Near-common-path interferometer for imaging Fourier-transform spectroscopy in wide-field microscopy

PubMed Central

Wadduwage, Dushan N.; Singh, Vijay Raj; Choi, Heejin; Yaqoob, Zahid; Heemskerk, Hans; Matsudaira, Paul; So, Peter T. C.

2017-01-01

Imaging Fourier-transform spectroscopy (IFTS) is a powerful method for biological hyperspectral analysis based on various imaging modalities, such as fluorescence or Raman. Since the measurements are taken in the Fourier space of the spectrum, it can also take advantage of compressed sensing strategies. IFTS has been readily implemented in high-throughput, high-content microscope systems based on wide-field imaging modalities. However, there are limitations in existing wide-field IFTS designs. Non-common-path approaches are less phase-stable. Alternatively, designs based on the common-path Sagnac interferometer are stable, but incompatible with high-throughput imaging. They require exhaustive sequential scanning over large interferometric path delays, making compressive strategic data acquisition impossible. In this paper, we present a novel phase-stable, near-common-path interferometer enabling high-throughput hyperspectral imaging based on strategic data acquisition. Our results suggest that this approach can improve throughput over those of many other wide-field spectral techniques by more than an order of magnitude without compromising phase stability. PMID:29392168

High-throughput measurement of rice tillers using a conveyor equipped with x-ray computed tomography

NASA Astrophysics Data System (ADS)

Yang, Wanneng; Xu, Xiaochun; Duan, Lingfeng; Luo, Qingming; Chen, Shangbin; Zeng, Shaoqun; Liu, Qian

2011-02-01

Tillering is one of the most important agronomic traits because the number of shoots per plant determines panicle number, a key component of grain yield. The conventional method of counting tillers is still manual. Under the condition of mass measurement, the accuracy and efficiency could be gradually degraded along with fatigue of experienced staff. Thus, manual measurement, including counting and recording, is not only time consuming but also lack objectivity. To automate this process, we developed a high-throughput facility, dubbed high-throughput system for measuring automatically rice tillers (H-SMART), for measuring rice tillers based on a conventional x-ray computed tomography (CT) system and industrial conveyor. Each pot-grown rice plant was delivered into the CT system for scanning via the conveyor equipment. A filtered back-projection algorithm was used to reconstruct the transverse section image of the rice culms. The number of tillers was then automatically extracted by image segmentation. To evaluate the accuracy of this system, three batches of rice at different growth stages (tillering, heading, or filling) were tested, yielding absolute mean absolute errors of 0.22, 0.36, and 0.36, respectively. Subsequently, the complete machine was used under industry conditions to estimate its efficiency, which was 4320 pots per continuous 24 h workday. Thus, the H-SMART could determine the number of tillers of pot-grown rice plants, providing three advantages over the manual tillering method: absence of human disturbance, automation, and high throughput. This facility expands the application of agricultural photonics in plant phenomics.

High-throughput measurement of rice tillers using a conveyor equipped with x-ray computed tomography.

PubMed

Yang, Wanneng; Xu, Xiaochun; Duan, Lingfeng; Luo, Qingming; Chen, Shangbin; Zeng, Shaoqun; Liu, Qian

2011-02-01

Tillering is one of the most important agronomic traits because the number of shoots per plant determines panicle number, a key component of grain yield. The conventional method of counting tillers is still manual. Under the condition of mass measurement, the accuracy and efficiency could be gradually degraded along with fatigue of experienced staff. Thus, manual measurement, including counting and recording, is not only time consuming but also lack objectivity. To automate this process, we developed a high-throughput facility, dubbed high-throughput system for measuring automatically rice tillers (H-SMART), for measuring rice tillers based on a conventional x-ray computed tomography (CT) system and industrial conveyor. Each pot-grown rice plant was delivered into the CT system for scanning via the conveyor equipment. A filtered back-projection algorithm was used to reconstruct the transverse section image of the rice culms. The number of tillers was then automatically extracted by image segmentation. To evaluate the accuracy of this system, three batches of rice at different growth stages (tillering, heading, or filling) were tested, yielding absolute mean absolute errors of 0.22, 0.36, and 0.36, respectively. Subsequently, the complete machine was used under industry conditions to estimate its efficiency, which was 4320 pots per continuous 24 h workday. Thus, the H-SMART could determine the number of tillers of pot-grown rice plants, providing three advantages over the manual tillering method: absence of human disturbance, automation, and high throughput. This facility expands the application of agricultural photonics in plant phenomics.

High throughput imaging cytometer with acoustic focussing† †Electronic supplementary information (ESI) available: High throughput imaging cytometer with acoustic focussing. See DOI: 10.1039/c5ra19497k Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file.

PubMed Central

Zmijan, Robert; Jonnalagadda, Umesh S.; Carugo, Dario; Kochi, Yu; Lemm, Elizabeth; Packham, Graham; Hill, Martyn

2015-01-01

We demonstrate an imaging flow cytometer that uses acoustic levitation to assemble cells and other particles into a sheet structure. This technique enables a high resolution, low noise CMOS camera to capture images of thousands of cells with each frame. While ultrasonic focussing has previously been demonstrated for 1D cytometry systems, extending the technology to a planar, much higher throughput format and integrating imaging is non-trivial, and represents a significant jump forward in capability, leading to diagnostic possibilities not achievable with current systems. A galvo mirror is used to track the images of the moving cells permitting exposure times of 10 ms at frame rates of 50 fps with motion blur of only a few pixels. At 80 fps, we demonstrate a throughput of 208 000 beads per second. We investigate the factors affecting motion blur and throughput, and demonstrate the system with fluorescent beads, leukaemia cells and a chondrocyte cell line. Cells require more time to reach the acoustic focus than beads, resulting in lower throughputs; however a longer device would remove this constraint. PMID:29456838

Optimization of wide-area ATM and local-area ethernet/FDDI network configurations for high-speed telemedicine communications employing NASA's ACTS.

PubMed

McDermott, W R; Tri, J L; Mitchell, M P; Levens, S P; Wondrow, M A; Huie, L M; Khandheria, B K; Gilbert, B K

1999-01-01

A high data rate terrestrial and satellite network was implemented to transfer medical images and data. This article describes the a optimization of the workstations and switching equipment incorporated into the network. Topics discussed in this article include tuning of the network software, the configuration of the Sun Microsystems workstations, the FORE Systems asynchronous transfer mode switches, as well as the throughput results of two telemedicine experiments undertaken by Mayo's physician staff. The technical staff was successful in achieving the data throughput needed by the telemedicine software; particularly important was the proper determination of peak throughput and TCP window sizes to ensure optimum use of the resources available on the Sun Microsystems and Hewlett Packard workstations.

Optimized Negative Staining: a High-throughput Protocol for Examining Small and Asymmetric Protein Structure by Electron Microscopy

DOE PAGES

Rames, Matthew; Yu, Yadong; Ren, Gang

2014-08-15

Structural determination of proteins is rather challenging for proteins with molecular masses between 40 - 200 kDa. Considering that more than half of natural proteins have a molecular mass between 40 - 200 kDa, a robust and high-throughput method with a nanometer resolution capability is needed. Negative staining (NS) electron microscopy (EM) is an easy, rapid, and qualitative approach which has frequently been used in research laboratories to examine protein structure and protein-protein interactions. Unfortunately, conventional NS protocols often generate structural artifacts on proteins, especially with lipoproteins that usually form presenting rouleaux artifacts. By using images of lipoproteins from cryo-electronmore » microscopy (cryo-EM) as a standard, the key parameters in NS specimen preparation conditions were recently screened and reported as the optimized NS protocol (OpNS), a modified conventional NS protocol. Artifacts like rouleaux can be greatly limited by OpNS, additionally providing high contrast along with reasonably high-resolution (near 1 nm) images of small and asymmetric proteins. These high-resolution and high contrast images are even favorable for an individual protein (a single object, no average) 3D reconstruction, such as a 160 kDa antibody, through the method of electron tomography. Moreover, OpNS can be a high-throughput tool to examine hundreds of samples of small proteins. For example, the previously published mechanism of 53 kDa cholesteryl ester transfer protein (CETP) involved the screening and imaging of hundreds of samples. Considering cryo-EM rarely successfully images proteins less than 200 kDa has yet to publish any study involving screening over one hundred sample conditions, it is fair to call OpNS a high-throughput method for studying small proteins. Hopefully the OpNS protocol presented here can be a useful tool to push the boundaries of EM and accelerate EM studies into small protein structure, dynamics and mechanisms.« less

Image Harvest: an open-source platform for high-throughput plant image processing and analysis

PubMed Central

Knecht, Avi C.; Campbell, Malachy T.; Caprez, Adam; Swanson, David R.; Walia, Harkamal

2016-01-01

High-throughput plant phenotyping is an effective approach to bridge the genotype-to-phenotype gap in crops. Phenomics experiments typically result in large-scale image datasets, which are not amenable for processing on desktop computers, thus creating a bottleneck in the image-analysis pipeline. Here, we present an open-source, flexible image-analysis framework, called Image Harvest (IH), for processing images originating from high-throughput plant phenotyping platforms. Image Harvest is developed to perform parallel processing on computing grids and provides an integrated feature for metadata extraction from large-scale file organization. Moreover, the integration of IH with the Open Science Grid provides academic researchers with the computational resources required for processing large image datasets at no cost. Image Harvest also offers functionalities to extract digital traits from images to interpret plant architecture-related characteristics. To demonstrate the applications of these digital traits, a rice (Oryza sativa) diversity panel was phenotyped and genome-wide association mapping was performed using digital traits that are used to describe different plant ideotypes. Three major quantitative trait loci were identified on rice chromosomes 4 and 6, which co-localize with quantitative trait loci known to regulate agronomically important traits in rice. Image Harvest is an open-source software for high-throughput image processing that requires a minimal learning curve for plant biologists to analyzephenomics datasets. PMID:27141917

High-throughput dual-colour precision imaging for brain-wide connectome with cytoarchitectonic landmarks at the cellular level

PubMed Central

Gong, Hui; Xu, Dongli; Yuan, Jing; Li, Xiangning; Guo, Congdi; Peng, Jie; Li, Yuxin; Schwarz, Lindsay A.; Li, Anan; Hu, Bihe; Xiong, Benyi; Sun, Qingtao; Zhang, Yalun; Liu, Jiepeng; Zhong, Qiuyuan; Xu, Tonghui; Zeng, Shaoqun; Luo, Qingming

2016-01-01

The precise annotation and accurate identification of neural structures are prerequisites for studying mammalian brain function. The orientation of neurons and neural circuits is usually determined by mapping brain images to coarse axial-sampling planar reference atlases. However, individual differences at the cellular level likely lead to position errors and an inability to orient neural projections at single-cell resolution. Here, we present a high-throughput precision imaging method that can acquire a co-localized brain-wide data set of both fluorescent-labelled neurons and counterstained cell bodies at a voxel size of 0.32 × 0.32 × 2.0 μm in 3 days for a single mouse brain. We acquire mouse whole-brain imaging data sets of multiple types of neurons and projections with anatomical annotation at single-neuron resolution. The results show that the simultaneous acquisition of labelled neural structures and cytoarchitecture reference in the same brain greatly facilitates precise tracing of long-range projections and accurate locating of nuclei. PMID:27374071

G protein-coupled receptor internalization assays in the high-content screening format.

PubMed

Haasen, Dorothea; Schnapp, Andreas; Valler, Martin J; Heilker, Ralf

2006-01-01

High-content screening (HCS), a combination of fluorescence microscopic imaging and automated image analysis, has become a frequently applied tool to study test compound effects in cellular disease-modeling systems. This chapter describes the measurement of G protein-coupled receptor (GPCR) internalization in the HCS format using a high-throughput, confocal cellular imaging device. GPCRs are the most successful group of therapeutic targets on the pharmaceutical market. Accordingly, the search for compounds that interfere with GPCR function in a specific and selective way is a major focus of the pharmaceutical industry today. This chapter describes methods for the ligand-induced internalization of GPCRs labeled previously with either a fluorophore-conjugated ligand or an antibody directed against an N-terminal tag of the GPCR. Both labeling techniques produce robust assay formats. Complementary to other functional GPCR drug discovery assays, internalization assays enable a pharmacological analysis of test compounds. We conclude that GPCR internalization assays represent a valuable medium/high-throughput screening format to determine the cellular activity of GPCR ligands.

Handheld Fluorescence Microscopy based Flow Analyzer.

PubMed

Saxena, Manish; Jayakumar, Nitin; Gorthi, Sai Siva

2016-03-01

Fluorescence microscopy has the intrinsic advantages of favourable contrast characteristics and high degree of specificity. Consequently, it has been a mainstay in modern biological inquiry and clinical diagnostics. Despite its reliable nature, fluorescence based clinical microscopy and diagnostics is a manual, labour intensive and time consuming procedure. The article outlines a cost-effective, high throughput alternative to conventional fluorescence imaging techniques. With system level integration of custom-designed microfluidics and optics, we demonstrate fluorescence microscopy based imaging flow analyzer. Using this system we have imaged more than 2900 FITC labeled fluorescent beads per minute. This demonstrates high-throughput characteristics of our flow analyzer in comparison to conventional fluorescence microscopy. The issue of motion blur at high flow rates limits the achievable throughput in image based flow analyzers. Here we address the issue by computationally deblurring the images and show that this restores the morphological features otherwise affected by motion blur. By further optimizing concentration of the sample solution and flow speeds, along with imaging multiple channels simultaneously, the system is capable of providing throughput of about 480 beads per second.

High-throughput imaging of heterogeneous cell organelles with an X-ray laser (CXIDB ID 25)

DOE Data Explorer

Hantke, Max, F.

2014-11-17

Preprocessed detector images that were used for the paper "High-throughput imaging of heterogeneous cell organelles with an X-ray laser". The CXI file contains the entire recorded data - including both hits and blanks. It also includes down-sampled images and LCLS machine parameters. Additionally, the Cheetah configuration file is attached that was used to create the pre-processed data.

Quantifying the Onset and Progression of Plant Senescence by Color Image Analysis for High Throughput Applications

PubMed Central

Cai, Jinhai; Okamoto, Mamoru; Atieno, Judith; Sutton, Tim; Li, Yongle; Miklavcic, Stanley J.

2016-01-01

Leaf senescence, an indicator of plant age and ill health, is an important phenotypic trait for the assessment of a plant’s response to stress. Manual inspection of senescence, however, is time consuming, inaccurate and subjective. In this paper we propose an objective evaluation of plant senescence by color image analysis for use in a high throughput plant phenotyping pipeline. As high throughput phenotyping platforms are designed to capture whole-of-plant features, camera lenses and camera settings are inappropriate for the capture of fine detail. Specifically, plant colors in images may not represent true plant colors, leading to errors in senescence estimation. Our algorithm features a color distortion correction and image restoration step prior to a senescence analysis. We apply our algorithm to two time series of images of wheat and chickpea plants to quantify the onset and progression of senescence. We compare our results with senescence scores resulting from manual inspection. We demonstrate that our procedure is able to process images in an automated way for an accurate estimation of plant senescence even from color distorted and blurred images obtained under high throughput conditions. PMID:27348807

A high performance hardware implementation image encryption with AES algorithm

NASA Astrophysics Data System (ADS)

Farmani, Ali; Jafari, Mohamad; Miremadi, Seyed Sohrab

2011-06-01

This paper describes implementation of a high-speed encryption algorithm with high throughput for encrypting the image. Therefore, we select a highly secured symmetric key encryption algorithm AES(Advanced Encryption Standard), in order to increase the speed and throughput using pipeline technique in four stages, control unit based on logic gates, optimal design of multiplier blocks in mixcolumn phase and simultaneous production keys and rounds. Such procedure makes AES suitable for fast image encryption. Implementation of a 128-bit AES on FPGA of Altra company has been done and the results are as follow: throughput, 6 Gbps in 471MHz. The time of encrypting in tested image with 32*32 size is 1.15ms.

Affordable Imaging Lab for Noninvasive Analysis of Biomass and Early Vigour in Cereal Crops

PubMed Central

2018-01-01

Plant phenotyping by imaging allows automated analysis of plants for various morphological and physiological traits. In this work, we developed a low-cost RGB imaging phenotyping lab (LCP lab) for low-throughput imaging and analysis using affordable imaging equipment and freely available software. LCP lab comprising RGB imaging and analysis pipeline is set up and demonstrated with early vigour analysis in wheat. Using this lab, a few hundred pots can be photographed in a day and the pots are tracked with QR codes. The software pipeline for both imaging and analysis is built from freely available software. The LCP lab was evaluated for early vigour analysis of five wheat cultivars. A high coefficient of determination (R2 0.94) was obtained between the dry weight and the projected leaf area of 20-day-old wheat plants and R2 of 0.9 for the relative growth rate between 10 and 20 days of plant growth. Detailed description for setting up such a lab is provided together with custom scripts built for imaging and analysis. The LCP lab is an affordable alternative for analysis of cereal crops when access to a high-throughput phenotyping facility is unavailable or when the experiments require growing plants in highly controlled climate chambers. The protocols described in this work are useful for building affordable imaging system for small-scale research projects and for education. PMID:29850536

Image Harvest: an open-source platform for high-throughput plant image processing and analysis.

PubMed

Knecht, Avi C; Campbell, Malachy T; Caprez, Adam; Swanson, David R; Walia, Harkamal

2016-05-01

High-throughput plant phenotyping is an effective approach to bridge the genotype-to-phenotype gap in crops. Phenomics experiments typically result in large-scale image datasets, which are not amenable for processing on desktop computers, thus creating a bottleneck in the image-analysis pipeline. Here, we present an open-source, flexible image-analysis framework, called Image Harvest (IH), for processing images originating from high-throughput plant phenotyping platforms. Image Harvest is developed to perform parallel processing on computing grids and provides an integrated feature for metadata extraction from large-scale file organization. Moreover, the integration of IH with the Open Science Grid provides academic researchers with the computational resources required for processing large image datasets at no cost. Image Harvest also offers functionalities to extract digital traits from images to interpret plant architecture-related characteristics. To demonstrate the applications of these digital traits, a rice (Oryza sativa) diversity panel was phenotyped and genome-wide association mapping was performed using digital traits that are used to describe different plant ideotypes. Three major quantitative trait loci were identified on rice chromosomes 4 and 6, which co-localize with quantitative trait loci known to regulate agronomically important traits in rice. Image Harvest is an open-source software for high-throughput image processing that requires a minimal learning curve for plant biologists to analyzephenomics datasets. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

High-throughput image analysis of tumor spheroids: a user-friendly software application to measure the size of spheroids automatically and accurately.

PubMed

Chen, Wenjin; Wong, Chung; Vosburgh, Evan; Levine, Arnold J; Foran, David J; Xu, Eugenia Y

2014-07-08

The increasing number of applications of three-dimensional (3D) tumor spheroids as an in vitro model for drug discovery requires their adaptation to large-scale screening formats in every step of a drug screen, including large-scale image analysis. Currently there is no ready-to-use and free image analysis software to meet this large-scale format. Most existing methods involve manually drawing the length and width of the imaged 3D spheroids, which is a tedious and time-consuming process. This study presents a high-throughput image analysis software application - SpheroidSizer, which measures the major and minor axial length of the imaged 3D tumor spheroids automatically and accurately; calculates the volume of each individual 3D tumor spheroid; then outputs the results in two different forms in spreadsheets for easy manipulations in the subsequent data analysis. The main advantage of this software is its powerful image analysis application that is adapted for large numbers of images. It provides high-throughput computation and quality-control workflow. The estimated time to process 1,000 images is about 15 min on a minimally configured laptop, or around 1 min on a multi-core performance workstation. The graphical user interface (GUI) is also designed for easy quality control, and users can manually override the computer results. The key method used in this software is adapted from the active contour algorithm, also known as Snakes, which is especially suitable for images with uneven illumination and noisy background that often plagues automated imaging processing in high-throughput screens. The complimentary "Manual Initialize" and "Hand Draw" tools provide the flexibility to SpheroidSizer in dealing with various types of spheroids and diverse quality images. This high-throughput image analysis software remarkably reduces labor and speeds up the analysis process. Implementing this software is beneficial for 3D tumor spheroids to become a routine in vitro model for drug screens in industry and academia.

Asymmetric-detection time-stretch optical microscopy (ATOM) for ultrafast high-contrast cellular imaging in flow

PubMed Central

Wong, Terence T. W.; Lau, Andy K. S.; Ho, Kenneth K. Y.; Tang, Matthew Y. H.; Robles, Joseph D. F.; Wei, Xiaoming; Chan, Antony C. S.; Tang, Anson H. L.; Lam, Edmund Y.; Wong, Kenneth K. Y.; Chan, Godfrey C. F.; Shum, Ho Cheung; Tsia, Kevin K.

2014-01-01

Accelerating imaging speed in optical microscopy is often realized at the expense of image contrast, image resolution, and detection sensitivity – a common predicament for advancing high-speed and high-throughput cellular imaging. We here demonstrate a new imaging approach, called asymmetric-detection time-stretch optical microscopy (ATOM), which can deliver ultrafast label-free high-contrast flow imaging with well delineated cellular morphological resolution and in-line optical image amplification to overcome the compromised imaging sensitivity at high speed. We show that ATOM can separately reveal the enhanced phase-gradient and absorption contrast in microfluidic live-cell imaging at a flow speed as high as ~10 m/s, corresponding to an imaging throughput of ~100,000 cells/sec. ATOM could thus be the enabling platform to meet the pressing need for intercalating optical microscopy in cellular assay, e.g. imaging flow cytometry – permitting high-throughput access to the morphological information of the individual cells simultaneously with a multitude of parameters obtained in the standard assay. PMID:24413677

A Dual-Mode Large-Arrayed CMOS ISFET Sensor for Accurate and High-Throughput pH Sensing in Biomedical Diagnosis.

PubMed

Huang, Xiwei; Yu, Hao; Liu, Xu; Jiang, Yu; Yan, Mei; Wu, Dongping

2015-09-01

The existing ISFET-based DNA sequencing detects hydrogen ions released during the polymerization of DNA strands on microbeads, which are scattered into microwell array above the ISFET sensor with unknown distribution. However, false pH detection happens at empty microwells due to crosstalk from neighboring microbeads. In this paper, a dual-mode CMOS ISFET sensor is proposed to have accurate pH detection toward DNA sequencing. Dual-mode sensing, optical and chemical modes, is realized by integrating a CMOS image sensor (CIS) with ISFET pH sensor, and is fabricated in a standard 0.18-μm CIS process. With accurate determination of microbead physical locations with CIS pixel by contact imaging, the dual-mode sensor can correlate local pH for one DNA slice at one location-determined microbead, which can result in improved pH detection accuracy. Moreover, toward a high-throughput DNA sequencing, a correlated-double-sampling readout that supports large array for both modes is deployed to reduce pixel-to-pixel nonuniformity such as threshold voltage mismatch. The proposed CMOS dual-mode sensor is experimentally examined to show a well correlated pH map and optical image for microbeads with a pH sensitivity of 26.2 mV/pH, a fixed pattern noise (FPN) reduction from 4% to 0.3%, and a readout speed of 1200 frames/s. A dual-mode CMOS ISFET sensor with suppressed FPN for accurate large-arrayed pH sensing is proposed and demonstrated with state-of-the-art measured results toward accurate and high-throughput DNA sequencing. The developed dual-mode CMOS ISFET sensor has great potential for future personal genome diagnostics with high accuracy and low cost.

Arraycount, an algorithm for automatic cell counting in microwell arrays.

