive algorithm based on the Boolean combinations (including Boolean union and Boolean intersection) of a series of plane widgets’ implicit functions. The algorithm is evaluated as highly efficient because the best time performance of the algorithm is linear, which applies to most of the cases in the computer-assisted surgical planning. Based on the above stated algorithm, a user-friendly module named SmartModelClip is developed on the basis of Slicer platform and VTK. A number of arbitrary clipping paths have been tested. Experimental results of presurgical planning for three types of Le Fort fractures and for tumor removal demonstrate the high reliability and efficiency of our recursive algorithm and robustness of the module.     

Numerical analysis of the hemodynamics and embolus capture of a greenfield vena cava filter

BACKGROUND: Vena Cava filters are used to prevent pulmonary embolism in patients with deep vein thrombosis who are unresponsive to anticoagulation therapy. Various filter designs exist in the market with different characteristics distinguishing them. An understanding of the characteristics of these filters is desirable in order to develop better designs. METHODS: A computational fluid dynamical study of the flow over an unoccluded stainless steel Greenfield Vena Cava filter (Boston Scientific, Watertown, MA) to determine its properties has been performed. Simulation of flow over a filter placed axisymmetrically in a rounded inferior vena cava has been performed at a Reynolds numbers of 1000 and the consequences of the flow (by studying parameters like shear stress and stagnation zones) have been discussed. Furthermore, a new finite element based numerical method has been developed that allows the study of capturing properties of Inferior Vena Cava filters. The key idea is the introduction of a thin-wire-model (TWM) that enables a drastic reduction in the computational cost while still maintaining control on the physics of the problem. This numerical technique has been applied to evaluate the embolus capture characteristic of a Greenfield filter. RESULTS: The flow around the unoccluded filter is found to be steady and laminar at the conditions studied. A recirculation/stagnation zone develops immediately downstream of the filter head. This zone is significantly larger when the central hole is occluded. The shear stress and stagnation zone properties for such a flow over a Greenfield filter are compared with existing literature (in vitro studies). A graph showing the regions wherein clots escape or get captured has been determined by a means of numerical simulations. The data has further been analyzed to determine the probability of clot capture as function of the clot size. CONCLUSIONS: The stagnation zone formed behind the head of the Greenfield filter is found to be smaller in size when compared to that of the same filter with the central hole occluded. A map of the shear stress distribution shows a small region having the potential for thrombogenesis. The non-Newtonian properties of blood are not seen to cause much variation in the flow field when compared to the Newtonian model. However variation in the cava size leads to a significant change in the shear stresses. This study also establishes a novel method wherein computational means are used to determine the efficacy of clot capturing of filters. These techniques can further be used to compare the different characteristics among filters.     

AraCyc: a biochemical pathway database for Arabidopsis

AraCyc is a database containing biochemical pathways of Arabidopsis, developed at The Arabidopsis Information Resource (http://www.arabidopsis.org). The aim of AraCyc is to represent Arabidopsis metabolism as completely as possible with a user-friendly Web-based interface. It presently features more than 170 pathways that include information on compounds, intermediates, cofactors, reactions, genes, proteins, and protein subcellular locations. The database uses Pathway Tools software, which allows the users to visualize a bird's eye view of all pathways in the database down to the individual chemical structures of the compounds. The database was built using Pathway Tools' Pathologic module with MetaCyc, a collection of pathways from more than 150 species, as a reference database. This initial build was manually refined and annotated. More than 20 plant-specific pathways, including carotenoid, brassinosteroid, and gibberellin biosyntheses have been added from the literature. A list of more than 40 plant pathways will be added in the coming months. The quality of the initial, automatic build of the database was compared with the manually improved version, and with EcoCyc, an Escherichia coli database using the same software system that has been manually annotated for many years. In addition, a Perl interface, PerlCyc, was developed that allows programmers to access Pathway Tools databases from the popular Perl language. AraCyc is available at the tools section of The Arabidopsis Information Resource Web site (http://www.arabidopsis.org/tools/aracyc).     