PubMed

Kachouie, Nezamoddin; Kang, Lifeng; Khademhosseini, Ali

2009-09-01

Microscale technologies have emerged as a powerful tool for studying and manipulating biological systems and miniaturizing experiments. However, the lack of software complementing these techniques has made it difficult to apply them for many high-throughput experiments. This work establishes Arraycount, an approach to automatically count cells in microwell arrays. The procedure consists of fluorescent microscope imaging of cells that are seeded in microwells of a microarray system and then analyzing images via computer to recognize the array and count cells inside each microwell. To start counting, green and red fluorescent images (representing live and dead cells, respectively) are extracted from the original image and processed separately. A template-matching algorithm is proposed in which pre-defined well and cell templates are matched against the red and green images to locate microwells and cells. Subsequently, local maxima in the correlation maps are determined and local maxima maps are thresholded. At the end, the software records the cell counts for each detected microwell on the original image in high-throughput. The automated counting was shown to be accurate compared with manual counting, with a difference of approximately 1-2 cells per microwell: based on cell concentration, the absolute difference between manual and automatic counting measurements was 2.5-13%.

High-Throughput Particle Uptake Analysis by Imaging Flow Cytometry

PubMed Central

Smirnov, Asya; Solga, Michael D.; Lannigan, Joanne; Criss, Alison K.

2017-01-01

Quantifying the efficiency of particle uptake by host cells is important in fields including infectious diseases, autoimmunity, cancer, developmental biology, and drug delivery. Here we present a protocol for high-throughput analysis of particle uptake using imaging flow cytometry, using the bacterium Neisseria gonorrhoeae attached and internalized to neutrophils as an example. Cells are exposed to fluorescently labeled bacteria, fixed, and stained with a bacteria-specific antibody of a different fluorophore. Thus in the absence of a permeabilizing agent, extracellular bacteria are double-labeled with two fluorophores while intracellular bacteria remain single-labeled. A spot count algorithm is used to determine the number of single- and double-labeled bacteria in individual cells, to calculate the percent of cells associated with bacteria, percent of cells with internalized bacteria, and percent of cell-associated bacteria that are internalized. These analyses quantify bacterial association and internalization across thousands of cells and can be applied to diverse experimental systems. PMID:28369762

Detecting adulterants in milk powder using high-throughput Raman chemical imaging

USDA-ARS?s Scientific Manuscript database

This study used a line-scan high-throughput Raman imaging system to authenticate milk powder. A 5 W 785 nm line laser (240 mm long and 1 mm wide) was used as a Raman excitation source. The system was used to acquire hyperspectral Raman images in a wavenumber range of 103–2881 cm-1 from the skim milk...

Lens-free shadow image based high-throughput continuous cell monitoring technique.

PubMed

Jin, Geonsoo; Yoo, In-Hwa; Pack, Seung Pil; Yang, Ji-Woon; Ha, Un-Hwan; Paek, Se-Hwan; Seo, Sungkyu

2012-01-01

A high-throughput continuous cell monitoring technique which does not require any labeling reagents or destruction of the specimen is demonstrated. More than 6000 human alveolar epithelial A549 cells are monitored for up to 72 h simultaneously and continuously with a single digital image within a cost and space effective lens-free shadow imaging platform. In an experiment performed within a custom built incubator integrated with the lens-free shadow imaging platform, the cell nucleus division process could be successfully characterized by calculating the signal-to-noise ratios (SNRs) and the shadow diameters (SDs) of the cell shadow patterns. The versatile nature of this platform also enabled a single cell viability test followed by live cell counting. This study firstly shows that the lens-free shadow imaging technique can provide a continuous cell monitoring without any staining/labeling reagent and destruction of the specimen. This high-throughput continuous cell monitoring technique based on lens-free shadow imaging may be widely utilized as a compact, low-cost, and high-throughput cell monitoring tool in the fields of drug and food screening or cell proliferation and viability testing. Copyright © 2012 Elsevier B.V. All rights reserved.

High-Throughput Method for Automated Colony and Cell Counting by Digital Image Analysis Based on Edge Detection

PubMed Central

Choudhry, Priya

2016-01-01

Counting cells and colonies is an integral part of high-throughput screens and quantitative cellular assays. Due to its subjective and time-intensive nature, manual counting has hindered the adoption of cellular assays such as tumor spheroid formation in high-throughput screens. The objective of this study was to develop an automated method for quick and reliable counting of cells and colonies from digital images. For this purpose, I developed an ImageJ macro Cell Colony Edge and a CellProfiler Pipeline Cell Colony Counting, and compared them to other open-source digital methods and manual counts. The ImageJ macro Cell Colony Edge is valuable in counting cells and colonies, and measuring their area, volume, morphology, and intensity. In this study, I demonstrate that Cell Colony Edge is superior to other open-source methods, in speed, accuracy and applicability to diverse cellular assays. It can fulfill the need to automate colony/cell counting in high-throughput screens, colony forming assays, and cellular assays. PMID:26848849

Present and future of membrane protein structure determination by electron crystallography.

PubMed

Ubarretxena-Belandia, Iban; Stokes, David L

2010-01-01

Membrane proteins are critical to cell physiology, playing roles in signaling, trafficking, transport, adhesion, and recognition. Despite their relative abundance in the proteome and their prevalence as targets of therapeutic drugs, structural information about membrane proteins is in short supply. This chapter describes the use of electron crystallography as a tool for determining membrane protein structures. Electron crystallography offers distinct advantages relative to the alternatives of X-ray crystallography and NMR spectroscopy. Namely, membrane proteins are placed in their native membranous environment, which is likely to favor a native conformation and allow changes in conformation in response to physiological ligands. Nevertheless, there are significant logistical challenges in finding appropriate conditions for inducing membrane proteins to form two-dimensional arrays within the membrane and in using electron cryo-microscopy to collect the data required for structure determination. A number of developments are described for high-throughput screening of crystallization trials and for automated imaging of crystals with the electron microscope. These tools are critical for exploring the necessary range of factors governing the crystallization process. There have also been recent software developments to facilitate the process of structure determination. However, further innovations in the algorithms used for processing images and electron diffraction are necessary to improve throughput and to make electron crystallography truly viable as a method for determining atomic structures of membrane proteins. Copyright © 2010 Elsevier Inc. All rights reserved.

Present and future of membrane protein structure determination by electron crystallography

PubMed Central

Ubarretxena-Belandia, Iban; Stokes, David L.

2011-01-01

Membrane proteins are critical to cell physiology, playing roles in signaling, trafficking, transport, adhesion, and recognition. Despite their relative abundance in the proteome and their prevalence as targets of therapeutic drugs, structural information about membrane proteins is in short supply. This review describes the use of electron crystallography as a tool for determining membrane protein structures. Electron crystallography offers distinct advantages relative to the alternatives of X-ray crystallography and NMR spectroscopy. Namely, membrane proteins are placed in their native membranous environment, which is likely to favor a native conformation and allow changes in conformation in response to physiological ligands. Nevertheless, there are significant logistical challenges in finding appropriate conditions for inducing membrane proteins to form two-dimensional arrays within the membrane and in using electron cryo-microscopy to collect the data required for structure determination. A number of developments are described for high-throughput screening of crystallization trials and for automated imaging of crystals with the electron microscope. These tools are critical for exploring the necessary range of factors governing the crystallization process. There have also been recent software developments to facilitate the process of structure determination. However, further innovations in the algorithms used for processing images and electron diffraction are necessary to improve throughput and to make electron crystallography truly viable as a method for determining atomic structures of membrane proteins. PMID:21115172

Subnuclear foci quantification using high-throughput 3D image cytometry

NASA Astrophysics Data System (ADS)

Wadduwage, Dushan N.; Parrish, Marcus; Choi, Heejin; Engelward, Bevin P.; Matsudaira, Paul; So, Peter T. C.

2015-07-01

Ionising radiation causes various types of DNA damages including double strand breaks (DSBs). DSBs are often recognized by DNA repair protein ATM which forms gamma-H2AX foci at the site of the DSBs that can be visualized using immunohistochemistry. However most of such experiments are of low throughput in terms of imaging and image analysis techniques. Most of the studies still use manual counting or classification. Hence they are limited to counting a low number of foci per cell (5 foci per nucleus) as the quantification process is extremely labour intensive. Therefore we have developed a high throughput instrumentation and computational pipeline specialized for gamma-H2AX foci quantification. A population of cells with highly clustered foci inside nuclei were imaged, in 3D with submicron resolution, using an in-house developed high throughput image cytometer. Imaging speeds as high as 800 cells/second in 3D were achieved by using HiLo wide-field depth resolved imaging and a remote z-scanning technique. Then the number of foci per cell nucleus were quantified using a 3D extended maxima transform based algorithm. Our results suggests that while most of the other 2D imaging and manual quantification studies can count only up to about 5 foci per nucleus our method is capable of counting more than 100. Moreover we show that 3D analysis is significantly superior compared to the 2D techniques.

Super-resolution Imaging of Chemical Synapses in the Brain

PubMed Central

Dani, Adish; Huang, Bo; Bergan, Joseph; Dulac, Catherine; Zhuang, Xiaowei

2010-01-01

Determination of the molecular architecture of synapses requires nanoscopic image resolution and specific molecular recognition, a task that has so far defied many conventional imaging approaches. Here we present a super-resolution fluorescence imaging method to visualize the molecular architecture of synapses in the brain. Using multicolor, three-dimensional stochastic optical reconstruction microscopy, the distributions of synaptic proteins can be measured with nanometer precision. Furthermore, the wide-field, volumetric imaging method enables high-throughput, quantitative analysis of a large number of synapses from different brain regions. To demonstrate the capabilities of this approach, we have determined the organization of ten protein components of the presynaptic active zone and the postsynaptic density. Variations in synapse morphology, neurotransmitter receptor composition, and receptor distribution were observed both among synapses and across different brain regions. Combination with optogenetics further allowed molecular events associated with synaptic plasticity to be resolved at the single-synapse level. PMID:21144999

Optical Layout Analysis of Polarization Interference Imaging Spectrometer by Jones Calculus in View of both Optical Throughput and Interference Fringe Visibility

NASA Astrophysics Data System (ADS)

Zhang, Xuanni; Zhang, Chunmin

2013-01-01

A polarization interference imaging spectrometer based on Savart polariscope was presented. Its optical throughput was analyzed by Jones calculus. The throughput expression was given, and clearly showed that the optical throughput mainly depended on the intensity of incident light, transmissivity, refractive index and the layout of optical system. The simulation and analysis gave the optimum layout in view of both optical throughput and interference fringe visibility, and verified that the layout of our former design was optimum. The simulation showed that a small deviation from the optimum layout influenced interference fringe visibility little for the optimum one, but influenced severely for others, so a small deviation is admissible in the optimum, and this can mitigate the manufacture difficulty. These results pave the way for further research and engineering design.

Label-free cell-cycle analysis by high-throughput quantitative phase time-stretch imaging flow cytometry

NASA Astrophysics Data System (ADS)

Mok, Aaron T. Y.; Lee, Kelvin C. M.; Wong, Kenneth K. Y.; Tsia, Kevin K.

2018-02-01

Biophysical properties of cells could complement and correlate biochemical markers to characterize a multitude of cellular states. Changes in cell size, dry mass and subcellular morphology, for instance, are relevant to cell-cycle progression which is prevalently evaluated by DNA-targeted fluorescence measurements. Quantitative-phase microscopy (QPM) is among the effective biophysical phenotyping tools that can quantify cell sizes and sub-cellular dry mass density distribution of single cells at high spatial resolution. However, limited camera frame rate and thus imaging throughput makes QPM incompatible with high-throughput flow cytometry - a gold standard in multiparametric cell-based assay. Here we present a high-throughput approach for label-free analysis of cell cycle based on quantitative-phase time-stretch imaging flow cytometry at a throughput of > 10,000 cells/s. Our time-stretch QPM system enables sub-cellular resolution even at high speed, allowing us to extract a multitude (at least 24) of single-cell biophysical phenotypes (from both amplitude and phase images). Those phenotypes can be combined to track cell-cycle progression based on a t-distributed stochastic neighbor embedding (t-SNE) algorithm. Using multivariate analysis of variance (MANOVA) discriminant analysis, cell-cycle phases can also be predicted label-free with high accuracy at >90% in G1 and G2 phase, and >80% in S phase. We anticipate that high throughput label-free cell cycle characterization could open new approaches for large-scale single-cell analysis, bringing new mechanistic insights into complex biological processes including diseases pathogenesis.

Frequency Based Design Partitioning to Achieve Higher Throughput in Digital Cross Correlator for Aperture Synthesis Passive MMW Imager.

PubMed

Asif, Muhammad; Guo, Xiangzhou; Zhang, Jing; Miao, Jungang

2018-04-17

Digital cross-correlation is central to many applications including but not limited to Digital Image Processing, Satellite Navigation and Remote Sensing. With recent advancements in digital technology, the computational demands of such applications have increased enormously. In this paper we are presenting a high throughput digital cross correlator, capable of processing 1-bit digitized stream, at the rate of up to 2 GHz, simultaneously on 64 channels i.e., approximately 4 Trillion correlation and accumulation operations per second. In order to achieve higher throughput, we have focused on frequency based partitioning of our design and tried to minimize and localize high frequency operations. This correlator is designed for a Passive Millimeter Wave Imager intended for the detection of contraband items concealed on human body. The goals are to increase the system bandwidth, achieve video rate imaging, improve sensitivity and reduce the size. Design methodology is detailed in subsequent sections, elaborating the techniques enabling high throughput. The design is verified for Xilinx Kintex UltraScale device in simulation and the implementation results are given in terms of device utilization and power consumption estimates. Our results show considerable improvements in throughput as compared to our baseline design, while the correlator successfully meets the functional requirements.

Strain Library Imaging Protocol for high-throughput, automated single-cell microscopy of large bacterial collections arrayed on multiwell plates.

PubMed

Shi, Handuo; Colavin, Alexandre; Lee, Timothy K; Huang, Kerwyn Casey

2017-02-01

Single-cell microscopy is a powerful tool for studying gene functions using strain libraries, but it suffers from throughput limitations. Here we describe the Strain Library Imaging Protocol (SLIP), which is a high-throughput, automated microscopy workflow for large strain collections that requires minimal user involvement. SLIP involves transferring arrayed bacterial cultures from multiwell plates onto large agar pads using inexpensive replicator pins and automatically imaging the resulting single cells. The acquired images are subsequently reviewed and analyzed by custom MATLAB scripts that segment single-cell contours and extract quantitative metrics. SLIP yields rich data sets on cell morphology and gene expression that illustrate the function of certain genes and the connections among strains in a library. For a library arrayed on 96-well plates, image acquisition can be completed within 4 min per plate.

Custom Super-Resolution Microscope for the Structural Analysis of Nanostructures

DTIC Science & Technology

2018-05-29

research community. As part of our validation of the new design approach, we performed two - color imaging of pairs of adjacent oligo probes hybridized...nanostructures and biological targets. Our microscope features a large field of view and custom optics that facilitate 3D imaging and enhanced contrast in...our imaging throughput by creating two microscopy platforms for high-throughput, super-resolution materials characterization, with the AO set-up being

High-Throughput Classification of Radiographs Using Deep Convolutional Neural Networks.

PubMed

Rajkomar, Alvin; Lingam, Sneha; Taylor, Andrew G; Blum, Michael; Mongan, John

2017-02-01

The study aimed to determine if computer vision techniques rooted in deep learning can use a small set of radiographs to perform clinically relevant image classification with high fidelity. One thousand eight hundred eighty-five chest radiographs on 909 patients obtained between January 2013 and July 2015 at our institution were retrieved and anonymized. The source images were manually annotated as frontal or lateral and randomly divided into training, validation, and test sets. Training and validation sets were augmented to over 150,000 images using standard image manipulations. We then pre-trained a series of deep convolutional networks based on the open-source GoogLeNet with various transformations of the open-source ImageNet (non-radiology) images. These trained networks were then fine-tuned using the original and augmented radiology images. The model with highest validation accuracy was applied to our institutional test set and a publicly available set. Accuracy was assessed by using the Youden Index to set a binary cutoff for frontal or lateral classification. This retrospective study was IRB approved prior to initiation. A network pre-trained on 1.2 million greyscale ImageNet images and fine-tuned on augmented radiographs was chosen. The binary classification method correctly classified 100 % (95 % CI 99.73-100 %) of both our test set and the publicly available images. Classification was rapid, at 38 images per second. A deep convolutional neural network created using non-radiological images, and an augmented set of radiographs is effective in highly accurate classification of chest radiograph view type and is a feasible, rapid method for high-throughput annotation.

Quasi-random array imaging collimator

DOEpatents

Fenimore, E.E.

1980-08-20

A hexagonally shaped quasi-random no-two-holes-touching imaging collimator. The quasi-random array imaging collimator eliminates contamination from small angle off-axis rays by using a no-two-holes-touching pattern which simultaneously provides for a self-supporting array increasing throughput by elimination of a substrate. The present invention also provides maximum throughput using hexagonally shaped holes in a hexagonal lattice pattern for diffraction limited applications. Mosaicking is also disclosed for reducing fabrication effort.

A high-throughput technique for determining grain boundary character non-destructively in microstructures with through-thickness grains

DOE PAGES

Seita, Matteo; Volpi, Marco; Patala, Srikanth; ...

2016-06-24

Grain boundaries (GBs) govern many properties of polycrystalline materials. However, because of their structural variability, our knowledge of GB constitutive relations is still very limited. We present a novel method to characterise the complete crystallography of individual GBs non-destructively, with high-throughput, and using commercially available tools. This method combines electron diffraction, optical reflectance and numerical image analysis to determine all five crystallographic parameters of numerous GBs in samples with through-thickness grains. We demonstrate the technique by measuring the crystallographic character of about 1,000 individual GBs in aluminum in a single run. Our method enables cost- and time-effective assembly of crystallography–propertymore » databases for thousands of individual GBs. Furthermore, such databases are essential for identifying GB constitutive relations and for predicting GB-related behaviours of polycrystalline solids.« less

High-Throughput Synthesis and Structure of Zeolite ZSM-43 with Two-Directional 8-Ring Channels.

PubMed

Willhammar, Tom; Su, Jie; Yun, Yifeng; Zou, Xiaodong; Afeworki, Mobae; Weston, Simon C; Vroman, Hilda B; Lonergan, William W; Strohmaier, Karl G

2017-08-07

The aluminosilicate zeolite ZSM-43 (where ZSM = Zeolite Socony Mobil) was first synthesized more than 3 decades ago, but its chemical structure remained unsolved because of its poor crystallinity and small crystal size. Here we present optimization of the ZSM-43 synthesis using a high-throughput approach and subsequent structure determination by the combination of electron crystallographic methods and powder X-ray diffraction. The synthesis required the use of a combination of both inorganic (Cs + and K + ) and organic (choline) structure-directing agents. High-throughput synthesis enabled a screening of the synthesis conditions, which made it possible to optimize the synthesis, despite its complexity, in order to obtain a material with significantly improved crystallinity. When both rotation electron diffraction and high-resolution transmission electron microscopy imaging techniques are applied, the structure of ZSM-43 could be determined. The structure of ZSM-43 is a new zeolite framework type and possesses a unique two-dimensional channel system limited by 8-ring channels. ZSM-43 is stable upon calcination, and sorption measurements show that the material is suitable for adsorption of carbon dioxide as well as methane.

Three applications of backscatter x-ray imaging technology to homeland defense

NASA Astrophysics Data System (ADS)

Chalmers, Alex

2005-05-01

A brief review of backscatter x-ray imaging and a description of three systems currently applying it to homeland defense missions (BodySearch, ZBV and ZBP). These missions include detection of concealed weapons, explosives and contraband on personnel, in vehicles and large cargo containers. An overview of the x-ray imaging subsystems is provided as well as sample images from each system. Key features such as x-ray safety, throughput and detection are discussed. Recent trends in operational modes are described that facilitate 100% inspection at high throughput chokepoints.

Automated image alignment for 2D gel electrophoresis in a high-throughput proteomics pipeline.

PubMed

Dowsey, Andrew W; Dunn, Michael J; Yang, Guang-Zhong

2008-04-01

The quest for high-throughput proteomics has revealed a number of challenges in recent years. Whilst substantial improvements in automated protein separation with liquid chromatography and mass spectrometry (LC/MS), aka 'shotgun' proteomics, have been achieved, large-scale open initiatives such as the Human Proteome Organization (HUPO) Brain Proteome Project have shown that maximal proteome coverage is only possible when LC/MS is complemented by 2D gel electrophoresis (2-DE) studies. Moreover, both separation methods require automated alignment and differential analysis to relieve the bioinformatics bottleneck and so make high-throughput protein biomarker discovery a reality. The purpose of this article is to describe a fully automatic image alignment framework for the integration of 2-DE into a high-throughput differential expression proteomics pipeline. The proposed method is based on robust automated image normalization (RAIN) to circumvent the drawbacks of traditional approaches. These use symbolic representation at the very early stages of the analysis, which introduces persistent errors due to inaccuracies in modelling and alignment. In RAIN, a third-order volume-invariant B-spline model is incorporated into a multi-resolution schema to correct for geometric and expression inhomogeneity at multiple scales. The normalized images can then be compared directly in the image domain for quantitative differential analysis. Through evaluation against an existing state-of-the-art method on real and synthetically warped 2D gels, the proposed analysis framework demonstrates substantial improvements in matching accuracy and differential sensitivity. High-throughput analysis is established through an accelerated GPGPU (general purpose computation on graphics cards) implementation. Supplementary material, software and images used in the validation are available at http://www.proteomegrid.org/rain/.

SPIM-fluid: open source light-sheet based platform for high-throughput imaging

PubMed Central

Gualda, Emilio J.; Pereira, Hugo; Vale, Tiago; Estrada, Marta Falcão; Brito, Catarina; Moreno, Nuno

2015-01-01

Light sheet fluorescence microscopy has recently emerged as the technique of choice for obtaining high quality 3D images of whole organisms/embryos with low photodamage and fast acquisition rates. Here we present an open source unified implementation based on Arduino and Micromanager, which is capable of operating Light Sheet Microscopes for automatized 3D high-throughput imaging on three-dimensional cell cultures and model organisms like zebrafish, oriented to massive drug screening. PMID:26601007

High throughput on-chip analysis of high-energy charged particle tracks using lensfree imaging

DOE Office of Scientific and Technical Information (OSTI.GOV)

Luo, Wei; Shabbir, Faizan; Gong, Chao

2015-04-13

We demonstrate a high-throughput charged particle analysis platform, which is based on lensfree on-chip microscopy for rapid ion track analysis using allyl diglycol carbonate, i.e., CR-39 plastic polymer as the sensing medium. By adopting a wide-area opto-electronic image sensor together with a source-shifting based pixel super-resolution technique, a large CR-39 sample volume (i.e., 4 cm × 4 cm × 0.1 cm) can be imaged in less than 1 min using a compact lensfree on-chip microscope, which detects partially coherent in-line holograms of the ion tracks recorded within the CR-39 detector. After the image capture, using highly parallelized reconstruction and ion track analysis algorithms running on graphics processingmore » units, we reconstruct and analyze the entire volume of a CR-39 detector within ∼1.5 min. This significant reduction in the entire imaging and ion track analysis time not only increases our throughput but also allows us to perform time-resolved analysis of the etching process to monitor and optimize the growth of ion tracks during etching. This computational lensfree imaging platform can provide a much higher throughput and more cost-effective alternative to traditional lens-based scanning optical microscopes for ion track analysis using CR-39 and other passive high energy particle detectors.« less

A space- and time-resolved single photon counting detector for fluorescence microscopy and spectroscopy

PubMed Central

Michalet, X.; Siegmund, O.H.W.; Vallerga, J.V.; Jelinsky, P.; Millaud, J.E.; Weiss, S.

2017-01-01

We have recently developed a wide-field photon-counting detector having high-temporal and high-spatial resolutions and capable of high-throughput (the H33D detector). Its design is based on a 25 mm diameter multi-alkali photocathode producing one photo electron per detected photon, which are then multiplied up to 107 times by a 3-microchannel plate stack. The resulting electron cloud is proximity focused on a cross delay line anode, which allows determining the incident photon position with high accuracy. The imaging and fluorescence lifetime measurement performances of the H33D detector installed on a standard epifluorescence microscope will be presented. We compare them to those of standard single-molecule detectors such as single-photon avalanche photodiode (SPAD) or electron-multiplying camera using model samples (fluorescent beads, quantum dots and live cells). Finally, we discuss the design and applications of future generation of H33D detectors for single-molecule imaging and high-throughput study of biomolecular interactions. PMID:29479130

Spotsizer: High-throughput quantitative analysis of microbial growth.

PubMed

Bischof, Leanne; Převorovský, Martin; Rallis, Charalampos; Jeffares, Daniel C; Arzhaeva, Yulia; Bähler, Jürg

2016-10-01

Microbial colony growth can serve as a useful readout in assays for studying complex genetic interactions or the effects of chemical compounds. Although computational tools for acquiring quantitative measurements of microbial colonies have been developed, their utility can be compromised by inflexible input image requirements, non-trivial installation procedures, or complicated operation. Here, we present the Spotsizer software tool for automated colony size measurements in images of robotically arrayed microbial colonies. Spotsizer features a convenient graphical user interface (GUI), has both single-image and batch-processing capabilities, and works with multiple input image formats and different colony grid types. We demonstrate how Spotsizer can be used for high-throughput quantitative analysis of fission yeast growth. The user-friendly Spotsizer tool provides rapid, accurate, and robust quantitative analyses of microbial growth in a high-throughput format. Spotsizer is freely available at https://data.csiro.au/dap/landingpage?pid=csiro:15330 under a proprietary CSIRO license.

A Near-Optimal Distributed QoS Constrained Routing Algorithm for Multichannel Wireless Sensor Networks

PubMed Central

Lin, Frank Yeong-Sung; Hsiao, Chiu-Han; Yen, Hong-Hsu; Hsieh, Yu-Jen

2013-01-01

One of the important applications in Wireless Sensor Networks (WSNs) is video surveillance that includes the tasks of video data processing and transmission. Processing and transmission of image and video data in WSNs has attracted a lot of attention in recent years. This is known as Wireless Visual Sensor Networks (WVSNs). WVSNs are distributed intelligent systems for collecting image or video data with unique performance, complexity, and quality of service challenges. WVSNs consist of a large number of battery-powered and resource constrained camera nodes. End-to-end delay is a very important Quality of Service (QoS) metric for video surveillance application in WVSNs. How to meet the stringent delay QoS in resource constrained WVSNs is a challenging issue that requires novel distributed and collaborative routing strategies. This paper proposes a Near-Optimal Distributed QoS Constrained (NODQC) routing algorithm to achieve an end-to-end route with lower delay and higher throughput. A Lagrangian Relaxation (LR)-based routing metric that considers the “system perspective” and “user perspective” is proposed to determine the near-optimal routing paths that satisfy end-to-end delay constraints with high system throughput. The empirical results show that the NODQC routing algorithm outperforms others in terms of higher system throughput with lower average end-to-end delay and delay jitter. In this paper, for the first time, the algorithm shows how to meet the delay QoS and at the same time how to achieve higher system throughput in stringently resource constrained WVSNs.

Multi-MHz laser-scanning single-cell fluorescence microscopy by spatiotemporally encoded virtual source array

PubMed Central

Wu, Jianglai; Tang, Anson H. L.; Mok, Aaron T. Y.; Yan, Wenwei; Chan, Godfrey C. F.; Wong, Kenneth K. Y.; Tsia, Kevin K.

2017-01-01

Apart from the spatial resolution enhancement, scaling of temporal resolution, equivalently the imaging throughput, of fluorescence microscopy is of equal importance in advancing cell biology and clinical diagnostics. Yet, this attribute has mostly been overlooked because of the inherent speed limitation of existing imaging strategies. To address the challenge, we employ an all-optical laser-scanning mechanism, enabled by an array of reconfigurable spatiotemporally-encoded virtual sources, to demonstrate ultrafast fluorescence microscopy at line-scan rate as high as 8 MHz. We show that this technique enables high-throughput single-cell microfluidic fluorescence imaging at 75,000 cells/second and high-speed cellular 2D dynamical imaging at 3,000 frames per second, outperforming the state-of-the-art high-speed cameras and the gold-standard laser scanning strategies. Together with its wide compatibility to the existing imaging modalities, this technology could empower new forms of high-throughput and high-speed biological fluorescence microscopy that was once challenged. PMID:28966855

Detection and quantification of adulterants in milk powder using high-throughput Raman chemical imaging technique

USDA-ARS?s Scientific Manuscript database

Milk is a vulnerable target for economically motivated adulteration. In this study, a line-scan high-throughput Raman imaging system was used to authenticate milk powder. A 5 W 785 nm line laser (240 mm long and 1 mm wide) was used as a Raman excitation source. The system was used to acquire hypersp...

High-Throughput Light Sheet Microscopy for the Automated Live Imaging of Larval Zebrafish

NASA Astrophysics Data System (ADS)

Baker, Ryan; Logan, Savannah; Dudley, Christopher; Parthasarathy, Raghuveer

The zebrafish is a model organism with a variety of useful properties; it is small and optically transparent, it reproduces quickly, it is a vertebrate, and there are a large variety of transgenic animals available. Because of these properties, the zebrafish is well suited to study using a variety of optical technologies including light sheet fluorescence microscopy (LSFM), which provides high-resolution three-dimensional imaging over large fields of view. Research progress, however, is often not limited by optical techniques but instead by the number of samples one can examine over the course of an experiment, which in the case of light sheet imaging has so far been severely limited. Here we present an integrated fluidic circuit and microscope which provides rapid, automated imaging of zebrafish using several imaging modes, including LSFM, Hyperspectral Imaging, and Differential Interference Contrast Microscopy. Using this system, we show that we can increase our imaging throughput by a factor of 10 compared to previous techniques. We also show preliminary results visualizing zebrafish immune response, which is sensitive to gut microbiota composition, and which shows a strong variability between individuals that highlights the utility of high throughput imaging. National Science Foundation, Award No. DBI-1427957.

An Automated High-throughput Array Microscope for Cancer Cell Mechanics

NASA Astrophysics Data System (ADS)

Cribb, Jeremy A.; Osborne, Lukas D.; Beicker, Kellie; Psioda, Matthew; Chen, Jian; O'Brien, E. Timothy; Taylor, Russell M., II; Vicci, Leandra; Hsiao, Joe Ping-Lin; Shao, Chong; Falvo, Michael; Ibrahim, Joseph G.; Wood, Kris C.; Blobe, Gerard C.; Superfine, Richard

2016-06-01

Changes in cellular mechanical properties correlate with the progression of metastatic cancer along the epithelial-to-mesenchymal transition (EMT). Few high-throughput methodologies exist that measure cell compliance, which can be used to understand the impact of genetic alterations or to screen the efficacy of chemotherapeutic agents. We have developed a novel array high-throughput microscope (AHTM) system that combines the convenience of the standard 96-well plate with the ability to image cultured cells and membrane-bound microbeads in twelve independently-focusing channels simultaneously, visiting all wells in eight steps. We use the AHTM and passive bead rheology techniques to determine the relative compliance of human pancreatic ductal epithelial (HPDE) cells, h-TERT transformed HPDE cells (HPNE), and four gain-of-function constructs related to EMT. The AHTM found HPNE, H-ras, Myr-AKT, and Bcl2 transfected cells more compliant relative to controls, consistent with parallel tests using atomic force microscopy and invasion assays, proving the AHTM capable of screening for changes in mechanical phenotype.

Quantitative volumetric imaging of normal, neoplastic and hyperplastic mouse prostate using ultrasound.

PubMed

Singh, Shalini; Pan, Chunliu; Wood, Ronald; Yeh, Chiuan-Ren; Yeh, Shuyuan; Sha, Kai; Krolewski, John J; Nastiuk, Kent L

2015-09-21

Genetically engineered mouse models are essential to the investigation of the molecular mechanisms underlying human prostate pathology and the effects of therapy on the diseased prostate. Serial in vivo volumetric imaging expands the scope and accuracy of experimental investigations of models of normal prostate physiology, benign prostatic hyperplasia and prostate cancer, which are otherwise limited by the anatomy of the mouse prostate. Moreover, accurate imaging of hyperplastic and tumorigenic prostates is now recognized as essential to rigorous pre-clinical trials of new therapies. Bioluminescent imaging has been widely used to determine prostate tumor size, but is semi-quantitative at best. Magnetic resonance imaging can determine prostate volume very accurately, but is expensive and has low throughput. We therefore sought to develop and implement a high throughput, low cost, and accurate serial imaging protocol for the mouse prostate. We developed a high frequency ultrasound imaging technique employing 3D reconstruction that allows rapid and precise assessment of mouse prostate volume. Wild-type mouse prostates were examined (n = 4) for reproducible baseline imaging, and treatment effects on volume were compared, and blinded data analyzed for intra- and inter-operator assessments of reproducibility by correlation and for Bland-Altman analysis. Examples of benign prostatic hyperplasia mouse model prostate (n = 2) and mouse prostate implantation of orthotopic human prostate cancer tumor and its growth (n =  ) are also demonstrated. Serial measurement volume of the mouse prostate revealed that high frequency ultrasound was very precise. Following endocrine manipulation, regression and regrowth of the prostate could be monitored with very low intra- and interobserver variability. This technique was also valuable to monitor the development of prostate growth in a model of benign prostatic hyperplasia. Additionally, we demonstrate accurate ultrasound image-guided implantation of orthotopic tumor xenografts and monitoring of subsequent tumor growth from ~10 to ~750 mm(3) volume. High frequency ultrasound imaging allows precise determination of normal, neoplastic and hyperplastic mouse prostate. Low cost and small image size allows incorporation of this imaging modality inside clean animal facilities, and thereby imaging of immunocompromised models. 3D reconstruction for volume determination is easily mastered, and both small and large relative changes in volume are accurately visualized. Ultrasound imaging does not rely on penetration of exogenous imaging agents, and so may therefore better measure poorly vascularized or necrotic diseased tissue, relative to bioluminescent imaging (IVIS). Our method is precise and reproducible with very low inter- and intra-observer variability. Because it is non-invasive, mouse models of prostatic disease states can be imaged serially, reducing inter-animal variability, and enhancing the power to detect small volume changes following therapeutic intervention.

Microfluidic Imaging Flow Cytometry by Asymmetric-detection Time-stretch Optical Microscopy (ATOM).

PubMed

Tang, Anson H L; Lai, Queenie T K; Chung, Bob M F; Lee, Kelvin C M; Mok, Aaron T Y; Yip, G K; Shum, Anderson H C; Wong, Kenneth K Y; Tsia, Kevin K

2017-06-28

Scaling the number of measurable parameters, which allows for multidimensional data analysis and thus higher-confidence statistical results, has been the main trend in the advanced development of flow cytometry. Notably, adding high-resolution imaging capabilities allows for the complex morphological analysis of cellular/sub-cellular structures. This is not possible with standard flow cytometers. However, it is valuable for advancing our knowledge of cellular functions and can benefit life science research, clinical diagnostics, and environmental monitoring. Incorporating imaging capabilities into flow cytometry compromises the assay throughput, primarily due to the limitations on speed and sensitivity in the camera technologies. To overcome this speed or throughput challenge facing imaging flow cytometry while preserving the image quality, asymmetric-detection time-stretch optical microscopy (ATOM) has been demonstrated to enable high-contrast, single-cell imaging with sub-cellular resolution, at an imaging throughput as high as 100,000 cells/s. Based on the imaging concept of conventional time-stretch imaging, which relies on all-optical image encoding and retrieval through the use of ultrafast broadband laser pulses, ATOM further advances imaging performance by enhancing the image contrast of unlabeled/unstained cells. This is achieved by accessing the phase-gradient information of the cells, which is spectrally encoded into single-shot broadband pulses. Hence, ATOM is particularly advantageous in high-throughput measurements of single-cell morphology and texture - information indicative of cell types, states, and even functions. Ultimately, this could become a powerful imaging flow cytometry platform for the biophysical phenotyping of cells, complementing the current state-of-the-art biochemical-marker-based cellular assay. This work describes a protocol to establish the key modules of an ATOM system (from optical frontend to data processing and visualization backend), as well as the workflow of imaging flow cytometry based on ATOM, using human cells and micro-algae as the examples.

A way toward analyzing high-content bioimage data by means of semantic annotation and visual data mining

NASA Astrophysics Data System (ADS)

Herold, Julia; Abouna, Sylvie; Zhou, Luxian; Pelengaris, Stella; Epstein, David B. A.; Khan, Michael; Nattkemper, Tim W.

2009-02-01

In the last years, bioimaging has turned from qualitative measurements towards a high-throughput and highcontent modality, providing multiple variables for each biological sample analyzed. We present a system which combines machine learning based semantic image annotation and visual data mining to analyze such new multivariate bioimage data. Machine learning is employed for automatic semantic annotation of regions of interest. The annotation is the prerequisite for a biological object-oriented exploration of the feature space derived from the image variables. With the aid of visual data mining, the obtained data can be explored simultaneously in the image as well as in the feature domain. Especially when little is known of the underlying data, for example in the case of exploring the effects of a drug treatment, visual data mining can greatly aid the process of data evaluation. We demonstrate how our system is used for image evaluation to obtain information relevant to diabetes study and screening of new anti-diabetes treatments. Cells of the Islet of Langerhans and whole pancreas in pancreas tissue samples are annotated and object specific molecular features are extracted from aligned multichannel fluorescence images. These are interactively evaluated for cell type classification in order to determine the cell number and mass. Only few parameters need to be specified which makes it usable also for non computer experts and allows for high-throughput analysis.

Strategies for high-throughput focused-beam ptychography

DOE PAGES

Jacobsen, Chris; Deng, Junjing; Nashed, Youssef

2017-08-08

X-ray ptychography is being utilized for a wide range of imaging experiments with a resolution beyond the limit of the X-ray optics used. Introducing a parameter for the ptychographic resolution gainG p(the ratio of the beam size over the achieved pixel size in the reconstructed image), strategies for data sampling and for increasing imaging throughput when the specimen is at the focus of an X-ray beam are considered. As a result, the tradeoffs between large and small illumination spots are examined.

A high throughput spectral image microscopy system

NASA Astrophysics Data System (ADS)

Gesley, M.; Puri, R.

2018-01-01

A high throughput spectral image microscopy system is configured for rapid detection of rare cells in large populations. To overcome flow cytometry rates and use of fluorophore tags, a system architecture integrates sample mechanical handling, signal processors, and optics in a non-confocal version of light absorption and scattering spectroscopic microscopy. Spectral images with native contrast do not require the use of exogeneous stain to render cells with submicron resolution. Structure may be characterized without restriction to cell clusters of differentiation.

Strategies for high-throughput focused-beam ptychography

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jacobsen, Chris; Deng, Junjing; Nashed, Youssef

X-ray ptychography is being utilized for a wide range of imaging experiments with a resolution beyond the limit of the X-ray optics used. Introducing a parameter for the ptychographic resolution gainG p(the ratio of the beam size over the achieved pixel size in the reconstructed image), strategies for data sampling and for increasing imaging throughput when the specimen is at the focus of an X-ray beam are considered. As a result, the tradeoffs between large and small illumination spots are examined.

High-throughput imaging of adult fluorescent zebrafish with an LED fluorescence macroscope

PubMed Central

Blackburn, Jessica S; Liu, Sali; Raimondi, Aubrey R; Ignatius, Myron S; Salthouse, Christopher D; Langenau, David M

2011-01-01

Zebrafish are a useful vertebrate model for the study of development, behavior, disease and cancer. A major advantage of zebrafish is that large numbers of animals can be economically used for experimentation; however, high-throughput methods for imaging live adult zebrafish had not been developed. Here, we describe protocols for building a light-emitting diode (LED) fluorescence macroscope and for using it to simultaneously image up to 30 adult animals that transgenically express a fluorescent protein, are transplanted with fluorescently labeled tumor cells or are tagged with fluorescent elastomers. These protocols show that the LED fluorescence macroscope is capable of distinguishing five fluorescent proteins and can image unanesthetized swimming adult zebrafish in multiple fluorescent channels simultaneously. The macroscope can be built and used for imaging within 1 day, whereas creating fluorescently labeled adult zebrafish requires 1 hour to several months, depending on the method chosen. The LED fluorescence macroscope provides a low-cost, high-throughput method to rapidly screen adult fluorescent zebrafish and it will be useful for imaging transgenic animals, screening for tumor engraftment, and tagging individual fish for long-term analysis. PMID:21293462

Androgen Receptor Functional Analyses by High Throughput Imaging: Determination of Ligand, Cell Cycle, and Mutation-Specific Effects

PubMed Central

Szafran, Adam T.; Szwarc, Maria; Marcelli, Marco; Mancini, Michael A.

2008-01-01

Background Understanding how androgen receptor (AR) function is modulated by exposure to steroids, growth factors or small molecules can have important mechanistic implications for AR-related disease therapies (e.g., prostate cancer, androgen insensitivity syndrome, AIS), and in the analysis of environmental endocrine disruptors. Methodology/Principal Findings We report the development of a high throughput (HT) image-based assay that quantifies AR subcellular and subnuclear distribution, and transcriptional reporter gene activity on a cell-by-cell basis. Furthermore, simultaneous analysis of DNA content allowed determination of cell cycle position and permitted the analysis of cell cycle dependent changes in AR function in unsynchronized cell populations. Assay quality for EC50 coefficients of variation were 5–24%, with Z' values reaching 0.91. This was achieved by the selective analysis of cells expressing physiological levels of AR, important because minor over-expression resulted in elevated nuclear speckling and decreased transcriptional reporter gene activity. A small screen of AR-binding ligands, including known agonists, antagonists, and endocrine disruptors, demonstrated that nuclear translocation and nuclear “speckling” were linked with transcriptional output, and specific ligands were noted to differentially affect measurements for wild type versus mutant AR, suggesting differing mechanisms of action. HT imaging of patient-derived AIS mutations demonstrated a proof-of-principle personalized medicine approach to rapidly identify ligands capable of restoring multiple AR functions. Conclusions/Significance HT imaging-based multiplex screening will provide a rapid, systems-level analysis of compounds/RNAi that may differentially affect wild type AR or clinically relevant AR mutations. PMID:18978937

Diffusion tensor imaging using multiple coils for mouse brain connectomics.

PubMed

Nouls, John C; Badea, Alexandra; Anderson, Robert B J; Cofer, Gary P; Allan Johnson, G

2018-06-01

The correlation between brain connectivity and psychiatric or neurological diseases has intensified efforts to develop brain connectivity mapping techniques on mouse models of human disease. The neural architecture of mouse brain specimens can be shown non-destructively and three-dimensionally by diffusion tensor imaging, which enables tractography, the establishment of a connectivity matrix and connectomics. However, experiments on cohorts of animals can be prohibitively long. To improve throughput in a 7-T preclinical scanner, we present a novel two-coil system in which each coil is shielded, placed off-isocenter along the axis of the magnet and connected to a receiver circuit of the scanner. Preservation of the quality factor of each coil is essential to signal-to-noise ratio (SNR) performance and throughput, because mouse brain specimen imaging at 7 T takes place in the coil-dominated noise regime. In that regime, we show a shielding configuration causing no SNR degradation in the two-coil system. To acquire data from several coils simultaneously, the coils are placed in the magnet bore, around the isocenter, in which gradient field distortions can bias diffusion tensor imaging metrics, affect tractography and contaminate measurements of the connectivity matrix. We quantified the experimental alterations in fractional anisotropy and eigenvector direction occurring in each coil. We showed that, when the coils were placed 12 mm away from the isocenter, measurements of the brain connectivity matrix appeared to be minimally altered by gradient field distortions. Simultaneous measurements on two mouse brain specimens demonstrated a full doubling of the diffusion tensor imaging throughput in practice. Each coil produced images devoid of shading or artifact. To further improve the throughput of mouse brain connectomics, we suggested a future expansion of the system to four coils. To better understand acceptable trade-offs between imaging throughput and connectivity matrix integrity, studies may seek to clarify how measurement variability, post-processing techniques and biological variability impact mouse brain connectomics. Copyright © 2018 John Wiley & Sons, Ltd.

Digital imaging of root traits (DIRT): a high-throughput computing and collaboration platform for field-based root phenomics.

PubMed

Das, Abhiram; Schneider, Hannah; Burridge, James; Ascanio, Ana Karine Martinez; Wojciechowski, Tobias; Topp, Christopher N; Lynch, Jonathan P; Weitz, Joshua S; Bucksch, Alexander

2015-01-01

Plant root systems are key drivers of plant function and yield. They are also under-explored targets to meet global food and energy demands. Many new technologies have been developed to characterize crop root system architecture (CRSA). These technologies have the potential to accelerate the progress in understanding the genetic control and environmental response of CRSA. Putting this potential into practice requires new methods and algorithms to analyze CRSA in digital images. Most prior approaches have solely focused on the estimation of root traits from images, yet no integrated platform exists that allows easy and intuitive access to trait extraction and analysis methods from images combined with storage solutions linked to metadata. Automated high-throughput phenotyping methods are increasingly used in laboratory-based efforts to link plant genotype with phenotype, whereas similar field-based studies remain predominantly manual low-throughput. Here, we present an open-source phenomics platform "DIRT", as a means to integrate scalable supercomputing architectures into field experiments and analysis pipelines. DIRT is an online platform that enables researchers to store images of plant roots, measure dicot and monocot root traits under field conditions, and share data and results within collaborative teams and the broader community. The DIRT platform seamlessly connects end-users with large-scale compute "commons" enabling the estimation and analysis of root phenotypes from field experiments of unprecedented size. DIRT is an automated high-throughput computing and collaboration platform for field based crop root phenomics. The platform is accessible at http://www.dirt.iplantcollaborative.org/ and hosted on the iPlant cyber-infrastructure using high-throughput grid computing resources of the Texas Advanced Computing Center (TACC). DIRT is a high volume central depository and high-throughput RSA trait computation platform for plant scientists working on crop roots. It enables scientists to store, manage and share crop root images with metadata and compute RSA traits from thousands of images in parallel. It makes high-throughput RSA trait computation available to the community with just a few button clicks. As such it enables plant scientists to spend more time on science rather than on technology. All stored and computed data is easily accessible to the public and broader scientific community. We hope that easy data accessibility will attract new tool developers and spur creative data usage that may even be applied to other fields of science.

Photon-Counting H33D Detector for Biological Fluorescence Imaging

PubMed Central

Michalet, X.; Siegmund, O.H.W.; Vallerga, J.V.; Jelinsky, P.; Millaud, J.E.; Weiss, S.