Detection and quantification of tau aggregation using a membrane filter assay

Aggregation of the microtubule-associated protein tau contributes to the formation of neurofibrillary lesions in Alzheimer's disease and is a useful marker of disease progression. Although filter trap assays have been employed to assess the extent of tau aggregation in cells and tissues as well as in vitro, their performance relative to other assay modalities has not been reported. To clarify this issue, the ability of the filter trap approach to quantify aggregation of purified recombinant full-length tau protein in vitro was examined as a function of membrane chemistry in a 96-well format. Results showed that nitrocellulose yielded the greatest assay sensitivity relative to polyvinylidene fluoride or cellulose acetate at equal membrane porosity. However, all combinations of filter chemistries, porosities, and monoclonal detection antibodies yielded nonlinear correlations between signal intensity and analyte concentration. When corrected for nonlinearity, the filter trap assay determined a value for the critical monomer concentration for tau aggregation that was statistically identical to determinations made by electron microscopy assay. The data suggest conditions under which filter trap assays can be used to estimate tau aggregation kinetics.     

An automated instrument for the performance of enzymatic DNA sequencing reactions

An instrument has been developed for the automation of enzymatic DNA sequencing reactions. Up to 96 DNA templates contained in a microtiter plate can be processed for either radioactive or fluorescence-based sequence analysis in a three-hour period. The quality of the resultant data is comparable to that obtained manually. The system is simple, flexible and is readily adapted to the use of new polymerases or modified experimental protocols.     

Method for rapid screening of pesticide mineralization in soil

A method has been developed for the analysis of (14)CO(2) evolution from the mineralization of (14)C-labelled organic compounds in soil samples. The new method is less space demanding and substantially cuts down laborious manual work compared to the traditional incubation bottle method used. Furthermore, the use of scintillation cocktail is largely reduced with the new method. In the new method, (14)CO(2) is trapped in filter paper held in the lid of a 20 ml glass vial by surface tension. The trapping solution used is Ca(OH)(2), which fixates CO(2) in the filter paper and the analysis of trapped (14)CO(2) is done using the Cyclone trade mark Storage Phosphor system. The lids are placed in a 32 well holder and exposed to a phosphor screen prior to scanning in a Cyclone trade mark scanner. The new filter method has been tested and compared to results obtained using the traditional method. The results show good agreement but due to a smaller capacity for CO(2) with the filter method compared to the traditional method, the interval between sampling has to be shorter using the filter method when the CO(2) development is high. The detection limits for the filter method is higher compared to the traditional method. With the filter method, the level of radioactivity has to exceed 300 dpm before detection is possible, while the same limit for the traditional method is around 30 dpm. On the other hand, the gas trapping faster and the efficiency is higher with the filter method.     

A text mining approach to detect mentions of protein glycosylation in biomedical text

Protein Glycosylation is an important post translational event that plays a pivotal role in protein folding and protein is trafficking. We describe a dictionary based and a rule based approach to mine ‘mentions‘ of protein glycosylation in text. The dictionary based approach relies on a set of manually curated dictionaries specially constructed to address this task. Abstracts are then screened for the ‘mentions‘ of words from these dictionaries which are further scored followed by classification on the basis of a threshold. The rule based approaches also relies on the words in the dictionary to arrive at the features which are used for classification. The performance of the system using both the approaches has been evaluated using a manually curated corpus of 3133 abstracts. The evaluation suggests that the performance of the Rule based approach supersedes that of the Dictionary based approach.     

Filter elution assays for DNA damage: practical and mechanistic significance of the DNA in the filter support wash.