2010-01-01

We have developed a photon-counting High-temporal and High-spatial resolution, High-throughput 3-Dimensional detector (H33D) for biological imaging of fluorescent samples. The design is based on a 25 mm diameter S20 photocathode followed by a 3-microchannel plate stack, and a cross delay line anode. We describe the bench performance of the H33D detector, as well as preliminary imaging results obtained with fluorescent beads, quantum dots and live cells and discuss applications of future generation detectors for single-molecule imaging and high-throughput study of biomolecular interactions. PMID:20151021

Raspberry Pi-powered imaging for plant phenotyping.

PubMed

Tovar, Jose C; Hoyer, J Steen; Lin, Andy; Tielking, Allison; Callen, Steven T; Elizabeth Castillo, S; Miller, Michael; Tessman, Monica; Fahlgren, Noah; Carrington, James C; Nusinow, Dmitri A; Gehan, Malia A

2018-03-01

Image-based phenomics is a powerful approach to capture and quantify plant diversity. However, commercial platforms that make consistent image acquisition easy are often cost-prohibitive. To make high-throughput phenotyping methods more accessible, low-cost microcomputers and cameras can be used to acquire plant image data. We used low-cost Raspberry Pi computers and cameras to manage and capture plant image data. Detailed here are three different applications of Raspberry Pi-controlled imaging platforms for seed and shoot imaging. Images obtained from each platform were suitable for extracting quantifiable plant traits (e.g., shape, area, height, color) en masse using open-source image processing software such as PlantCV. This protocol describes three low-cost platforms for image acquisition that are useful for quantifying plant diversity. When coupled with open-source image processing tools, these imaging platforms provide viable low-cost solutions for incorporating high-throughput phenomics into a wide range of research programs.

Fast globally optimal segmentation of cells in fluorescence microscopy images.

PubMed

Bergeest, Jan-Philip; Rohr, Karl

2011-01-01

Accurate and efficient segmentation of cells in fluorescence microscopy images is of central importance for the quantification of protein expression in high-throughput screening applications. We propose a new approach for segmenting cell nuclei which is based on active contours and convex energy functionals. Compared to previous work, our approach determines the global solution. Thus, the approach does not suffer from local minima and the segmentation result does not depend on the initialization. We also suggest a numeric approach for efficiently computing the solution. The performance of our approach has been evaluated using fluorescence microscopy images of different cell types. We have also performed a quantitative comparison with previous segmentation approaches.

Integrated Analysis Platform: An Open-Source Information System for High-Throughput Plant Phenotyping1[C][W][OPEN

PubMed Central

Klukas, Christian; Chen, Dijun; Pape, Jean-Michel

2014-01-01

High-throughput phenotyping is emerging as an important technology to dissect phenotypic components in plants. Efficient image processing and feature extraction are prerequisites to quantify plant growth and performance based on phenotypic traits. Issues include data management, image analysis, and result visualization of large-scale phenotypic data sets. Here, we present Integrated Analysis Platform (IAP), an open-source framework for high-throughput plant phenotyping. IAP provides user-friendly interfaces, and its core functions are highly adaptable. Our system supports image data transfer from different acquisition environments and large-scale image analysis for different plant species based on real-time imaging data obtained from different spectra. Due to the huge amount of data to manage, we utilized a common data structure for efficient storage and organization of data for both input data and result data. We implemented a block-based method for automated image processing to extract a representative list of plant phenotypic traits. We also provide tools for build-in data plotting and result export. For validation of IAP, we performed an example experiment that contains 33 maize (Zea mays ‘Fernandez’) plants, which were grown for 9 weeks in an automated greenhouse with nondestructive imaging. Subsequently, the image data were subjected to automated analysis with the maize pipeline implemented in our system. We found that the computed digital volume and number of leaves correlate with our manually measured data in high accuracy up to 0.98 and 0.95, respectively. In summary, IAP provides a multiple set of functionalities for import/export, management, and automated analysis of high-throughput plant phenotyping data, and its analysis results are highly reliable. PMID:24760818

High-speed Fourier ptychographic microscopy based on programmable annular illuminations.

PubMed

Sun, Jiasong; Zuo, Chao; Zhang, Jialin; Fan, Yao; Chen, Qian

2018-05-16

High-throughput quantitative phase imaging (QPI) is essential to cellular phenotypes characterization as it allows high-content cell analysis and avoids adverse effects of staining reagents on cellular viability and cell signaling. Among different approaches, Fourier ptychographic microscopy (FPM) is probably the most promising technique to realize high-throughput QPI by synthesizing a wide-field, high-resolution complex image from multiple angle-variably illuminated, low-resolution images. However, the large dataset requirement in conventional FPM significantly limits its imaging speed, resulting in low temporal throughput. Moreover, the underlying theoretical mechanism as well as optimum illumination scheme for high-accuracy phase imaging in FPM remains unclear. Herein, we report a high-speed FPM technique based on programmable annular illuminations (AIFPM). The optical-transfer-function (OTF) analysis of FPM reveals that the low-frequency phase information can only be correctly recovered if the LEDs are precisely located at the edge of the objective numerical aperture (NA) in the frequency space. By using only 4 low-resolution images corresponding to 4 tilted illuminations matching a 10×, 0.4 NA objective, we present the high-speed imaging results of in vitro Hela cells mitosis and apoptosis at a frame rate of 25 Hz with a full-pitch resolution of 655 nm at a wavelength of 525 nm (effective NA = 0.8) across a wide field-of-view (FOV) of 1.77 mm 2 , corresponding to a space-bandwidth-time product of 411 megapixels per second. Our work reveals an important capability of FPM towards high-speed high-throughput imaging of in vitro live cells, achieving video-rate QPI performance across a wide range of scales, both spatial and temporal.

Nonlinear Optical Characterization of Membrane Protein Microcrystals and Nanocrystals.

PubMed

Newman, Justin A; Simpson, Garth J

2016-01-01

Nonlinear optical methods such as second harmonic generation (SHG) and two-photon excited UV fluorescence (TPE-UVF) imaging are promising approaches to address bottlenecks in the membrane protein structure determination pipeline. The general principles of SHG and TPE-UVF are discussed here along with instrument design considerations. Comparisons to conventional methods in high throughput crystallization condition screening and crystal quality assessment prior to X-ray diffraction are also discussed.

Ultrafast Microfluidic Cellular Imaging by Optical Time-Stretch.

PubMed

Lau, Andy K S; Wong, Terence T W; Shum, Ho Cheung; Wong, Kenneth K Y; Tsia, Kevin K

2016-01-01

There is an unmet need in biomedicine for measuring a multitude of parameters of individual cells (i.e., high content) in a large population efficiently (i.e., high throughput). This is particularly driven by the emerging interest in bringing Big-Data analysis into this arena, encompassing pathology, drug discovery, rare cancer cell detection, emulsion microdroplet assays, to name a few. This momentum is particularly evident in recent advancements in flow cytometry. They include scaling of the number of measurable colors from the labeled cells and incorporation of imaging capability to access the morphological information of the cells. However, an unspoken predicament appears in the current technologies: higher content comes at the expense of lower throughput, and vice versa. For example, accessing additional spatial information of individual cells, imaging flow cytometers only achieve an imaging throughput ~1000 cells/s, orders of magnitude slower than the non-imaging flow cytometers. In this chapter, we introduce an entirely new imaging platform, namely optical time-stretch microscopy, for ultrahigh speed and high contrast label-free single-cell (in a ultrafast microfluidic flow up to 10 m/s) imaging and analysis with an ultra-fast imaging line-scan rate as high as tens of MHz. Based on this technique, not only morphological information of the individual cells can be obtained in an ultrafast manner, quantitative evaluation of cellular information (e.g., cell volume, mass, refractive index, stiffness, membrane tension) at nanometer scale based on the optical phase is also possible. The technology can also be integrated with conventional fluorescence measurements widely adopted in the non-imaging flow cytometers. Therefore, these two combinatorial and complementary measurement capabilities in long run is an attractive platform for addressing the pressing need for expanding the "parameter space" in high-throughput single-cell analysis. This chapter provides the general guidelines of constructing the optical system for time stretch imaging, fabrication and design of the microfluidic chip for ultrafast fluidic flow, as well as the image acquisition and processing.

Segment and fit thresholding: a new method for image analysis applied to microarray and immunofluorescence data.

PubMed

Ensink, Elliot; Sinha, Jessica; Sinha, Arkadeep; Tang, Huiyuan; Calderone, Heather M; Hostetter, Galen; Winter, Jordan; Cherba, David; Brand, Randall E; Allen, Peter J; Sempere, Lorenzo F; Haab, Brian B

2015-10-06

Experiments involving the high-throughput quantification of image data require algorithms for automation. A challenge in the development of such algorithms is to properly interpret signals over a broad range of image characteristics, without the need for manual adjustment of parameters. Here we present a new approach for locating signals in image data, called Segment and Fit Thresholding (SFT). The method assesses statistical characteristics of small segments of the image and determines the best-fit trends between the statistics. Based on the relationships, SFT identifies segments belonging to background regions; analyzes the background to determine optimal thresholds; and analyzes all segments to identify signal pixels. We optimized the initial settings for locating background and signal in antibody microarray and immunofluorescence data and found that SFT performed well over multiple, diverse image characteristics without readjustment of settings. When used for the automated analysis of multicolor, tissue-microarray images, SFT correctly found the overlap of markers with known subcellular localization, and it performed better than a fixed threshold and Otsu's method for selected images. SFT promises to advance the goal of full automation in image analysis.

Segment and Fit Thresholding: A New Method for Image Analysis Applied to Microarray and Immunofluorescence Data

PubMed Central

Ensink, Elliot; Sinha, Jessica; Sinha, Arkadeep; Tang, Huiyuan; Calderone, Heather M.; Hostetter, Galen; Winter, Jordan; Cherba, David; Brand, Randall E.; Allen, Peter J.; Sempere, Lorenzo F.; Haab, Brian B.

2016-01-01

Certain experiments involve the high-throughput quantification of image data, thus requiring algorithms for automation. A challenge in the development of such algorithms is to properly interpret signals over a broad range of image characteristics, without the need for manual adjustment of parameters. Here we present a new approach for locating signals in image data, called Segment and Fit Thresholding (SFT). The method assesses statistical characteristics of small segments of the image and determines the best-fit trends between the statistics. Based on the relationships, SFT identifies segments belonging to background regions; analyzes the background to determine optimal thresholds; and analyzes all segments to identify signal pixels. We optimized the initial settings for locating background and signal in antibody microarray and immunofluorescence data and found that SFT performed well over multiple, diverse image characteristics without readjustment of settings. When used for the automated analysis of multi-color, tissue-microarray images, SFT correctly found the overlap of markers with known subcellular localization, and it performed better than a fixed threshold and Otsu’s method for selected images. SFT promises to advance the goal of full automation in image analysis. PMID:26339978

A Review of Imaging Techniques for Plant Phenotyping

PubMed Central

Li, Lei; Zhang, Qin; Huang, Danfeng

2014-01-01

Given the rapid development of plant genomic technologies, a lack of access to plant phenotyping capabilities limits our ability to dissect the genetics of quantitative traits. Effective, high-throughput phenotyping platforms have recently been developed to solve this problem. In high-throughput phenotyping platforms, a variety of imaging methodologies are being used to collect data for quantitative studies of complex traits related to the growth, yield and adaptation to biotic or abiotic stress (disease, insects, drought and salinity). These imaging techniques include visible imaging (machine vision), imaging spectroscopy (multispectral and hyperspectral remote sensing), thermal infrared imaging, fluorescence imaging, 3D imaging and tomographic imaging (MRT, PET and CT). This paper presents a brief review on these imaging techniques and their applications in plant phenotyping. The features used to apply these imaging techniques to plant phenotyping are described and discussed in this review. PMID:25347588

Factors that determine the optimum dose for sub-20nm resist systems: DUV, EUV, and e-beam options

NASA Astrophysics Data System (ADS)

Preil, Moshe

2012-03-01

As EUV and e-beam direct write (EBDW) technologies move closer to insertion into pilot production, questions regarding cost effectiveness take on increasing importance. One of the most critical questions is determining the optimum dose which balances the requirements for cost-effective throughput vs. imaging performance. To date most of the dose requirements have been dictated by the hardware side of the industry. The exposure tool manufacturers have a vested interest in specifying the fastest resists possible in order to maximize the throughput even if it comes at the expense of optimum resist performance. This is especially true for both EUV and EBDW where source power is severely limited. We will explore the cost-benefit tradeoffs which drive the equipment side of the industry, and show how these considerations lead to the current throughput and dose requirements for volume production tools. We will then show how the resulting low doses may lead to shot noise problems and a resulting penalty in resist performance. By comparison to the history of 248 nm DUV resist development we will illustrate how setting unrealistic initial targets for resist dose may lead to unacceptable tradeoffs in resist performance and subsequently long delays in the development of production worthy resists.

QR-on-a-chip: a computer-recognizable micro-pattern engraved microfluidic device for high-throughput image acquisition.

PubMed

Yun, Kyungwon; Lee, Hyunjae; Bang, Hyunwoo; Jeon, Noo Li

2016-02-21

This study proposes a novel way to achieve high-throughput image acquisition based on a computer-recognizable micro-pattern implemented on a microfluidic device. We integrated the QR code, a two-dimensional barcode system, onto the microfluidic device to simplify imaging of multiple ROIs (regions of interest). A standard QR code pattern was modified to arrays of cylindrical structures of polydimethylsiloxane (PDMS). Utilizing the recognition of the micro-pattern, the proposed system enables: (1) device identification, which allows referencing additional information of the device, such as device imaging sequences or the ROIs and (2) composing a coordinate system for an arbitrarily located microfluidic device with respect to the stage. Based on these functionalities, the proposed method performs one-step high-throughput imaging for data acquisition in microfluidic devices without further manual exploration and locating of the desired ROIs. In our experience, the proposed method significantly reduced the time for the preparation of an acquisition. We expect that the method will innovatively improve the prototype device data acquisition and analysis.

XML-based data model and architecture for a knowledge-based grid-enabled problem-solving environment for high-throughput biological imaging.

PubMed

Ahmed, Wamiq M; Lenz, Dominik; Liu, Jia; Paul Robinson, J; Ghafoor, Arif

2008-03-01

High-throughput biological imaging uses automated imaging devices to collect a large number of microscopic images for analysis of biological systems and validation of scientific hypotheses. Efficient manipulation of these datasets for knowledge discovery requires high-performance computational resources, efficient storage, and automated tools for extracting and sharing such knowledge among different research sites. Newly emerging grid technologies provide powerful means for exploiting the full potential of these imaging techniques. Efficient utilization of grid resources requires the development of knowledge-based tools and services that combine domain knowledge with analysis algorithms. In this paper, we first investigate how grid infrastructure can facilitate high-throughput biological imaging research, and present an architecture for providing knowledge-based grid services for this field. We identify two levels of knowledge-based services. The first level provides tools for extracting spatiotemporal knowledge from image sets and the second level provides high-level knowledge management and reasoning services. We then present cellular imaging markup language, an extensible markup language-based language for modeling of biological images and representation of spatiotemporal knowledge. This scheme can be used for spatiotemporal event composition, matching, and automated knowledge extraction and representation for large biological imaging datasets. We demonstrate the expressive power of this formalism by means of different examples and extensive experimental results.

EOS 2D/3D X-ray imaging system: a systematic review and economic evaluation.

PubMed

McKenna, C; Wade, R; Faria, R; Yang, H; Stirk, L; Gummerson, N; Sculpher, M; Woolacott, N

2012-01-01

EOS is a biplane X-ray imaging system manufactured by EOS Imaging (formerly Biospace Med, Paris, France). It uses slot-scanning technology to produce a high-quality image with less irradiation than standard imaging techniques. To determine the clinical effectiveness and cost-effectiveness of EOS two-dimensional (2D)/three-dimensional (3D) X-ray imaging system for the evaluation and monitoring of scoliosis and other relevant orthopaedic conditions. For the systematic review of EOS, electronic databases (MEDLINE, Allied and Complementary Medicine Database, BIOSIS Previews, Cumulative Index to Nursing and Allied Health Literature, The Cochrane Library, EMBASE, Health Management Information Consortium, Inspec, ISI Science Citation Index and PASCAL), clinical trials registries and the manufacturer's website were searched from 1993 to November 2010. A systematic review of studies comparing EOS with standard X-ray [film, computed radiography (CR) or digital radiography] in any orthopaedic condition was performed. A narrative synthesis was undertaken. A decision-analytic model was developed to assess the cost-effectiveness of EOS in the relevant indications compared with standard X-ray and incorporated the clinical effectiveness of EOS and the adverse effects of radiation. The model incorporated a lifetime horizon to estimate outcomes in terms of quality-adjusted life-years (QALYs) and costs from the perspective of the NHS. Three studies met the inclusion criteria for the review. Two studies compared EOS with film X-ray and one study compared EOS with CR. The three included studies were small and of limited quality. One study used an earlier version of the technology, the Charpak system. Both studies comparing EOS with film X-ray found image quality to be comparable or better with EOS overall. Radiation dose was considerably lower with EOS: ratio of means for posteroanterior spine was 5.2 (13.1 for the study using the Charpak system); ratio of means for the lateral spine was 6.2 (15.1 for the study using the Charpak system). The study comparing EOS with CR found image quality to be comparable or better with EOS. Radiation dose was considerably lower with EOS than CR; ratio of means for the centre of the back was 5.9 and for the proximal lateral point 8.8. The lowest ratio of means was at the nape of the neck, which was 2.9. No other outcomes were assessed in the included studies, such as implications for patient management from the nature and quality of the image. Patient throughput is the major determinant of the cost-effectiveness of EOS. The average cost per procedure of EOS decreases with utilisation. Using estimates of patient throughput at national level from Hospital Episode Statistics data suggests that EOS is not cost-effective for the indications considered. Throughput in the region of 15,100 to 26,500 (corresponding to a workload of 60 to 106 patient appointments per working day) for EOS compared with a throughput of only 7530 for CR (30 patient appointments per working day) is needed to achieve an incremental cost-effectiveness ratio of £30,000 per QALY. EOS can be shown to be cost-effective only when compared with CR if the utilisation for EOS is about double the utilisation of CR. The main limitation of the systematic review of the clinical effectiveness of EOS was the limited number and quality of the data available. In particular, there were no studies assessing the potential health benefits arising from the quality and nature of the image, over and above those associated with reduced radiation exposure. Uncertainty in the model inputs was not fully explored owing to a lack of reporting of standard deviations or confidence intervals in the published literature for most of the parameters. As a result, uncertainty in the cost-effectiveness results was not presented. Radiation dose is considerably lower with EOS than standard X-ray, whereas image quality remains comparable or better with EOS. However, the long-term health benefits from reduced radiation exposure with EOS are very small and there was a lack of data on other potential patient health benefits. The implications of any changes in the quality and nature of the EOS image compared with standard X-ray, for patient health outcomes, needs to be assessed. Given the higher cost of an EOS machine, utilisation is the major determinant of cost-effectiveness. Estimates of patient throughput at national level suggest that EOS is not cost-effective. The National Institute for Health Research Health Technology Assessment programme.

A review of snapshot multidimensional optical imaging: measuring photon tags in parallel

PubMed Central

Gao, Liang; Wang, Lihong V.

2015-01-01

Multidimensional optical imaging has seen remarkable growth in the past decade. Rather than measuring only the two-dimensional spatial distribution of light, as in conventional photography, multidimensional optical imaging captures light in up to nine dimensions, providing unprecedented information about incident photons’ spatial coordinates, emittance angles, wavelength, time, and polarization. Multidimensional optical imaging can be accomplished either by scanning or parallel acquisition. Compared with scanning-based imagers, parallel acquisition—also dubbed snapshot imaging—has a prominent advantage in maximizing optical throughput, particularly when measuring a datacube of high dimensions. Here, we first categorize snapshot multidimensional imagers based on their acquisition and image reconstruction strategies, then highlight the snapshot advantage in the context of optical throughput, and finally we discuss their state-of-the-art implementations and applications. PMID:27134340

Concepts for on board satellite image registration. Volume 4: Impact of data set selection on satellite on board signal processing

NASA Technical Reports Server (NTRS)

Ruedger, W. H.; Aanstoos, J. V.; Snyder, W. E.

1982-01-01

The NASA NEEDS program goals present a requirement for on-board signal processing to achieve user-compatible, information-adaptive data acquisition. This volume addresses the impact of data set selection on data formatting required for efficient telemetering of the acquired satellite sensor data. More specifically, the FILE algorithm developed by Martin-Marietta provides a means for the determination of those pixels from the data stream effects an improvement in the achievable system throughput. It will be seen that based on the lack of statistical stationarity in cloud cover, spatial distribution periods exist where data acquisition rates exceed the throughput capability. The study therefore addresses various approaches to data compression and truncation as applicable to this sensor mission.

Three-dimensional Imaging and Scanning: Current and Future Applications for Pathology

PubMed Central

Farahani, Navid; Braun, Alex; Jutt, Dylan; Huffman, Todd; Reder, Nick; Liu, Zheng; Yagi, Yukako; Pantanowitz, Liron

2017-01-01

Imaging is vital for the assessment of physiologic and phenotypic details. In the past, biomedical imaging was heavily reliant on analog, low-throughput methods, which would produce two-dimensional images. However, newer, digital, and high-throughput three-dimensional (3D) imaging methods, which rely on computer vision and computer graphics, are transforming the way biomedical professionals practice. 3D imaging has been useful in diagnostic, prognostic, and therapeutic decision-making for the medical and biomedical professions. Herein, we summarize current imaging methods that enable optimal 3D histopathologic reconstruction: Scanning, 3D scanning, and whole slide imaging. Briefly mentioned are emerging platforms, which combine robotics, sectioning, and imaging in their pursuit to digitize and automate the entire microscopy workflow. Finally, both current and emerging 3D imaging methods are discussed in relation to current and future applications within the context of pathology. PMID:28966836

Using high-throughput barcode sequencing to efficiently map connectomes

PubMed Central

Peikon, Ian D.; Kebschull, Justus M.; Vagin, Vasily V.; Ravens, Diana I.; Sun, Yu-Chi; Brouzes, Eric; Corrêa, Ivan R.; Bressan, Dario

2017-01-01

Abstract The function of a neural circuit is determined by the details of its synaptic connections. At present, the only available method for determining a neural wiring diagram with single synapse precision—a ‘connectome’—is based on imaging methods that are slow, labor-intensive and expensive. Here, we present SYNseq, a method for converting the connectome into a form that can exploit the speed and low cost of modern high-throughput DNA sequencing. In SYNseq, each neuron is labeled with a unique random nucleotide sequence—an RNA ‘barcode’—which is targeted to the synapse using engineered proteins. Barcodes in pre- and postsynaptic neurons are then associated through protein-protein crosslinking across the synapse, extracted from the tissue, and joined into a form suitable for sequencing. Although our failure to develop an efficient barcode joining scheme precludes the widespread application of this approach, we expect that with further development SYNseq will enable tracing of complex circuits at high speed and low cost. PMID:28449067

High performance gel imaging with a commercial single lens reflex camera

NASA Astrophysics Data System (ADS)

Slobodan, J.; Corbett, R.; Wye, N.; Schein, J. E.; Marra, M. A.; Coope, R. J. N.

2011-03-01

A high performance gel imaging system was constructed using a digital single lens reflex camera with epi-illumination to image 19 × 23 cm agarose gels with up to 10,000 DNA bands each. It was found to give equivalent performance to a laser scanner in this high throughput DNA fingerprinting application using the fluorophore SYBR Green®. The specificity and sensitivity of the imager and scanner were within 1% using the same band identification software. Low and high cost color filters were also compared and it was found that with care, good results could be obtained with inexpensive dyed acrylic filters in combination with more costly dielectric interference filters, but that very poor combinations were also possible. Methods for determining resolution, dynamic range, and optical efficiency for imagers are also proposed to facilitate comparison between systems.

Astronomical Polarimetry with the RIT Polarization Imaging Camera

NASA Astrophysics Data System (ADS)

Vorobiev, Dmitry V.; Ninkov, Zoran; Brock, Neal

2018-06-01

In the last decade, imaging polarimeters based on micropolarizer arrays have been developed for use in terrestrial remote sensing and metrology applications. Micropolarizer-based sensors are dramatically smaller and more mechanically robust than other polarimeters with similar spectral response and snapshot capability. To determine the suitability of these new polarimeters for astronomical applications, we developed the RIT Polarization Imaging Camera to investigate the performance of these devices, with a special attention to the low signal-to-noise regime. We characterized the device performance in the lab, by determining the relative throughput, efficiency, and orientation of every pixel, as a function of wavelength. Using the resulting pixel response model, we developed demodulation procedures for aperture photometry and imaging polarimetry observing modes. We found that, using the current calibration, RITPIC is capable of detecting polarization signals as small as ∼0.3%. The relative ease of data collection, calibration, and analysis provided by these sensors suggest than they may become an important tool for a number of astronomical targets.

Quantitative monitoring of Arabidopsis thaliana growth and development using high-throughput plant phenotyping

PubMed Central

Arend, Daniel; Lange, Matthias; Pape, Jean-Michel; Weigelt-Fischer, Kathleen; Arana-Ceballos, Fernando; Mücke, Ingo; Klukas, Christian; Altmann, Thomas; Scholz, Uwe; Junker, Astrid

2016-01-01

With the implementation of novel automated, high throughput methods and facilities in the last years, plant phenomics has developed into a highly interdisciplinary research domain integrating biology, engineering and bioinformatics. Here we present a dataset of a non-invasive high throughput plant phenotyping experiment, which uses image- and image analysis- based approaches to monitor the growth and development of 484 Arabidopsis thaliana plants (thale cress). The result is a comprehensive dataset of images and extracted phenotypical features. Such datasets require detailed documentation, standardized description of experimental metadata as well as sustainable data storage and publication in order to ensure the reproducibility of experiments, data reuse and comparability among the scientific community. Therefore the here presented dataset has been annotated using the standardized ISA-Tab format and considering the recently published recommendations for the semantical description of plant phenotyping experiments. PMID:27529152

Quantitative monitoring of Arabidopsis thaliana growth and development using high-throughput plant phenotyping.

PubMed

Arend, Daniel; Lange, Matthias; Pape, Jean-Michel; Weigelt-Fischer, Kathleen; Arana-Ceballos, Fernando; Mücke, Ingo; Klukas, Christian; Altmann, Thomas; Scholz, Uwe; Junker, Astrid

2016-08-16

With the implementation of novel automated, high throughput methods and facilities in the last years, plant phenomics has developed into a highly interdisciplinary research domain integrating biology, engineering and bioinformatics. Here we present a dataset of a non-invasive high throughput plant phenotyping experiment, which uses image- and image analysis- based approaches to monitor the growth and development of 484 Arabidopsis thaliana plants (thale cress). The result is a comprehensive dataset of images and extracted phenotypical features. Such datasets require detailed documentation, standardized description of experimental metadata as well as sustainable data storage and publication in order to ensure the reproducibility of experiments, data reuse and comparability among the scientific community. Therefore the here presented dataset has been annotated using the standardized ISA-Tab format and considering the recently published recommendations for the semantical description of plant phenotyping experiments.

High-throughput behavioral screening method for detecting auditory response defects in zebrafish.

PubMed

Bang, Pascal I; Yelick, Pamela C; Malicki, Jarema J; Sewell, William F

2002-08-30

We have developed an automated, high-throughput behavioral screening method for detecting hearing defects in zebrafish. Our assay monitors a rapid escape reflex in response to a loud sound. With this approach, 36 adult zebrafish, restrained in visually isolated compartments, can be simultaneously assessed for responsiveness to near-field 400 Hz sinusoidal tone bursts. Automated, objective determinations of responses are achieved with a computer program that obtains images at precise times relative to the acoustic stimulus. Images taken with a CCD video camera before and after stimulus presentation are subtracted to reveal a response to the sound. Up to 108 fish can be screened per hour. Over 6500 fish were tested to validate the reliability of the assay. We found that 1% of these animals displayed hearing deficits. The phenotypes of non-responders were further assessed with radiological analysis for defects in the gross morphology of the auditory system. Nearly all of those showed abnormalities in conductive elements of the auditory system: the swim bladder or Weberian ossicles. Copyright 2002 Elsevier Science B.V.

Nile Red Detection of Bacterial Hydrocarbons and Ketones in a High-Throughput Format

PubMed Central

Pinzon, Neissa M.; Aukema, Kelly G.; Gralnick, Jeffrey A.; Wackett, Lawrence P.

2011-01-01

ABSTRACT A method for use in high-throughput screening of bacteria for the production of long-chain hydrocarbons and ketones by monitoring fluorescent light emission in the presence of Nile red is described. Nile red has previously been used to screen for polyhydroxybutyrate (PHB) and fatty acid esters, but this is the first report of screening for recombinant bacteria making hydrocarbons or ketones. The microtiter plate assay was evaluated using wild-type and recombinant strains of Shewanella oneidensis and Escherichia coli expressing the enzyme OleA, previously shown to initiate hydrocarbon biosynthesis. The strains expressing exogenous Stenotrophomonas maltophilia oleA, with increased levels of ketone production as determined by gas chromatography-mass spectrometry, were distinguished with Nile red fluorescence. Confocal microscopy images of S. oneidensis oleA-expressing strains stained with Nile red were consistent with a membrane localization of the ketones. This differed from Nile red staining of bacterial PHB or algal lipid droplets that showed intracellular inclusion bodies. These results demonstrated the applicability of Nile red in a high-throughput technique for the detection of bacterial hydrocarbons and ketones. PMID:21712420

Quantitative digital image analysis of chromogenic assays for high throughput screening of alpha-amylase mutant libraries.

PubMed

Shankar, Manoharan; Priyadharshini, Ramachandran; Gunasekaran, Paramasamy

2009-08-01

An image analysis-based method for high throughput screening of an alpha-amylase mutant library using chromogenic assays was developed. Assays were performed in microplates and high resolution images of the assay plates were read using the Virtual Microplate Reader (VMR) script to quantify the concentration of the chromogen. This method is fast and sensitive in quantifying 0.025-0.3 mg starch/ml as well as 0.05-0.75 mg glucose/ml. It was also an effective screening method for improved alpha-amylase activity with a coefficient of variance of 18%.

Wavelength Scanning with a Tilting Interference Filter for Glow-Discharge Elemental Imaging.

PubMed

Storey, Andrew P; Ray, Steven J; Hoffmann, Volker; Voronov, Maxim; Engelhard, Carsten; Buscher, Wolfgang; Hieftje, Gary M

2017-06-01

Glow discharges have long been used for depth profiling and bulk analysis of solid samples. In addition, over the past decade, several methods of obtaining lateral surface elemental distributions have been introduced, each with its own strengths and weaknesses. Challenges for each of these techniques are acceptable optical throughput and added instrumental complexity. Here, these problems are addressed with a tilting-filter instrument. A pulsed glow discharge is coupled to an optical system comprising an adjustable-angle tilting filter, collimating and imaging lenses, and a gated, intensified charge-coupled device (CCD) camera, which together provide surface elemental mapping of solid samples. The tilting-filter spectrometer is instrumentally simpler, produces less image distortion, and achieves higher optical throughput than a monochromator-based instrument, but has a much more limited tunable spectral range and poorer spectral resolution. As a result, the tilting-filter spectrometer is limited to single-element or two-element determinations, and only when the target spectral lines fall within an appropriate spectral range and can be spectrally discerned. Spectral interferences that result from heterogeneous impurities can be flagged and overcome by observing the spatially resolved signal response across the available tunable spectral range. The instrument has been characterized and evaluated for the spatially resolved analysis of glow-discharge emission from selected but representative samples.

Real-time Image Processing for Microscopy-based Label-free Imaging Flow Cytometry in a Microfluidic Chip.

PubMed

Heo, Young Jin; Lee, Donghyeon; Kang, Junsu; Lee, Keondo; Chung, Wan Kyun

2017-09-14

Imaging flow cytometry (IFC) is an emerging technology that acquires single-cell images at high-throughput for analysis of a cell population. Rich information that comes from high sensitivity and spatial resolution of a single-cell microscopic image is beneficial for single-cell analysis in various biological applications. In this paper, we present a fast image-processing pipeline (R-MOD: Real-time Moving Object Detector) based on deep learning for high-throughput microscopy-based label-free IFC in a microfluidic chip. The R-MOD pipeline acquires all single-cell images of cells in flow, and identifies the acquired images as a real-time process with minimum hardware that consists of a microscope and a high-speed camera. Experiments show that R-MOD has the fast and reliable accuracy (500 fps and 93.3% mAP), and is expected to be used as a powerful tool for biomedical and clinical applications.

Non-destructive Determination of Shikimic Acid Concentration in Transgenic Maize Exhibiting Glyphosate Tolerance Using Chlorophyll Fluorescence and Hyperspectral Imaging

PubMed Central

Feng, Xuping; Yu, Chenliang; Chen, Yue; Peng, Jiyun; Ye, Lanhan; Shen, Tingting; Wen, Haiyong; He, Yong

2018-01-01

The development of transgenic glyphosate-tolerant crops has revolutionized weed control in crops in many regions of the world. The early, non-destructive identification of superior plant phenotypes is an important stage in plant breeding programs. Here, glyphosate-tolerant transgenic maize and its parental wild-type control were studied at 2, 4, 6, and 8 days after glyphosate treatment. Visible and near-infrared hyperspectral imaging and chlorophyll fluorescence imaging techniques were applied to monitor the performance of plants. In our research, transgenic maize, which was highly tolerant to glyphosate, was phenotyped using these high-throughput non-destructive methods to validate low levels of shikimic acid accumulation and high photochemical efficiency of photosystem II as reflected by maximum quantum yield and non-photochemical quenching in response to glyphosate. For hyperspectral imaging analysis, the combination of spectroscopy and chemometric methods was used to predict shikimic acid concentration. Our results indicated that a partial least-squares regression model, built on optimal wavelengths, effectively predicted shikimic acid concentrations, with a coefficient of determination value of 0.79 for the calibration set, and 0.82 for the prediction set. Moreover, shikimic acid concentration estimates from hyperspectral images were visualized on the prediction maps by spectral features, which could help in developing a simple multispectral imaging instrument for non-destructive phenotyping. Specific physiological effects of glyphosate affected the photochemical processes of maize, which induced substantial changes in chlorophyll fluorescence characteristics. A new data-driven method, combining mean fluorescence parameters and featuring a screening approach, provided a satisfactory relationship between fluorescence parameters and shikimic acid content. The glyphosate-tolerant transgenic plants can be identified with the developed discrimination model established on important wavelengths or sensitive fluorescence parameters 6 days after glyphosate treatment. The overall results indicated that both hyperspectral imaging and chlorophyll fluorescence imaging techniques could provide useful tools for stress phenotyping in maize breeding programs and could enable the detection and evaluation of superior genotypes, such as glyphosate tolerance, with a non-destructive high-throughput technique. PMID:29686693

Non-destructive Determination of Shikimic Acid Concentration in Transgenic Maize Exhibiting Glyphosate Tolerance Using Chlorophyll Fluorescence and Hyperspectral Imaging.

PubMed

Feng, Xuping; Yu, Chenliang; Chen, Yue; Peng, Jiyun; Ye, Lanhan; Shen, Tingting; Wen, Haiyong; He, Yong

2018-01-01

The development of transgenic glyphosate-tolerant crops has revolutionized weed control in crops in many regions of the world. The early, non-destructive identification of superior plant phenotypes is an important stage in plant breeding programs. Here, glyphosate-tolerant transgenic maize and its parental wild-type control were studied at 2, 4, 6, and 8 days after glyphosate treatment. Visible and near-infrared hyperspectral imaging and chlorophyll fluorescence imaging techniques were applied to monitor the performance of plants. In our research, transgenic maize, which was highly tolerant to glyphosate, was phenotyped using these high-throughput non-destructive methods to validate low levels of shikimic acid accumulation and high photochemical efficiency of photosystem II as reflected by maximum quantum yield and non-photochemical quenching in response to glyphosate. For hyperspectral imaging analysis, the combination of spectroscopy and chemometric methods was used to predict shikimic acid concentration. Our results indicated that a partial least-squares regression model, built on optimal wavelengths, effectively predicted shikimic acid concentrations, with a coefficient of determination value of 0.79 for the calibration set, and 0.82 for the prediction set. Moreover, shikimic acid concentration estimates from hyperspectral images were visualized on the prediction maps by spectral features, which could help in developing a simple multispectral imaging instrument for non-destructive phenotyping. Specific physiological effects of glyphosate affected the photochemical processes of maize, which induced substantial changes in chlorophyll fluorescence characteristics. A new data-driven method, combining mean fluorescence parameters and featuring a screening approach, provided a satisfactory relationship between fluorescence parameters and shikimic acid content. The glyphosate-tolerant transgenic plants can be identified with the developed discrimination model established on important wavelengths or sensitive fluorescence parameters 6 days after glyphosate treatment. The overall results indicated that both hyperspectral imaging and chlorophyll fluorescence imaging techniques could provide useful tools for stress phenotyping in maize breeding programs and could enable the detection and evaluation of superior genotypes, such as glyphosate tolerance, with a non-destructive high-throughput technique.

Depth-resolved incoherent and coherent wide-field high-content imaging (Conference Presentation)

NASA Astrophysics Data System (ADS)

So, Peter T.

2016-03-01

Recent advances in depth-resolved wide-field imaging technique has enabled many high throughput applications in biology and medicine. Depth resolved imaging of incoherent signals can be readily accomplished with structured light illumination or nonlinear temporal focusing. The integration of these high throughput systems with novel spectroscopic resolving elements further enable high-content information extraction. We will introduce a novel near common-path interferometer and demonstrate its uses in toxicology and cancer biology applications. The extension of incoherent depth-resolved wide-field imaging to coherent modality is non-trivial. Here, we will cover recent advances in wide-field 3D resolved mapping of refractive index, absorbance, and vibronic components in biological specimens.

Comprehensive optical and data management infrastructure for high-throughput light-sheet microscopy of whole mouse brains.

PubMed

Müllenbroich, M Caroline; Silvestri, Ludovico; Onofri, Leonardo; Costantini, Irene; Hoff, Marcel Van't; Sacconi, Leonardo; Iannello, Giulio; Pavone, Francesco S

2015-10-01

Comprehensive mapping and quantification of neuronal projections in the central nervous system requires high-throughput imaging of large volumes with microscopic resolution. To this end, we have developed a confocal light-sheet microscope that has been optimized for three-dimensional (3-D) imaging of structurally intact clarified whole-mount mouse brains. We describe the optical and electromechanical arrangement of the microscope and give details on the organization of the microscope management software. The software orchestrates all components of the microscope, coordinates critical timing and synchronization, and has been written in a versatile and modular structure using the LabVIEW language. It can easily be adapted and integrated to other microscope systems and has been made freely available to the light-sheet community. The tremendous amount of data routinely generated by light-sheet microscopy further requires novel strategies for data handling and storage. To complete the full imaging pipeline of our high-throughput microscope, we further elaborate on big data management from streaming of raw images up to stitching of 3-D datasets. The mesoscale neuroanatomy imaged at micron-scale resolution in those datasets allows characterization and quantification of neuronal projections in unsectioned mouse brains.

Automatic Segmentation of High-Throughput RNAi Fluorescent Cellular Images

PubMed Central

Yan, Pingkum; Zhou, Xiaobo; Shah, Mubarak; Wong, Stephen T. C.

2010-01-01

High-throughput genome-wide RNA interference (RNAi) screening is emerging as an essential tool to assist biologists in understanding complex cellular processes. The large number of images produced in each study make manual analysis intractable; hence, automatic cellular image analysis becomes an urgent need, where segmentation is the first and one of the most important steps. In this paper, a fully automatic method for segmentation of cells from genome-wide RNAi screening images is proposed. Nuclei are first extracted from the DNA channel by using a modified watershed algorithm. Cells are then extracted by modeling the interaction between them as well as combining both gradient and region information in the Actin and Rac channels. A new energy functional is formulated based on a novel interaction model for segmenting tightly clustered cells with significant intensity variance and specific phenotypes. The energy functional is minimized by using a multiphase level set method, which leads to a highly effective cell segmentation method. Promising experimental results demonstrate that automatic segmentation of high-throughput genome-wide multichannel screening can be achieved by using the proposed method, which may also be extended to other multichannel image segmentation problems. PMID:18270043

High throughput dual-wavelength temperature distribution imaging via compressive imaging

NASA Astrophysics Data System (ADS)

Yao, Xu-Ri; Lan, Ruo-Ming; Liu, Xue-Feng; Zhu, Ge; Zheng, Fu; Yu, Wen-Kai; Zhai, Guang-Jie

2018-03-01

Thermal imaging is an essential tool in a wide variety of research areas. In this work we demonstrate high-throughput double-wavelength temperature distribution imaging using a modified single-pixel camera without the requirement of a beam splitter (BS). A digital micro-mirror device (DMD) is utilized to display binary masks and split the incident radiation, which eliminates the necessity of a BS. Because the spatial resolution is dictated by the DMD, this thermal imaging system has the advantage of perfect spatial registration between the two images, which limits the need for the pixel registration and fine adjustments. Two bucket detectors, which measures the total light intensity reflected from the DMD, are employed in this system and yield an improvement in the detection efficiency of the narrow-band radiation. A compressive imaging algorithm is utilized to achieve under-sampling recovery. A proof-of-principle experiment was presented to demonstrate the feasibility of this structure.

Fluorescence imaging technology (FI) for high-throughput screening of selenide-modified nano-TiO2 catalysts.

PubMed

Wang, Liping; Lee, Jianchao; Zhang, Meijuan; Duan, Qiannan; Zhang, Jiarui; Qi, Hailang

2016-02-18

A high-throughput screening (HTS) method based on fluorescence imaging (FI) was implemented to evaluate the catalytic performance of selenide-modified nano-TiO2. Chemical ink-jet printing (IJP) technology was reformed to fabricate a catalyst library comprising 1405 (Ni(a)Cu(b)Cd(c)Ce(d)In(e)Y(f))Se(x)/TiO2 (M6Se/Ti) composite photocatalysts. Nineteen M6Se/Tis were screened out from the 1405 candidates efficiently.

Large-scale microfluidics providing high-resolution and high-throughput screening of Caenorhabditis elegans poly-glutamine aggregation model

NASA Astrophysics Data System (ADS)

Mondal, Sudip; Hegarty, Evan; Martin, Chris; Gökçe, Sertan Kutal; Ghorashian, Navid; Ben-Yakar, Adela

2016-10-01

Next generation drug screening could benefit greatly from in vivo studies, using small animal models such as Caenorhabditis elegans for hit identification and lead optimization. Current in vivo assays can operate either at low throughput with high resolution or with low resolution at high throughput. To enable both high-throughput and high-resolution imaging of C. elegans, we developed an automated microfluidic platform. This platform can image 15 z-stacks of ~4,000 C. elegans from 96 different populations using a large-scale chip with a micron resolution in 16 min. Using this platform, we screened ~100,000 animals of the poly-glutamine aggregation model on 25 chips. We tested the efficacy of ~1,000 FDA-approved drugs in improving the aggregation phenotype of the model and identified four confirmed hits. This robust platform now enables high-content screening of various C. elegans disease models at the speed and cost of in vitro cell-based assays.

High throughput imaging and analysis for biological interpretation of agricultural plants and environmental interaction

NASA Astrophysics Data System (ADS)

Hong, Hyundae; Benac, Jasenka; Riggsbee, Daniel; Koutsky, Keith

2014-03-01

High throughput (HT) phenotyping of crops is essential to increase yield in environments deteriorated by climate change. The controlled environment of a greenhouse offers an ideal platform to study the genotype to phenotype linkages for crop screening. Advanced imaging technologies are used to study plants' responses to resource limitations such as water and nutrient deficiency. Advanced imaging technologies coupled with automation make HT phenotyping in the greenhouse not only feasible, but practical. Monsanto has a state of the art automated greenhouse (AGH) facility. Handling of the soil, pots water and nutrients are all completely automated. Images of the plants are acquired by multiple hyperspectral and broadband cameras. The hyperspectral cameras cover wavelengths from visible light through short wave infra-red (SWIR). Inhouse developed software analyzes the images to measure plant morphological and biochemical properties. We measure phenotypic metrics like plant area, height, and width as well as biomass. Hyperspectral imaging allows us to measure biochemcical metrics such as chlorophyll, anthocyanin, and foliar water content. The last 4 years of AGH operations on crops like corn, soybean, and cotton have demonstrated successful application of imaging and analysis technologies for high throughput plant phenotyping. Using HT phenotyping, scientists have been showing strong correlations to environmental conditions, such as water and nutrient deficits, as well as the ability to tease apart distinct differences in the genetic backgrounds of crops.

Architecture for a PACS primary diagnosis workstation

NASA Astrophysics Data System (ADS)

Shastri, Kaushal; Moran, Byron

1990-08-01

A major factor in determining the overall utility of a medical Picture Archiving and Communications (PACS) system is the functionality of the diagnostic workstation. Meyer-Ebrecht and Wendler [1] have proposed a modular picture computer architecture with high throughput and Perry et.al [2] have defined performance requirements for radiology workstations. In order to be clinically useful, a primary diagnosis workstation must not only provide functions of current viewing systems (e.g. mechanical alternators [3,4]) such as acceptable image quality, simultaneous viewing of multiple images, and rapid switching of image banks; but must also provide a diagnostic advantage over the current systems. This includes window-level functions on any image, simultaneous display of multi-modality images, rapid image manipulation, image processing, dynamic image display (cine), electronic image archival, hardcopy generation, image acquisition, network support, and an easy user interface. Implementation of such a workstation requires an underlying hardware architecture which provides high speed image transfer channels, local storage facilities, and image processing functions. This paper describes the hardware architecture of the Siemens Diagnostic Reporting Console (DRC) which meets these requirements.