The alkaline and neutral (or nondenaturing) filter elution assays are popular methods for the measurement of DNA strand breakage and its repair in eukaryotic cells. In both alkaline and neutral elution, it is recommended practice to wash the filter support after removal of the filter and to analyze the DNA recovered by this procedure together with that remaining on the filter as uneluted DNA, although it is not obvious why the DNA in the filter support wash should be so interpreted. We have observed that the sum of the DNA on the filter and that recovered in the filter support wash is approximately constant when the pH of the alkaline filter elution assay for total strand breaks is increased from 12.1 to 12.6, whereas the fraction on the filter itself is markedly smaller at the higher pH. This behavior characterized DNA elution from undamaged cells, as well as from cells treated with various DNA-damaging agents. These findings are consistent with the "tug-of-war" mechanism that has been proposed for alkaline elution, but are inconsistent with the simplest mechanism of the "sieve" class. In the neutral filter elution assay for double-strand breaks, by contrast, the distribution of DNA between the filter and the filter support wash is pH-independent. This suggests that single- and double-stranded DNA segments traverse a filter by different physical mechanisms. Our observations underscore the importance of carrying out the filter support wash and the analysis of the DNA it contains as uneluted DNA in alkaline elution, while indicating that a different analysis of this DNA might be appropriate for neutral elution.     

Comparative surface ultrastructure of adult Onchocerca volvulus recovered from human nodules by dissection or collagenase digestion

The surface ultrastructure of male and female adult worms of Onchocerca volvulus obtained from human nodules by the technique of collagenase digestion has been compared with that of worms excised manually without the aid of enzyme treatment. No topographical differences have been identified.     

Segmentation of left ventricle in short-axis echocardiographic sequences by weighted radial edge filtering and adaptive recovery of dropout regions

In this paper, we present a weighted radial edge filtering algorithm with adaptive recovery of dropout regions for the semi-automatic delineation of endocardial contours in short-axis echocardiographic image sequences. The proposed algorithm requires minimal user intervention at the end diastolic frame of the image sequence for specifying the candidate points of the contour. The region of interest is identified by fitting an ellipse in the region defined by the specified points. Subsequently, the ellipse centre is used for originating the radial lines for filtering. A weighted radial edge filter is employed for the detection of edge points. The outliers are corrected by global as well as local statistics. Dropout regions are recovered by incorporating the important temporal information from the previous frame by means of recursive least squares adaptive filter. This ensures fairly accurate segmentation of the cardiac structures for further determination of the functional cardiac parameters. The proposed algorithm was applied to 10 data-sets over a full cardiac cycle and the results were validated by comparing computer-generated boundaries to those manually outlined by two experts using Hausdorff distance (HD) measure, radial mean square error (rmse) and contour similarity index. The rmse was 1.83 mm with a HD of 5.12 ± 1.21 mm. We have also compared our results with two existing approaches, level set and optical flow. The results indicate an improvement when compared with ground truth due to incorporation of temporal clues. The weighted radial edge filtering algorithm in conjunction with adaptive dropout recovery offers semi-automatic segmentation of heart chambers in 2D echocardiography sequences for accurate assessment of global left ventricular function to guide therapy and staging of the cardiovascular diseases.     

A protocol for Papanicolaou staining of cytologic specimens following flow analysis

A protocol has been developed for restaining cytologic specimens that have been analyzed on a multidimensional slit-scan flow system. The technique involves Papanicolaou staining of cells on a membrane filter that has been previously stained with acridine orange and fixed with glutaraldehyde buffer. The specimen and staining solutions were sequentially added to a 5-micrometers pore size, 47-mm diameter Gelman "Metricel" filter while it remained in a glass filtration apparatus. The practice of retaining the filter in the filtration apparatus throughout the staining procedure minimizes cell loss and eliminates specimen cross contamination when compared with conventional filter dip staining. The availability of this postflow specimen Papanicolaou staining protocol permits accurate determination of the performance characteristics of a multidimensional slit-scan flow system and should be useful whenever staining of a limited number of cells with minimal cell loss is desired.