3D imaging of optically cleared tissue using a simplified CLARITY method and on-chip microscopy

PubMed Central

Zhang, Yibo; Shin, Yoonjung; Sung, Kevin; Yang, Sam; Chen, Harrison; Wang, Hongda; Teng, Da; Rivenson, Yair; Kulkarni, Rajan P.; Ozcan, Aydogan

2017-01-01

High-throughput sectioning and optical imaging of tissue samples using traditional immunohistochemical techniques can be costly and inaccessible in resource-limited areas. We demonstrate three-dimensional (3D) imaging and phenotyping in optically transparent tissue using lens-free holographic on-chip microscopy as a low-cost, simple, and high-throughput alternative to conventional approaches. The tissue sample is passively cleared using a simplified CLARITY method and stained using 3,3′-diaminobenzidine to target cells of interest, enabling bright-field optical imaging and 3D sectioning of thick samples. The lens-free computational microscope uses pixel super-resolution and multi-height phase recovery algorithms to digitally refocus throughout the cleared tissue and obtain a 3D stack of complex-valued images of the sample, containing both phase and amplitude information. We optimized the tissue-clearing and imaging system by finding the optimal illumination wavelength, tissue thickness, sample preparation parameters, and the number of heights of the lens-free image acquisition and implemented a sparsity-based denoising algorithm to maximize the imaging volume and minimize the amount of the acquired data while also preserving the contrast-to-noise ratio of the reconstructed images. As a proof of concept, we achieved 3D imaging of neurons in a 200-μm-thick cleared mouse brain tissue over a wide field of view of 20.5 mm2. The lens-free microscope also achieved more than an order-of-magnitude reduction in raw data compared to a conventional scanning optical microscope imaging the same sample volume. Being low cost, simple, high-throughput, and data-efficient, we believe that this CLARITY-enabled computational tissue imaging technique could find numerous applications in biomedical diagnosis and research in low-resource settings. PMID:28819645

Optimizing ultrafast illumination for multiphoton-excited fluorescence imaging

PubMed Central

Stoltzfus, Caleb R.; Rebane, Aleksander

2016-01-01

We study the optimal conditions for high throughput two-photon excited fluorescence (2PEF) and three-photon excited fluorescence (3PEF) imaging using femtosecond lasers. We derive relations that allow maximization of the rate of imaging depending on the average power, pulse repetition rate, and noise characteristics of the laser, as well as on the size and structure of the sample. We perform our analysis using ~100 MHz, ~1 MHz and 1 kHz pulse rates and using both a tightly-focused illumination beam with diffraction-limited image resolution, as well loosely focused illumination with a relatively low image resolution, where the latter utilizes separate illumination and fluorescence detection beam paths. Our theoretical estimates agree with the experiments, which makes our approach especially useful for optimizing high throughput imaging of large samples with a field-of-view up to 10x10 cm2. PMID:27231620

High resolution light-sheet based high-throughput imaging cytometry system enables visualization of intra-cellular organelles

NASA Astrophysics Data System (ADS)

Regmi, Raju; Mohan, Kavya; Mondal, Partha Pratim

2014-09-01

Visualization of intracellular organelles is achieved using a newly developed high throughput imaging cytometry system. This system interrogates the microfluidic channel using a sheet of light rather than the existing point-based scanning techniques. The advantages of the developed system are many, including, single-shot scanning of specimens flowing through the microfluidic channel at flow rate ranging from micro- to nano- lit./min. Moreover, this opens-up in-vivo imaging of sub-cellular structures and simultaneous cell counting in an imaging cytometry system. We recorded a maximum count of 2400 cells/min at a flow-rate of 700 nl/min, and simultaneous visualization of fluorescently-labeled mitochondrial network in HeLa cells during flow. The developed imaging cytometry system may find immediate application in biotechnology, fluorescence microscopy and nano-medicine.

Recent advances in high-throughput QCL-based infrared microspectral imaging (Conference Presentation)

NASA Astrophysics Data System (ADS)

Rowlette, Jeremy A.; Fotheringham, Edeline; Nichols, David; Weida, Miles J.; Kane, Justin; Priest, Allen; Arnone, David B.; Bird, Benjamin; Chapman, William B.; Caffey, David B.; Larson, Paul; Day, Timothy

2017-02-01

The field of infrared spectral imaging and microscopy is advancing rapidly due in large measure to the recent commercialization of the first high-throughput, high-spatial-definition quantum cascade laser (QCL) microscope. Having speed, resolution and noise performance advantages while also eliminating the need for cryogenic cooling, its introduction has established a clear path to translating the well-established diagnostic capability of infrared spectroscopy into clinical and pre-clinical histology, cytology and hematology workflows. Demand for even higher throughput while maintaining high-spectral fidelity and low-noise performance continues to drive innovation in QCL-based spectral imaging instrumentation. In this talk, we will present for the first time, recent technological advances in tunable QCL photonics which have led to an additional 10X enhancement in spectral image data collection speed while preserving the high spectral fidelity and SNR exhibited by the first generation of QCL microscopes. This new approach continues to leverage the benefits of uncooled microbolometer focal plane array cameras, which we find to be essential for ensuring both reproducibility of data across instruments and achieving the high-reliability needed in clinical applications. We will discuss the physics underlying these technological advancements as well as the new biomedical applications these advancements are enabling, including automated whole-slide infrared chemical imaging on clinically relevant timescales.

Using high-throughput barcode sequencing to efficiently map connectomes.

PubMed

Peikon, Ian D; Kebschull, Justus M; Vagin, Vasily V; Ravens, Diana I; Sun, Yu-Chi; Brouzes, Eric; Corrêa, Ivan R; Bressan, Dario; Zador, Anthony M

2017-07-07

The function of a neural circuit is determined by the details of its synaptic connections. At present, the only available method for determining a neural wiring diagram with single synapse precision-a 'connectome'-is based on imaging methods that are slow, labor-intensive and expensive. Here, we present SYNseq, a method for converting the connectome into a form that can exploit the speed and low cost of modern high-throughput DNA sequencing. In SYNseq, each neuron is labeled with a unique random nucleotide sequence-an RNA 'barcode'-which is targeted to the synapse using engineered proteins. Barcodes in pre- and postsynaptic neurons are then associated through protein-protein crosslinking across the synapse, extracted from the tissue, and joined into a form suitable for sequencing. Although our failure to develop an efficient barcode joining scheme precludes the widespread application of this approach, we expect that with further development SYNseq will enable tracing of complex circuits at high speed and low cost. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Real-time Full-spectral Imaging and Affinity Measurements from 50 Microfluidic Channels using Nanohole Surface Plasmon Resonance†

PubMed Central

Lee, Si Hoon; Lindquist, Nathan C.; Wittenberg, Nathan J.; Jordan, Luke R.; Oh, Sang-Hyun

2012-01-01

With recent advances in high-throughput proteomics and systems biology, there is a growing demand for new instruments that can precisely quantify a wide range of receptor-ligand binding kinetics in a high-throughput fashion. Here we demonstrate a surface plasmon resonance (SPR) imaging spectroscopy instrument capable of extracting binding kinetics and affinities from 50 parallel microfluidic channels simultaneously. The instrument utilizes large-area (~cm2) metallic nanohole arrays as SPR sensing substrates and combines a broadband light source, a high-resolution imaging spectrometer and a low-noise CCD camera to extract spectral information from every channel in real time with a refractive index resolution of 7.7 × 10−6. To demonstrate the utility of our instrument for quantifying a wide range of biomolecular interactions, each parallel microfluidic channel is coated with a biomimetic supported lipid membrane containing ganglioside (GM1) receptors. The binding kinetics of cholera toxin b (CTX-b) to GM1 are then measured in a single experiment from 50 channels. By combining the highly parallel microfluidic device with large-area periodic nanohole array chips, our SPR imaging spectrometer system enables high-throughput, label-free, real-time SPR biosensing, and its full-spectral imaging capability combined with nanohole arrays could enable integration of SPR imaging with concurrent surface-enhanced Raman spectroscopy. PMID:22895607

High-throughput microsphiltration to assess red blood cell deformability and screen for malaria transmission-blocking drugs.

PubMed

Duez, Julien; Carucci, Mario; Garcia-Barbazan, Irene; Corral, Matias; Perez, Oscar; Presa, Jesus Luis; Henry, Benoit; Roussel, Camille; Ndour, Papa Alioune; Rosa, Noemi Bahamontes; Sanz, Laura; Gamo, Francisco-Javier; Buffet, Pierre

2018-06-01

The mechanical retention of rigid erythrocytes in the spleen is central in major hematological diseases such as hereditary spherocytosis, sickle-cell disease and malaria. Here, we describe the use of microsphiltration (microsphere filtration) to assess erythrocyte deformability in hundreds to thousands of samples in parallel, by filtering them through microsphere layers in 384-well plates adapted for the discovery of compounds that stiffen Plasmodium falciparum gametocytes, with the aim of interrupting malaria transmission. Compound-exposed gametocytes are loaded into microsphiltration plates, filtered and then transferred to imaging plates for analysis. High-content imaging detects viable gametocytes upstream and downstream from filters and quantifies spleen-like retention. This screening assay takes 3-4 d. Unlike currently available methods used to assess red blood cell (RBC) deformability, microsphiltration enables high-throughput pharmacological screening (tens of thousands of compounds tested in a matter of months) and involves a cell mechanical challenge that induces a physiologically relevant dumbbell-shape deformation. It therefore directly assesses the ability of RBCs to cross inter-endothelial splenic slits in vivo. This protocol has potential applications in quality control for transfusion and in determination of phenotypic markers of erythrocytes in hematological diseases.

Quantitative imaging features: extension of the oncology medical image database

NASA Astrophysics Data System (ADS)

Patel, M. N.; Looney, P. T.; Young, K. C.; Halling-Brown, M. D.

2015-03-01

Radiological imaging is fundamental within the healthcare industry and has become routinely adopted for diagnosis, disease monitoring and treatment planning. With the advent of digital imaging modalities and the rapid growth in both diagnostic and therapeutic imaging, the ability to be able to harness this large influx of data is of paramount importance. The Oncology Medical Image Database (OMI-DB) was created to provide a centralized, fully annotated dataset for research. The database contains both processed and unprocessed images, associated data, and annotations and where applicable expert determined ground truths describing features of interest. Medical imaging provides the ability to detect and localize many changes that are important to determine whether a disease is present or a therapy is effective by depicting alterations in anatomic, physiologic, biochemical or molecular processes. Quantitative imaging features are sensitive, specific, accurate and reproducible imaging measures of these changes. Here, we describe an extension to the OMI-DB whereby a range of imaging features and descriptors are pre-calculated using a high throughput approach. The ability to calculate multiple imaging features and data from the acquired images would be valuable and facilitate further research applications investigating detection, prognosis, and classification. The resultant data store contains more than 10 million quantitative features as well as features derived from CAD predictions. Theses data can be used to build predictive models to aid image classification, treatment response assessment as well as to identify prognostic imaging biomarkers.

Cytopathological image analysis using deep-learning networks in microfluidic microscopy.

PubMed

Gopakumar, G; Hari Babu, K; Mishra, Deepak; Gorthi, Sai Siva; Sai Subrahmanyam, Gorthi R K

2017-01-01

Cytopathologic testing is one of the most critical steps in the diagnosis of diseases, including cancer. However, the task is laborious and demands skill. Associated high cost and low throughput drew considerable interest in automating the testing process. Several neural network architectures were designed to provide human expertise to machines. In this paper, we explore and propose the feasibility of using deep-learning networks for cytopathologic analysis by performing the classification of three important unlabeled, unstained leukemia cell lines (K562, MOLT, and HL60). The cell images used in the classification are captured using a low-cost, high-throughput cell imaging technique: microfluidics-based imaging flow cytometry. We demonstrate that without any conventional fine segmentation followed by explicit feature extraction, the proposed deep-learning algorithms effectively classify the coarsely localized cell lines. We show that the designed deep belief network as well as the deeply pretrained convolutional neural network outperform the conventionally used decision systems and are important in the medical domain, where the availability of labeled data is limited for training. We hope that our work enables the development of a clinically significant high-throughput microfluidic microscopy-based tool for disease screening/triaging, especially in resource-limited settings.

Real-time traffic sign detection and recognition

NASA Astrophysics Data System (ADS)

Herbschleb, Ernst; de With, Peter H. N.

2009-01-01

The continuous growth of imaging databases increasingly requires analysis tools for extraction of features. In this paper, a new architecture for the detection of traffic signs is proposed. The architecture is designed to process a large database with tens of millions of images with a resolution up to 4,800x2,400 pixels. Because of the size of the database, a high reliability as well as a high throughput is required. The novel architecture consists of a three-stage algorithm with multiple steps per stage, combining both color and specific spatial information. The first stage contains an area-limitation step which is performance critical in both the detection rate as the overall processing time. The second stage locates suggestions for traffic signs using recently published feature processing. The third stage contains a validation step to enhance reliability of the algorithm. During this stage, the traffic signs are recognized. Experiments show a convincing detection rate of 99%. With respect to computational speed, the throughput for line-of-sight images of 800×600 pixels is 35 Hz and for panorama images it is 4 Hz. Our novel architecture outperforms existing algorithms, with respect to both detection rate and throughput

Clinical and laboratory applications of slide-based cytometry with the LSC, SFM, and the iCYTE imaging cytometer instruments

NASA Astrophysics Data System (ADS)

Bocsi, Jozsef; Luther, Ed; Mittag, Anja; Jensen, Ingo; Sack, Ulrich; Lenz, Dominik; Trezl, Lajos; Varga, Viktor S.; Molnar, Beea; Tarnok, Attila

2004-06-01

Background: Slide based cytometry (SBC) is a technology for the rapid stoichiometric analysis of cells fixed to surfaces. Its applications are highly versatile and ranges from the clinics to high throughput drug discovery. SBC is realized in different instruments such as the Laser Scanning Cytometer (LSC) and Scanning Fluorescent Microscope (SFM) and the novel inverted microscope based iCyte image cytometer (Compucyte Corp.). Methods: Fluorochrome labeled specimens were immobilized on microscopic slides. They were placed on a conventional fluorescence microscope and analyzed by photomultiplayers or digital camera. Data comparable to flow cytometry were generated. In addition, each individual event could be visualized. Applications: The major advantage of instruments is the combination of two features: a) the minimal sample volume needed, and b) the connection of fluorescence data and morphological information. Rare cells were detected, frequency of apoptosis by myricetin formaldehyde and H2O2 mixtures was determined;. Conclusion: LSC, SFM and the novel iCyte have a wide spectrum of applicability in SBC and can be introduced as a standard technology for multiple settings. In addition, the iCyte and SFM instrument is suited for high throughput screening by automation and may be in future adapted to telepathology due to their high quality images. (This study was supported by the IZKF-Leipzig, Germany and T 034245 OTKA, Hungary)

High-throughput flow alignment of barcoded hydrogel microparticles†

PubMed Central

Chapin, Stephen C.; Pregibon, Daniel C.

2010-01-01

Suspension (particle-based) arrays offer several advantages over conventional planar arrays in the detection and quantification of biomolecules, including the use of smaller sample volumes, more favorable probe-target binding kinetics, and rapid probe-set modification. We present a microfluidic system for the rapid alignment of multifunctional hydrogel microparticles designed to bear one or several biomolecule probe regions, as well as a graphical code to identify the embedded probes. Using high-speed imaging, we have developed and optimized a flow-through system that (1) allows for a high particle throughput, (2) ensures proper particle alignment for decoding and target quantification, and (3) can be reliably operated continuously without clogging. A tapered channel flanked by side focusing streams is used to orient the flexible, tablet-shaped particles into a well-ordered flow in the center of the channel. The effects of channel geometry, particle geometry, particle composition, particle loading density, and barcode design are explored to determine the best combination for eventual use in biological assays. Particles in the optimized system move at velocities of ~50 cm s−1 and with throughputs of ~40 particles s−1. Simple physical models and CFD simulations have been used to investigate flow behavior in the device. PMID:19823726

Polarization-resolved second-harmonic generation microscopy as a method to visualize protein-crystal domains

PubMed Central

DeWalt, Emma L.; Begue, Victoria J.; Ronau, Judith A.; Sullivan, Shane Z.; Das, Chittaranjan; Simpson, Garth J.

2013-01-01

Polarization-resolved second-harmonic generation (PR-SHG) microscopy is described and applied to identify the presence of multiple crystallographic domains within protein-crystal conglomerates, which was confirmed by synchrotron X-ray diffraction. Principal component analysis (PCA) of PR-SHG images resulted in principal component 2 (PC2) images with areas of contrasting negative and positive values for conglomerated crystals and PC2 images exhibiting uniformly positive or uniformly negative values for single crystals. Qualitative assessment of PC2 images allowed the identification of domains of different internal ordering within protein-crystal samples as well as differentiation between multi-domain conglomerated crystals and single crystals. PR-SHG assessments of crystalline domains were in good agreement with spatially resolved synchrotron X-ray diffraction measurements. These results have implications for improving the productive throughput of protein structure determination through early identification of multi-domain crystals. PMID:23275165

Use of a Fluorometric Imaging Plate Reader in high-throughput screening

NASA Astrophysics Data System (ADS)

Groebe, Duncan R.; Gopalakrishnan, Sujatha; Hahn, Holly; Warrior, Usha; Traphagen, Linda; Burns, David J.

1999-04-01

High-throughput screening (HTS) efforts at Abbott Laboratories have been greatly facilitated by the use of a Fluorometric Imaging Plate Reader. The FLIPR consists of an incubated cabinet with integrated 96-channel pipettor and fluorometer. An argon laser is used to excite fluorophores in a 96-well microtiter plate and the emitted fluorometer. An argon laser is used to excite fluorophores in a 96-well microtiter plate and the emitted fluorescence is imaged by a cooled CCD camera. The image data is downloaded from the camera and processed to average the signal form each well of the microtiter pate for each time point. The data is presented in real time on the computer screen, facilitating interpretation and trouble-shooting. In addition to fluorescence, the camera can also detect luminescence form firefly luciferase.

Determining the distribution of probes between different subcellular locations through automated unmixing of subcellular patterns.

PubMed

Peng, Tao; Bonamy, Ghislain M C; Glory-Afshar, Estelle; Rines, Daniel R; Chanda, Sumit K; Murphy, Robert F

2010-02-16

Many proteins or other biological macromolecules are localized to more than one subcellular structure. The fraction of a protein in different cellular compartments is often measured by colocalization with organelle-specific fluorescent markers, requiring availability of fluorescent probes for each compartment and acquisition of images for each in conjunction with the macromolecule of interest. Alternatively, tailored algorithms allow finding particular regions in images and quantifying the amount of fluorescence they contain. Unfortunately, this approach requires extensive hand-tuning of algorithms and is often cell type-dependent. Here we describe a machine-learning approach for estimating the amount of fluorescent signal in different subcellular compartments without hand tuning, requiring only the acquisition of separate training images of markers for each compartment. In testing on images of cells stained with mixtures of probes for different organelles, we achieved a 93% correlation between estimated and expected amounts of probes in each compartment. We also demonstrated that the method can be used to quantify drug-dependent protein translocations. The method enables automated and unbiased determination of the distributions of protein across cellular compartments, and will significantly improve imaging-based high-throughput assays and facilitate proteome-scale localization efforts.

Adapting the γ-H2AX assay for automated processing in human lymphocytes. 1. Technological aspects.

PubMed

Turner, Helen C; Brenner, David J; Chen, Youhua; Bertucci, Antonella; Zhang, Jian; Wang, Hongliang; Lyulko, Oleksandra V; Xu, Yanping; Shuryak, Igor; Schaefer, Julia; Simaan, Nabil; Randers-Pehrson, Gerhard; Yao, Y Lawrence; Amundson, Sally A; Garty, Guy

2011-03-01

The immunofluorescence-based detection of γ-H2AX is a reliable and sensitive method for quantitatively measuring DNA double-strand breaks (DSBs) in irradiated samples. Since H2AX phosphorylation is highly linear with radiation dose, this well-established biomarker is in current use in radiation biodosimetry. At the Center for High-Throughput Minimally Invasive Radiation Biodosimetry, we have developed a fully automated high-throughput system, the RABIT (Rapid Automated Biodosimetry Tool), that can be used to measure γ-H2AX yields from fingerstick-derived samples of blood. The RABIT workstation has been designed to fully automate the γ-H2AX immunocytochemical protocol, from the isolation of human blood lymphocytes in heparin-coated PVC capillaries to the immunolabeling of γ-H2AX protein and image acquisition to determine fluorescence yield. High throughput is achieved through the use of purpose-built robotics, lymphocyte handling in 96-well filter-bottomed plates, and high-speed imaging. The goal of the present study was to optimize and validate the performance of the RABIT system for the reproducible and quantitative detection of γ-H2AX total fluorescence in lymphocytes in a multiwell format. Validation of our biodosimetry platform was achieved by the linear detection of a dose-dependent increase in γ-H2AX fluorescence in peripheral blood samples irradiated ex vivo with γ rays over the range 0 to 8 Gy. This study demonstrates for the first time the optimization and use of our robotically based biodosimetry workstation to successfully quantify γ-H2AX total fluorescence in irradiated peripheral lymphocytes.

Automated image-based phenotypic analysis in zebrafish embryos

PubMed Central

Vogt, Andreas; Cholewinski, Andrzej; Shen, Xiaoqiang; Nelson, Scott; Lazo, John S.; Tsang, Michael; Hukriede, Neil A.

2009-01-01

Presently, the zebrafish is the only vertebrate model compatible with contemporary paradigms of drug discovery. Zebrafish embryos are amenable to automation necessary for high-throughput chemical screens, and optical transparency makes them potentially suited for image-based screening. However, the lack of tools for automated analysis of complex images presents an obstacle to utilizing the zebrafish as a high-throughput screening model. We have developed an automated system for imaging and analyzing zebrafish embryos in multi-well plates regardless of embryo orientation and without user intervention. Images of fluorescent embryos were acquired on a high-content reader and analyzed using an artificial intelligence-based image analysis method termed Cognition Network Technology (CNT). CNT reliably detected transgenic fluorescent embryos (Tg(fli1:EGFP)y1) arrayed in 96-well plates and quantified intersegmental blood vessel development in embryos treated with small molecule inhibitors of anigiogenesis. The results demonstrate it is feasible to adapt image-based high-content screening methodology to measure complex whole organism phenotypes. PMID:19235725

Comparison of Human Induced Pluripotent Stem Cell-Derived Neurons and Rat Primary CorticalNeurons as In Vitro Models of Neurite Outgrowth

EPA Science Inventory

High-throughput assays that can quantify chemical-induced changes at the cellular and molecular level have been recommended for use in chemical safety assessment. High-throughput, high content imaging assays for the key cellular events of neurodevelopment have been proposed to ra...

Achieving High Throughput for Data Transfer over ATM Networks

NASA Technical Reports Server (NTRS)

Johnson, Marjory J.; Townsend, Jeffrey N.

1996-01-01

File-transfer rates for ftp are often reported to be relatively slow, compared to the raw bandwidth available in emerging gigabit networks. While a major bottleneck is disk I/O, protocol issues impact performance as well. Ftp was developed and optimized for use over the TCP/IP protocol stack of the Internet. However, TCP has been shown to run inefficiently over ATM. In an effort to maximize network throughput, data-transfer protocols can be developed to run over UDP or directly over IP, rather than over TCP. If error-free transmission is required, techniques for achieving reliable transmission can be included as part of the transfer protocol. However, selected image-processing applications can tolerate a low level of errors in images that are transmitted over a network. In this paper we report on experimental work to develop a high-throughput protocol for unreliable data transfer over ATM networks. We attempt to maximize throughput by keeping the communications pipe full, but still keep packet loss under five percent. We use the Bay Area Gigabit Network Testbed as our experimental platform.

Pipeline for illumination correction of images for high-throughput microscopy.

PubMed

Singh, S; Bray, M-A; Jones, T R; Carpenter, A E

2014-12-01

The presence of systematic noise in images in high-throughput microscopy experiments can significantly impact the accuracy of downstream results. Among the most common sources of systematic noise is non-homogeneous illumination across the image field. This often adds an unacceptable level of noise, obscures true quantitative differences and precludes biological experiments that rely on accurate fluorescence intensity measurements. In this paper, we seek to quantify the improvement in the quality of high-content screen readouts due to software-based illumination correction. We present a straightforward illumination correction pipeline that has been used by our group across many experiments. We test the pipeline on real-world high-throughput image sets and evaluate the performance of the pipeline at two levels: (a) Z'-factor to evaluate the effect of the image correction on a univariate readout, representative of a typical high-content screen, and (b) classification accuracy on phenotypic signatures derived from the images, representative of an experiment involving more complex data mining. We find that applying the proposed post-hoc correction method improves performance in both experiments, even when illumination correction has already been applied using software associated with the instrument. To facilitate the ready application and future development of illumination correction methods, we have made our complete test data sets as well as open-source image analysis pipelines publicly available. This software-based solution has the potential to improve outcomes for a wide-variety of image-based HTS experiments. © 2014 The Authors. Journal of Microscopy published by John Wiley & Sons Ltd on behalf of Royal Microscopical Society.

High-speed cell recognition algorithm for ultrafast flow cytometer imaging system.

PubMed

Zhao, Wanyue; Wang, Chao; Chen, Hongwei; Chen, Minghua; Yang, Sigang

2018-04-01

An optical time-stretch flow imaging system enables high-throughput examination of cells/particles with unprecedented high speed and resolution. A significant amount of raw image data is produced. A high-speed cell recognition algorithm is, therefore, highly demanded to analyze large amounts of data efficiently. A high-speed cell recognition algorithm consisting of two-stage cascaded detection and Gaussian mixture model (GMM) classification is proposed. The first stage of detection extracts cell regions. The second stage integrates distance transform and the watershed algorithm to separate clustered cells. Finally, the cells detected are classified by GMM. We compared the performance of our algorithm with support vector machine. Results show that our algorithm increases the running speed by over 150% without sacrificing the recognition accuracy. This algorithm provides a promising solution for high-throughput and automated cell imaging and classification in the ultrafast flow cytometer imaging platform. (2018) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE).

High-speed cell recognition algorithm for ultrafast flow cytometer imaging system

NASA Astrophysics Data System (ADS)

Zhao, Wanyue; Wang, Chao; Chen, Hongwei; Chen, Minghua; Yang, Sigang

2018-04-01

An optical time-stretch flow imaging system enables high-throughput examination of cells/particles with unprecedented high speed and resolution. A significant amount of raw image data is produced. A high-speed cell recognition algorithm is, therefore, highly demanded to analyze large amounts of data efficiently. A high-speed cell recognition algorithm consisting of two-stage cascaded detection and Gaussian mixture model (GMM) classification is proposed. The first stage of detection extracts cell regions. The second stage integrates distance transform and the watershed algorithm to separate clustered cells. Finally, the cells detected are classified by GMM. We compared the performance of our algorithm with support vector machine. Results show that our algorithm increases the running speed by over 150% without sacrificing the recognition accuracy. This algorithm provides a promising solution for high-throughput and automated cell imaging and classification in the ultrafast flow cytometer imaging platform.

Cellular resolution functional imaging in behaving rats using voluntary head restraint

PubMed Central

Scott, Benjamin B.; Brody, Carlos D.; Tank, David W.

2013-01-01

SUMMARY High-throughput operant conditioning systems for rodents provide efficient training on sophisticated behavioral tasks. Combining these systems with technologies for cellular resolution functional imaging would provide a powerful approach to study neural dynamics during behavior. Here we describe an integrated two-photon microscope and behavioral apparatus that allows cellular resolution functional imaging of cortical regions during epochs of voluntary head restraint. Rats were trained to initiate periods of restraint up to 8 seconds in duration, which provided the mechanical stability necessary for in vivo imaging while allowing free movement between behavioral trials. A mechanical registration system repositioned the head to within a few microns, allowing the same neuronal populations to be imaged on each trial. In proof-of-principle experiments, calcium dependent fluorescence transients were recorded from GCaMP-labeled cortical neurons. In contrast to previous methods for head restraint, this system can also be incorporated into high-throughput operant conditioning systems. PMID:24055015

SmartGrain: high-throughput phenotyping software for measuring seed shape through image analysis.

PubMed

Tanabata, Takanari; Shibaya, Taeko; Hori, Kiyosumi; Ebana, Kaworu; Yano, Masahiro

2012-12-01

Seed shape and size are among the most important agronomic traits because they affect yield and market price. To obtain accurate seed size data, a large number of measurements are needed because there is little difference in size among seeds from one plant. To promote genetic analysis and selection for seed shape in plant breeding, efficient, reliable, high-throughput seed phenotyping methods are required. We developed SmartGrain software for high-throughput measurement of seed shape. This software uses a new image analysis method to reduce the time taken in the preparation of seeds and in image capture. Outlines of seeds are automatically recognized from digital images, and several shape parameters, such as seed length, width, area, and perimeter length, are calculated. To validate the software, we performed a quantitative trait locus (QTL) analysis for rice (Oryza sativa) seed shape using backcrossed inbred lines derived from a cross between japonica cultivars Koshihikari and Nipponbare, which showed small differences in seed shape. SmartGrain removed areas of awns and pedicels automatically, and several QTLs were detected for six shape parameters. The allelic effect of a QTL for seed length detected on chromosome 11 was confirmed in advanced backcross progeny; the cv Nipponbare allele increased seed length and, thus, seed weight. High-throughput measurement with SmartGrain reduced sampling error and made it possible to distinguish between lines with small differences in seed shape. SmartGrain could accurately recognize seed not only of rice but also of several other species, including Arabidopsis (Arabidopsis thaliana). The software is free to researchers.

A Versatile Cell Death Screening Assay Using Dye-Stained Cells and Multivariate Image Analysis.

PubMed

Collins, Tony J; Ylanko, Jarkko; Geng, Fei; Andrews, David W

2015-11-01

A novel dye-based method for measuring cell death in image-based screens is presented. Unlike conventional high- and medium-throughput cell death assays that measure only one form of cell death accurately, using multivariate analysis of micrographs of cells stained with the inexpensive mix, red dye nonyl acridine orange, and a nuclear stain, it was possible to quantify cell death induced by a variety of different agonists even without a positive control. Surprisingly, using a single known cytotoxic agent as a positive control for training a multivariate classifier allowed accurate quantification of cytotoxicity for mechanistically unrelated compounds enabling generation of dose-response curves. Comparison with low throughput biochemical methods suggested that cell death was accurately distinguished from cell stress induced by low concentrations of the bioactive compounds Tunicamycin and Brefeldin A. High-throughput image-based format analyses of more than 300 kinase inhibitors correctly identified 11 as cytotoxic with only 1 false positive. The simplicity and robustness of this dye-based assay makes it particularly suited to live cell screening for toxic compounds.

A Versatile Cell Death Screening Assay Using Dye-Stained Cells and Multivariate Image Analysis

PubMed Central

Collins, Tony J.; Ylanko, Jarkko; Geng, Fei

2015-01-01

Abstract A novel dye-based method for measuring cell death in image-based screens is presented. Unlike conventional high- and medium-throughput cell death assays that measure only one form of cell death accurately, using multivariate analysis of micrographs of cells stained with the inexpensive mix, red dye nonyl acridine orange, and a nuclear stain, it was possible to quantify cell death induced by a variety of different agonists even without a positive control. Surprisingly, using a single known cytotoxic agent as a positive control for training a multivariate classifier allowed accurate quantification of cytotoxicity for mechanistically unrelated compounds enabling generation of dose–response curves. Comparison with low throughput biochemical methods suggested that cell death was accurately distinguished from cell stress induced by low concentrations of the bioactive compounds Tunicamycin and Brefeldin A. High-throughput image-based format analyses of more than 300 kinase inhibitors correctly identified 11 as cytotoxic with only 1 false positive. The simplicity and robustness of this dye-based assay makes it particularly suited to live cell screening for toxic compounds. PMID:26422066

A compact imaging spectroscopic system for biomolecular detections on plasmonic chips.

PubMed

Lo, Shu-Cheng; Lin, En-Hung; Wei, Pei-Kuen; Tsai, Wan-Shao

2016-10-17

In this study, we demonstrate a compact imaging spectroscopic system for high-throughput detection of biomolecular interactions on plasmonic chips, based on a curved grating as the key element of light diffraction and light focusing. Both the curved grating and the plasmonic chips are fabricated on flexible plastic substrates using a gas-assisted thermal-embossing method. A fiber-coupled broadband light source and a camera are included in the system. Spectral resolution within 1 nm is achieved in sensing environmental index solutions and protein bindings. The detected sensitivities of the plasmonic chip are comparable with a commercial spectrometer. An extra one-dimensional scanning stage enables high-throughput detection of protein binding on a designed plasmonic chip consisting of several nanoslit arrays with different periods. The detected resonance wavelengths match well with the grating equation under an air environment. Wavelength shifts between 1 and 9 nm are detected for antigens of various concentrations binding with antibodies. A simple, mass-productive and cost-effective method has been demonstrated on the imaging spectroscopic system for real-time, label-free, highly sensitive and high-throughput screening of biomolecular interactions.

High Throughput In vivo Analysis of Plant Leaf Chemical Properties Using Hyperspectral Imaging

PubMed Central

Pandey, Piyush; Ge, Yufeng; Stoerger, Vincent; Schnable, James C.

2017-01-01

Image-based high-throughput plant phenotyping in greenhouse has the potential to relieve the bottleneck currently presented by phenotypic scoring which limits the throughput of gene discovery and crop improvement efforts. Numerous studies have employed automated RGB imaging to characterize biomass and growth of agronomically important crops. The objective of this study was to investigate the utility of hyperspectral imaging for quantifying chemical properties of maize and soybean plants in vivo. These properties included leaf water content, as well as concentrations of macronutrients nitrogen (N), phosphorus (P), potassium (K), magnesium (Mg), calcium (Ca), and sulfur (S), and micronutrients sodium (Na), iron (Fe), manganese (Mn), boron (B), copper (Cu), and zinc (Zn). Hyperspectral images were collected from 60 maize and 60 soybean plants, each subjected to varying levels of either water deficit or nutrient limitation stress with the goal of creating a wide range of variation in the chemical properties of plant leaves. Plants were imaged on an automated conveyor belt system using a hyperspectral imager with a spectral range from 550 to 1,700 nm. Images were processed to extract reflectance spectrum from each plant and partial least squares regression models were developed to correlate spectral data with chemical data. Among all the chemical properties investigated, water content was predicted with the highest accuracy [R2 = 0.93 and RPD (Ratio of Performance to Deviation) = 3.8]. All macronutrients were also quantified satisfactorily (R2 from 0.69 to 0.92, RPD from 1.62 to 3.62), with N predicted best followed by P, K, and S. The micronutrients group showed lower prediction accuracy (R2 from 0.19 to 0.86, RPD from 1.09 to 2.69) than the macronutrient groups. Cu and Zn were best predicted, followed by Fe and Mn. Na and B were the only two properties that hyperspectral imaging was not able to quantify satisfactorily (R2 < 0.3 and RPD < 1.2). This study suggested the potential usefulness of hyperspectral imaging as a high-throughput phenotyping technology for plant chemical traits. Future research is needed to test the method more thoroughly by designing experiments to vary plant nutrients individually and cover more plant species, genotypes, and growth stages. PMID:28824683

High Throughput In vivo Analysis of Plant Leaf Chemical Properties Using Hyperspectral Imaging.

PubMed

Pandey, Piyush; Ge, Yufeng; Stoerger, Vincent; Schnable, James C

2017-01-01

Image-based high-throughput plant phenotyping in greenhouse has the potential to relieve the bottleneck currently presented by phenotypic scoring which limits the throughput of gene discovery and crop improvement efforts. Numerous studies have employed automated RGB imaging to characterize biomass and growth of agronomically important crops. The objective of this study was to investigate the utility of hyperspectral imaging for quantifying chemical properties of maize and soybean plants in vivo . These properties included leaf water content, as well as concentrations of macronutrients nitrogen (N), phosphorus (P), potassium (K), magnesium (Mg), calcium (Ca), and sulfur (S), and micronutrients sodium (Na), iron (Fe), manganese (Mn), boron (B), copper (Cu), and zinc (Zn). Hyperspectral images were collected from 60 maize and 60 soybean plants, each subjected to varying levels of either water deficit or nutrient limitation stress with the goal of creating a wide range of variation in the chemical properties of plant leaves. Plants were imaged on an automated conveyor belt system using a hyperspectral imager with a spectral range from 550 to 1,700 nm. Images were processed to extract reflectance spectrum from each plant and partial least squares regression models were developed to correlate spectral data with chemical data. Among all the chemical properties investigated, water content was predicted with the highest accuracy [ R 2 = 0.93 and RPD (Ratio of Performance to Deviation) = 3.8]. All macronutrients were also quantified satisfactorily ( R 2 from 0.69 to 0.92, RPD from 1.62 to 3.62), with N predicted best followed by P, K, and S. The micronutrients group showed lower prediction accuracy ( R 2 from 0.19 to 0.86, RPD from 1.09 to 2.69) than the macronutrient groups. Cu and Zn were best predicted, followed by Fe and Mn. Na and B were the only two properties that hyperspectral imaging was not able to quantify satisfactorily ( R 2 < 0.3 and RPD < 1.2). This study suggested the potential usefulness of hyperspectral imaging as a high-throughput phenotyping technology for plant chemical traits. Future research is needed to test the method more thoroughly by designing experiments to vary plant nutrients individually and cover more plant species, genotypes, and growth stages.

Snapshot imaging Fraunhofer line discriminator for detection of plant fluorescence

NASA Astrophysics Data System (ADS)

Gupta Roy, S.; Kudenov, M. W.

2015-05-01

Non-invasive quantification of plant health is traditionally accomplished using reflectance based metrics, such as the normalized difference vegetative index (NDVI). However, measuring plant fluorescence (both active and passive) to determine photochemistry of plants has gained importance. Due to better cost efficiency, lower power requirements, and simpler scanning synchronization, detecting passive fluorescence is preferred over active fluorescence. In this paper, we propose a high speed imaging approach for measuring passive plant fluorescence, within the hydrogen alpha Fraunhofer line at ~656 nm, using a Snapshot Imaging Fraunhofer Line Discriminator (SIFOLD). For the first time, the advantage of snapshot imaging for high throughput Fraunhofer Line Discrimination (FLD) is cultivated by our system, which is based on a multiple-image Fourier transform spectrometer and a spatial heterodyne interferometer (SHI). The SHI is a Sagnac interferometer, which is dispersion compensated using blazed diffraction gratings. We present data and techniques for calibrating the SIFOLD to any particular wavelength. This technique can be applied to quantify plant fluorescence at low cost and reduced complexity of data collection.

Image-based computational quantification and visualization of genetic alterations and tumour heterogeneity

PubMed Central

Zhong, Qing; Rüschoff, Jan H.; Guo, Tiannan; Gabrani, Maria; Schüffler, Peter J.; Rechsteiner, Markus; Liu, Yansheng; Fuchs, Thomas J.; Rupp, Niels J.; Fankhauser, Christian; Buhmann, Joachim M.; Perner, Sven; Poyet, Cédric; Blattner, Miriam; Soldini, Davide; Moch, Holger; Rubin, Mark A.; Noske, Aurelia; Rüschoff, Josef; Haffner, Michael C.; Jochum, Wolfram; Wild, Peter J.

2016-01-01

Recent large-scale genome analyses of human tissue samples have uncovered a high degree of genetic alterations and tumour heterogeneity in most tumour entities, independent of morphological phenotypes and histopathological characteristics. Assessment of genetic copy-number variation (CNV) and tumour heterogeneity by fluorescence in situ hybridization (ISH) provides additional tissue morphology at single-cell resolution, but it is labour intensive with limited throughput and high inter-observer variability. We present an integrative method combining bright-field dual-colour chromogenic and silver ISH assays with an image-based computational workflow (ISHProfiler), for accurate detection of molecular signals, high-throughput evaluation of CNV, expressive visualization of multi-level heterogeneity (cellular, inter- and intra-tumour heterogeneity), and objective quantification of heterogeneous genetic deletions (PTEN) and amplifications (19q12, HER2) in diverse human tumours (prostate, endometrial, ovarian and gastric), using various tissue sizes and different scanners, with unprecedented throughput and reproducibility. PMID:27052161

Image-based computational quantification and visualization of genetic alterations and tumour heterogeneity.

PubMed

Zhong, Qing; Rüschoff, Jan H; Guo, Tiannan; Gabrani, Maria; Schüffler, Peter J; Rechsteiner, Markus; Liu, Yansheng; Fuchs, Thomas J; Rupp, Niels J; Fankhauser, Christian; Buhmann, Joachim M; Perner, Sven; Poyet, Cédric; Blattner, Miriam; Soldini, Davide; Moch, Holger; Rubin, Mark A; Noske, Aurelia; Rüschoff, Josef; Haffner, Michael C; Jochum, Wolfram; Wild, Peter J

2016-04-07

Recent large-scale genome analyses of human tissue samples have uncovered a high degree of genetic alterations and tumour heterogeneity in most tumour entities, independent of morphological phenotypes and histopathological characteristics. Assessment of genetic copy-number variation (CNV) and tumour heterogeneity by fluorescence in situ hybridization (ISH) provides additional tissue morphology at single-cell resolution, but it is labour intensive with limited throughput and high inter-observer variability. We present an integrative method combining bright-field dual-colour chromogenic and silver ISH assays with an image-based computational workflow (ISHProfiler), for accurate detection of molecular signals, high-throughput evaluation of CNV, expressive visualization of multi-level heterogeneity (cellular, inter- and intra-tumour heterogeneity), and objective quantification of heterogeneous genetic deletions (PTEN) and amplifications (19q12, HER2) in diverse human tumours (prostate, endometrial, ovarian and gastric), using various tissue sizes and different scanners, with unprecedented throughput and reproducibility.

High-throughput ultraviolet photoacoustic microscopy with multifocal excitation

NASA Astrophysics Data System (ADS)

Imai, Toru; Shi, Junhui; Wong, Terence T. W.; Li, Lei; Zhu, Liren; Wang, Lihong V.

2018-03-01

Ultraviolet photoacoustic microscopy (UV-PAM) is a promising intraoperative tool for surgical margin assessment (SMA), one that can provide label-free histology-like images with high resolution. In this study, using a microlens array and a one-dimensional (1-D) array ultrasonic transducer, we developed a high-throughput multifocal UV-PAM (MF-UV-PAM). Our new system achieved a 1.6 ± 0.2 μm lateral resolution and produced images 40 times faster than the previously developed point-by-point scanning UV-PAM. MF-UV-PAM provided a readily comprehensible photoacoustic image of a mouse brain slice with specific absorption contrast in ˜16 min, highlighting cell nuclei. Individual cell nuclei could be clearly resolved, showing its practical potential for intraoperative SMA.

Quantitative image analysis of immunohistochemical stains using a CMYK color model

PubMed Central

Pham, Nhu-An; Morrison, Andrew; Schwock, Joerg; Aviel-Ronen, Sarit; Iakovlev, Vladimir; Tsao, Ming-Sound; Ho, James; Hedley, David W

2007-01-01

Background Computer image analysis techniques have decreased effects of observer biases, and increased the sensitivity and the throughput of immunohistochemistry (IHC) as a tissue-based procedure for the evaluation of diseases. Methods We adapted a Cyan/Magenta/Yellow/Key (CMYK) model for automated computer image analysis to quantify IHC stains in hematoxylin counterstained histological sections. Results The spectral characteristics of the chromogens AEC, DAB and NovaRed as well as the counterstain hematoxylin were first determined using CMYK, Red/Green/Blue (RGB), normalized RGB and Hue/Saturation/Lightness (HSL) color models. The contrast of chromogen intensities on a 0–255 scale (24-bit image file) as well as compared to the hematoxylin counterstain was greatest using the Yellow channel of a CMYK color model, suggesting an improved sensitivity for IHC evaluation compared to other color models. An increase in activated STAT3 levels due to growth factor stimulation, quantified using the Yellow channel image analysis was associated with an increase detected by Western blotting. Two clinical image data sets were used to compare the Yellow channel automated method with observer-dependent methods. First, a quantification of DAB-labeled carbonic anhydrase IX hypoxia marker in 414 sections obtained from 138 biopsies of cervical carcinoma showed strong association between Yellow channel and positive color selection results. Second, a linear relationship was also demonstrated between Yellow intensity and visual scoring for NovaRed-labeled epidermal growth factor receptor in 256 non-small cell lung cancer biopsies. Conclusion The Yellow channel image analysis method based on a CMYK color model is independent of observer biases for threshold and positive color selection, applicable to different chromogens, tolerant of hematoxylin, sensitive to small changes in IHC intensity and is applicable to simple automation procedures. These characteristics are advantageous for both basic as well as clinical research in an unbiased, reproducible and high throughput evaluation of IHC intensity. PMID:17326824

A simple and sensitive high-throughput GFP screening in woody and herbaceous plants.

PubMed

Hily, Jean-Michel; Liu, Zongrang

2009-03-01

Green fluorescent protein (GFP) has been used widely as a powerful bioluminescent reporter, but its visualization by existing methods in tissues or whole plants and its utilization for high-throughput screening remains challenging in many species. Here, we report a fluorescence image analyzer-based method for GFP detection and its utility for high-throughput screening of transformed plants. Of three detection methods tested, the Typhoon fluorescence scanner was able to detect GFP fluorescence in all Arabidopsis thaliana tissues and apple leaves, while regular fluorescence microscopy detected it only in Arabidopsis flowers and siliques but barely in the leaves of either Arabidopsis or apple. The hand-held UV illumination method failed in all tissues of both species. Additionally, the Typhoon imager was able to detect GFP fluorescence in both green and non-green tissues of Arabidopsis seedlings as well as in imbibed seeds, qualifying it as a high-throughput screening tool, which was further demonstrated by screening the seedlings of primary transformed T(0) seeds. Of the 30,000 germinating Arabidopsis seedlings screened, at least 69 GFP-positive lines were identified, accounting for an approximately 0.23% transformation efficiency. About 14,000 seedlings grown in 16 Petri plates could be screened within an hour, making the screening process significantly more efficient and robust than any other existing high-throughput screening method for transgenic plants.

Overview of machine vision methods in x-ray imaging and microtomography

NASA Astrophysics Data System (ADS)

Buzmakov, Alexey; Zolotov, Denis; Chukalina, Marina; Nikolaev, Dmitry; Gladkov, Andrey; Ingacheva, Anastasia; Yakimchuk, Ivan; Asadchikov, Victor

2018-04-01

Digital X-ray imaging became widely used in science, medicine, non-destructive testing. This allows using modern digital images analysis for automatic information extraction and interpretation. We give short review of scientific applications of machine vision in scientific X-ray imaging and microtomography, including image processing, feature detection and extraction, images compression to increase camera throughput, microtomography reconstruction, visualization and setup adjustment.

A high-throughput in vitro ring assay for vasoactivity using magnetic 3D bioprinting

PubMed Central

Tseng, Hubert; Gage, Jacob A.; Haisler, William L.; Neeley, Shane K.; Shen, Tsaiwei; Hebel, Chris; Barthlow, Herbert G.; Wagoner, Matthew; Souza, Glauco R.

2016-01-01

Vasoactive liabilities are typically assayed using wire myography, which is limited by its high cost and low throughput. To meet the demand for higher throughput in vitro alternatives, this study introduces a magnetic 3D bioprinting-based vasoactivity assay. The principle behind this assay is the magnetic printing of vascular smooth muscle cells into 3D rings that functionally represent blood vessel segments, whose contraction can be altered by vasodilators and vasoconstrictors. A cost-effective imaging modality employing a mobile device is used to capture contraction with high throughput. The goal of this study was to validate ring contraction as a measure of vasoactivity, using a small panel of known vasoactive drugs. In vitro responses of the rings matched outcomes predicted by in vivo pharmacology, and were supported by immunohistochemistry. Altogether, this ring assay robustly models vasoactivity, which could meet the need for higher throughput in vitro alternatives. PMID:27477945

Quantitative high-throughput population dynamics in continuous-culture by automated microscopy.

PubMed

Merritt, Jason; Kuehn, Seppe

2016-09-12

We present a high-throughput method to measure abundance dynamics in microbial communities sustained in continuous-culture. Our method uses custom epi-fluorescence microscopes to automatically image single cells drawn from a continuously-cultured population while precisely controlling culture conditions. For clonal populations of Escherichia coli our instrument reveals history-dependent resilience and growth rate dependent aggregation.

An Automated Method for High-Throughput Screening of Arabidopsis Rosette Growth in Multi-Well Plates and Its Validation in Stress Conditions.

PubMed

De Diego, Nuria; Fürst, Tomáš; Humplík, Jan F; Ugena, Lydia; Podlešáková, Kateřina; Spíchal, Lukáš

2017-01-01

High-throughput plant phenotyping platforms provide new possibilities for automated, fast scoring of several plant growth and development traits, followed over time using non-invasive sensors. Using Arabidops is as a model offers important advantages for high-throughput screening with the opportunity to extrapolate the results obtained to other crops of commercial interest. In this study we describe the development of a highly reproducible high-throughput Arabidopsis in vitro bioassay established using our OloPhen platform, suitable for analysis of rosette growth in multi-well plates. This method was successfully validated on example of multivariate analysis of Arabidopsis rosette growth in different salt concentrations and the interaction with varying nutritional composition of the growth medium. Several traits such as changes in the rosette area, relative growth rate, survival rate and homogeneity of the population are scored using fully automated RGB imaging and subsequent image analysis. The assay can be used for fast screening of the biological activity of chemical libraries, phenotypes of transgenic or recombinant inbred lines, or to search for potential quantitative trait loci. It is especially valuable for selecting genotypes or growth conditions that improve plant stress tolerance.

Development of a High-Throughput Microwave Imaging System for Concealed Weapons Detection

DTIC Science & Technology

2016-07-15

hardware. Index Terms—Microwave imaging, multistatic radar, Fast Fourier Transform (FFT). I. INTRODUCTION Near-field microwave imaging is a non-ionizing...configuration, but its computational demands are extreme. Fast Fourier Transform (FFT) imaging has long been used to efficiently construct images sampled with...Simulated image of 25 point scatterers imaged at range 1.5m, with array layout depicted in Fig. 3. Left: image formed with Equation (5) ( Fourier

Improved high-throughput quantification of luminescent microplate assays using a common Western-blot imaging system.

PubMed

Hawkins, Liam J; Storey, Kenneth B

2017-01-01

Common Western-blot imaging systems have previously been adapted to measure signals from luminescent microplate assays. This can be a cost saving measure as Western-blot imaging systems are common laboratory equipment and could substitute a dedicated luminometer if one is not otherwise available. One previously unrecognized limitation is that the signals captured by the cameras in these systems are not equal for all wells. Signals are dependent on the angle of incidence to the camera, and thus the location of the well on the microplate. Here we show that: •The position of a well on a microplate significantly affects the signal captured by a common Western-blot imaging system from a luminescent assay.•The effect of well position can easily be corrected for.•This method can be applied to commercially available luminescent assays, allowing for high-throughput quantification of a wide range of biological processes and biochemical reactions.

CellCognition: time-resolved phenotype annotation in high-throughput live cell imaging.

PubMed

Held, Michael; Schmitz, Michael H A; Fischer, Bernd; Walter, Thomas; Neumann, Beate; Olma, Michael H; Peter, Matthias; Ellenberg, Jan; Gerlich, Daniel W

2010-09-01

Fluorescence time-lapse imaging has become a powerful tool to investigate complex dynamic processes such as cell division or intracellular trafficking. Automated microscopes generate time-resolved imaging data at high throughput, yet tools for quantification of large-scale movie data are largely missing. Here we present CellCognition, a computational framework to annotate complex cellular dynamics. We developed a machine-learning method that combines state-of-the-art classification with hidden Markov modeling for annotation of the progression through morphologically distinct biological states. Incorporation of time information into the annotation scheme was essential to suppress classification noise at state transitions and confusion between different functional states with similar morphology. We demonstrate generic applicability in different assays and perturbation conditions, including a candidate-based RNA interference screen for regulators of mitotic exit in human cells. CellCognition is published as open source software, enabling live-cell imaging-based screening with assays that directly score cellular dynamics.

Targeted post-mortem computed tomography cardiac angiography: proof of concept.

PubMed

Saunders, Sarah L; Morgan, Bruno; Raj, Vimal; Robinson, Claire E; Rutty, Guy N

2011-07-01

With the increasing use and availability of multi-detector computed tomography and magnetic resonance imaging in autopsy practice, there has been an international push towards the development of the so-called near virtual autopsy. However, currently, a significant obstacle to the consideration as to whether or not near virtual autopsies could one day replace the conventional invasive autopsy is the failure of post-mortem imaging to yield detailed information concerning the coronary arteries. To date, a cost-effective, practical solution to allow high throughput imaging has not been presented within the forensic literature. We present a proof of concept paper describing a simple, quick, cost-effective, manual, targeted in situ post-mortem cardiac angiography method using a minimally invasive approach, to be used with multi-detector computed tomography for high throughput cadaveric imaging which can be used in permanent or temporary mortuaries.

Automated sample area definition for high-throughput microscopy.

PubMed

Zeder, M; Ellrott, A; Amann, R

2011-04-01

High-throughput screening platforms based on epifluorescence microscopy are powerful tools in a variety of scientific fields. Although some applications are based on imaging geometrically defined samples such as microtiter plates, multiwell slides, or spotted gene arrays, others need to cope with inhomogeneously located samples on glass slides. The analysis of microbial communities in aquatic systems by sample filtration on membrane filters followed by multiple fluorescent staining, or the investigation of tissue sections are examples. Therefore, we developed a strategy for flexible and fast definition of sample locations by the acquisition of whole slide overview images and automated sample recognition by image analysis. Our approach was tested on different microscopes and the computer programs are freely available (http://www.technobiology.ch). Copyright © 2011 International Society for Advancement of Cytometry.

Using iterative cluster merging with improved gap statistics to perform online phenotype discovery in the context of high-throughput RNAi screens

PubMed Central

Yin, Zheng; Zhou, Xiaobo; Bakal, Chris; Li, Fuhai; Sun, Youxian; Perrimon, Norbert; Wong, Stephen TC

2008-01-01

Background The recent emergence of high-throughput automated image acquisition technologies has forever changed how cell biologists collect and analyze data. Historically, the interpretation of cellular phenotypes in different experimental conditions has been dependent upon the expert opinions of well-trained biologists. Such qualitative analysis is particularly effective in detecting subtle, but important, deviations in phenotypes. However, while the rapid and continuing development of automated microscope-based technologies now facilitates the acquisition of trillions of cells in thousands of diverse experimental conditions, such as in the context of RNA interference (RNAi) or small-molecule screens, the massive size of these datasets precludes human analysis. Thus, the development of automated methods which aim to identify novel and biological relevant phenotypes online is one of the major challenges in high-throughput image-based screening. Ideally, phenotype discovery methods should be designed to utilize prior/existing information and tackle three challenging tasks, i.e. restoring pre-defined biological meaningful phenotypes, differentiating novel phenotypes from known ones and clarifying novel phenotypes from each other. Arbitrarily extracted information causes biased analysis, while combining the complete existing datasets with each new image is intractable in high-throughput screens. Results Here we present the design and implementation of a novel and robust online phenotype discovery method with broad applicability that can be used in diverse experimental contexts, especially high-throughput RNAi screens. This method features phenotype modelling and iterative cluster merging using improved gap statistics. A Gaussian Mixture Model (GMM) is employed to estimate the distribution of each existing phenotype, and then used as reference distribution in gap statistics. This method is broadly applicable to a number of different types of image-based datasets derived from a wide spectrum of experimental conditions and is suitable to adaptively process new images which are continuously added to existing datasets. Validations were carried out on different dataset, including published RNAi screening using Drosophila embryos [Additional files 1, 2], dataset for cell cycle phase identification using HeLa cells [Additional files 1, 3, 4] and synthetic dataset using polygons, our methods tackled three aforementioned tasks effectively with an accuracy range of 85%–90%. When our method is implemented in the context of a Drosophila genome-scale RNAi image-based screening of cultured cells aimed to identifying the contribution of individual genes towards the regulation of cell-shape, it efficiently discovers meaningful new phenotypes and provides novel biological insight. We also propose a two-step procedure to modify the novelty detection method based on one-class SVM, so that it can be used to online phenotype discovery. In different conditions, we compared the SVM based method with our method using various datasets and our methods consistently outperformed SVM based method in at least two of three tasks by 2% to 5%. These results demonstrate that our methods can be used to better identify novel phenotypes in image-based datasets from a wide range of conditions and organisms. Conclusion We demonstrate that our method can detect various novel phenotypes effectively in complex datasets. Experiment results also validate that our method performs consistently under different order of image input, variation of starting conditions including the number and composition of existing phenotypes, and dataset from different screens. In our findings, the proposed method is suitable for online phenotype discovery in diverse high-throughput image-based genetic and chemical screens. PMID:18534020

Recovering the dynamics of root growth and development using novel image acquisition and analysis methods

PubMed Central

Wells, Darren M.; French, Andrew P.; Naeem, Asad; Ishaq, Omer; Traini, Richard; Hijazi, Hussein; Bennett, Malcolm J.; Pridmore, Tony P.

2012-01-01

Roots are highly responsive to environmental signals encountered in the rhizosphere, such as nutrients, mechanical resistance and gravity. As a result, root growth and development is very plastic. If this complex and vital process is to be understood, methods and tools are required to capture the dynamics of root responses. Tools are needed which are high-throughput, supporting large-scale experimental work, and provide accurate, high-resolution, quantitative data. We describe and demonstrate the efficacy of the high-throughput and high-resolution root imaging systems recently developed within the Centre for Plant Integrative Biology (CPIB). This toolset includes (i) robotic imaging hardware to generate time-lapse datasets from standard cameras under infrared illumination and (ii) automated image analysis methods and software to extract quantitative information about root growth and development both from these images and via high-resolution light microscopy. These methods are demonstrated using data gathered during an experimental study of the gravitropic response of Arabidopsis thaliana. PMID:22527394

Recovering the dynamics of root growth and development using novel image acquisition and analysis methods.

PubMed

Wells, Darren M; French, Andrew P; Naeem, Asad; Ishaq, Omer; Traini, Richard; Hijazi, Hussein I; Hijazi, Hussein; Bennett, Malcolm J; Pridmore, Tony P

2012-06-05

Roots are highly responsive to environmental signals encountered in the rhizosphere, such as nutrients, mechanical resistance and gravity. As a result, root growth and development is very plastic. If this complex and vital process is to be understood, methods and tools are required to capture the dynamics of root responses. Tools are needed which are high-throughput, supporting large-scale experimental work, and provide accurate, high-resolution, quantitative data. We describe and demonstrate the efficacy of the high-throughput and high-resolution root imaging systems recently developed within the Centre for Plant Integrative Biology (CPIB). This toolset includes (i) robotic imaging hardware to generate time-lapse datasets from standard cameras under infrared illumination and (ii) automated image analysis methods and software to extract quantitative information about root growth and development both from these images and via high-resolution light microscopy. These methods are demonstrated using data gathered during an experimental study of the gravitropic response of Arabidopsis thaliana.

Efficient globally optimal segmentation of cells in fluorescence microscopy images using level sets and convex energy functionals.

PubMed

Bergeest, Jan-Philip; Rohr, Karl

2012-10-01

In high-throughput applications, accurate and efficient segmentation of cells in fluorescence microscopy images is of central importance for the quantification of protein expression and the understanding of cell function. We propose an approach for segmenting cell nuclei which is based on active contours using level sets and convex energy functionals. Compared to previous work, our approach determines the global solution. Thus, the approach does not suffer from local minima and the segmentation result does not depend on the initialization. We consider three different well-known energy functionals for active contour-based segmentation and introduce convex formulations of these functionals. We also suggest a numeric approach for efficiently computing the solution. The performance of our approach has been evaluated using fluorescence microscopy images from different experiments comprising different cell types. We have also performed a quantitative comparison with previous segmentation approaches. Copyright © 2012 Elsevier B.V. All rights reserved.

High Throughput Determination of Critical Human Dosing Parameters (SOT)

EPA Science Inventory

High throughput toxicokinetics (HTTK) is a rapid approach that uses in vitro data to estimate TK for hundreds of environmental chemicals. Reverse dosimetry (i.e., reverse toxicokinetics or RTK) based on HTTK data converts high throughput in vitro toxicity screening (HTS) data int...

High Throughput Determinations of Critical Dosing Parameters (IVIVE workshop)

EPA Science Inventory

High throughput toxicokinetics (HTTK) is an approach that allows for rapid estimations of TK for hundreds of environmental chemicals. HTTK-based reverse dosimetry (i.e, reverse toxicokinetics or RTK) is used in order to convert high throughput in vitro toxicity screening (HTS) da...

A novel high-throughput imaging system for automated analyses of avoidance behavior in zebrafish larvae

PubMed Central

Pelkowski, Sean D.; Kapoor, Mrinal; Richendrfer, Holly A.; Wang, Xingyue; Colwill, Ruth M.; Creton, Robbert

2011-01-01

Early brain development can be influenced by numerous genetic and environmental factors, with long-lasting effects on brain function and behavior. The identification of these factors is facilitated by recent innovations in high-throughput screening. However, large-scale screening in whole organisms remains challenging, in particular when studying changes in brain function or behavior in vertebrate model systems. In this study, we present a novel imaging system for high-throughput analyses of behavior in zebrafish larvae. The three-camera system can image twelve multiwell plates simultaneously and is unique in its ability to provide local visual stimuli in the wells of a multiwell plate. The acquired images are converted into a series of coordinates, which characterize the location and orientation of the larvae. The developed imaging techniques were tested by measuring avoidance behaviors in seven-day-old zebrafish larvae. The system effectively quantified larval avoidance and revealed an increased edge preference in response to a blue or red ‘bouncing ball’ stimulus. Larvae also avoid a bouncing ball stimulus when it is counter-balanced with a stationary ball, but do not avoid blinking balls counter-balanced with a stationary ball. These results indicate that the seven-day-old larvae respond specifically to movement, rather than color, size, or local changes in light intensity. The imaging system and assays for measuring avoidance behavior may be used to screen for genetic and environmental factors that cause developmental brain disorders and for novel drugs that could prevent or treat these disorders. PMID:21549762

A novel high-throughput imaging system for automated analyses of avoidance behavior in zebrafish larvae.

PubMed

Pelkowski, Sean D; Kapoor, Mrinal; Richendrfer, Holly A; Wang, Xingyue; Colwill, Ruth M; Creton, Robbert

2011-09-30

Early brain development can be influenced by numerous genetic and environmental factors, with long-lasting effects on brain function and behavior. The identification of these factors is facilitated by recent innovations in high-throughput screening. However, large-scale screening in whole organisms remains challenging, in particular when studying changes in brain function or behavior in vertebrate model systems. In this study, we present a novel imaging system for high-throughput analyses of behavior in zebrafish larvae. The three-camera system can image 12 multiwell plates simultaneously and is unique in its ability to provide local visual stimuli in the wells of a multiwell plate. The acquired images are converted into a series of coordinates, which characterize the location and orientation of the larvae. The developed imaging techniques were tested by measuring avoidance behaviors in seven-day-old zebrafish larvae. The system effectively quantified larval avoidance and revealed an increased edge preference in response to a blue or red 'bouncing ball' stimulus. Larvae also avoid a bouncing ball stimulus when it is counter-balanced with a stationary ball, but do not avoid blinking balls counter-balanced with a stationary ball. These results indicate that the seven-day-old larvae respond specifically to movement, rather than color, size, or local changes in light intensity. The imaging system and assays for measuring avoidance behavior may be used to screen for genetic and environmental factors that cause developmental brain disorders and for novel drugs that could prevent or treat these disorders. Copyright © 2011 Elsevier B.V. All rights reserved.

A Novel, Poly-Etalon, Fabry-Perot for Planetary Research

NASA Technical Reports Server (NTRS)

Kerr, Robert B.; Doe, Richard; Noto, John

1997-01-01

In an effort to develop a mechanically robust, high throughput and solid state spectrometer several liquid crystal Fabry-Perot etalons were constructed. The etalons were tested for spectral response, radiation resistance and optical transmission. The first year of this project was spent developing and understanding the properties of the liquid crystal etalons; in the second year an intensified all-sky imaging system was developed around a pair of LC etalons. The imaging system, developed jointly with SRI International represents a unique brassboard to demonstrate the use of LC etalons as tunable filters. The first set of etalons constructed in year one of this project were tested for spectral response and throughput while etalon surrogates were exposed to proton radiation simulating the exposure of an object in Low Earth Orbit (LEO). The 2" diameter etalons had a measure finesse of approximately 10 and were tunable over five orders. Liquid crystals exposed to proton irradiation showed no signs of damage. In year two two larger diameter (3") etalons were constructed with gaps of 3 and 5 microns. This pair of etalons is for use in a high resolution, all-sky spectral imager. The WATUMI imager system follows the heritage of all sky, narrow band, intensified imagers however it includes two LC Fabry-Perot etalons to provide tunability and the ability to switch wavelengths rapidly, an import consideration in auroral airglow imaging. This work also resulted in two publications and one poster presentation. The instrument will be uniquely capable, with superior throughput and speed, to measure optical airglow of multiple emission lines in harsh conditions.

Automated crystallographic system for high-throughput protein structure determination.

PubMed

Brunzelle, Joseph S; Shafaee, Padram; Yang, Xiaojing; Weigand, Steve; Ren, Zhong; Anderson, Wayne F

2003-07-01

High-throughput structural genomic efforts require software that is highly automated, distributive and requires minimal user intervention to determine protein structures. Preliminary experiments were set up to test whether automated scripts could utilize a minimum set of input parameters and produce a set of initial protein coordinates. From this starting point, a highly distributive system was developed that could determine macromolecular structures at a high throughput rate, warehouse and harvest the associated data. The system uses a web interface to obtain input data and display results. It utilizes a relational database to store the initial data needed to start the structure-determination process as well as generated data. A distributive program interface administers the crystallographic programs which determine protein structures. Using a test set of 19 protein targets, 79% were determined automatically.

AI-augmented time stretch microscopy

NASA Astrophysics Data System (ADS)

Mahjoubfar, Ata; Chen, Claire L.; Lin, Jiahao; Jalali, Bahram

2017-02-01

Cell reagents used in biomedical analysis often change behavior of the cells that they are attached to, inhibiting their native signaling. On the other hand, label-free cell analysis techniques have long been viewed as challenging either due to insufficient accuracy by limited features, or because of low throughput as a sacrifice of improved precision. We present a recently developed artificial-intelligence augmented microscope, which builds upon high-throughput time stretch quantitative phase imaging (TS-QPI) and deep learning to perform label-free cell classification with record high-accuracy. Our system captures quantitative optical phase and intensity images simultaneously by frequency multiplexing, extracts multiple biophysical features of the individual cells from these images fused, and feeds these features into a supervised machine learning model for classification. The enhanced performance of our system compared to other label-free assays is demonstrated by classification of white blood T-cells versus colon cancer cells and lipid accumulating algal strains for biofuel production, which is as much as five-fold reduction in inaccuracy. This system obtains the accuracy required in practical applications such as personalized drug development, while the cells remain intact and the throughput is not sacrificed. Here, we introduce a data acquisition scheme based on quadrature phase demodulation that enables interruptionless storage of TS-QPI cell images. Our proof of principle demonstration is capable of saving 40 TB of cell images in about four hours, i.e. pictures of every single cell in 10 mL of a sample.

Diamond Turned High Precision PIAA Optics and Four Mirror PIAA System for High Contrast Imaging of Exo-planets

NASA Technical Reports Server (NTRS)

Balasubramanian, Kunjithapatham; Cady, Eric; Pueyo, Laurent; Ana, Xin; Shaklan, Stuart; Guyon, Olivier; Belikov, Ruslan

2011-01-01

Off-axis, high-sag PIAA optics for high contrast imaging present challenges in manufacturing and testing. With smaller form factors and consequently smaller surface deformations (< 80 microns), diamond turned fabrication of these mirrors becomes feasible. Though such a design reduces the system throughput, it still provides 2(lambda)D inner working angle. We report on the design, fabrication, measurements, and initial assessment of the novel PIAA optics in a coronagraph testbed. We also describe, for the first time, a four mirror PIAA coronagraph that relaxes apodizer requirements and significantly improves throughput while preserving the low-cost benefits.

High-Throughput Phenotyping of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes and Neurons Using Electric Field Stimulation and High-Speed Fluorescence Imaging

PubMed Central

Daily, Neil J.; Du, Zhong-Wei

2017-01-01

Abstract Electrophysiology of excitable cells, including muscle cells and neurons, has been measured by making direct contact with a single cell using a micropipette electrode. To increase the assay throughput, optical devices such as microscopes and microplate readers have been used to analyze electrophysiology of multiple cells. We have established a high-throughput (HTP) analysis of action potentials (APs) in highly enriched motor neurons and cardiomyocytes (CMs) that are differentiated from human induced pluripotent stem cells (iPSCs). A multichannel electric field stimulation (EFS) device enabled the ability to electrically stimulate cells and measure dynamic changes in APs of excitable cells ultra-rapidly (>100 data points per second) by imaging entire 96-well plates. We found that the activities of both neurons and CMs and their response to EFS and chemicals are readily discerned by our fluorescence imaging-based HTP phenotyping assay. The latest generation of calcium (Ca2+) indicator dyes, FLIPR Calcium 6 and Cal-520, with the HTP device enables physiological analysis of human iPSC-derived samples highlighting its potential application for understanding disease mechanisms and discovering new therapeutic treatments. PMID:28525289

High-throughput two-dimensional root system phenotyping platform facilitates genetic analysis of root growth and development.

PubMed

Clark, Randy T; Famoso, Adam N; Zhao, Keyan; Shaff, Jon E; Craft, Eric J; Bustamante, Carlos D; McCouch, Susan R; Aneshansley, Daniel J; Kochian, Leon V

2013-02-01

High-throughput phenotyping of root systems requires a combination of specialized techniques and adaptable plant growth, root imaging and software tools. A custom phenotyping platform was designed to capture images of whole root systems, and novel software tools were developed to process and analyse these images. The platform and its components are adaptable to a wide range root phenotyping studies using diverse growth systems (hydroponics, paper pouches, gel and soil) involving several plant species, including, but not limited to, rice, maize, sorghum, tomato and Arabidopsis. The RootReader2D software tool is free and publicly available and was designed with both user-guided and automated features that increase flexibility and enhance efficiency when measuring root growth traits from specific roots or entire root systems during large-scale phenotyping studies. To demonstrate the unique capabilities and high-throughput capacity of this phenotyping platform for studying root systems, genome-wide association studies on rice (Oryza sativa) and maize (Zea mays) root growth were performed and root traits related to aluminium (Al) tolerance were analysed on the parents of the maize nested association mapping (NAM) population. © 2012 Blackwell Publishing Ltd.

High-throughput linear optical stretcher for mechanical characterization of blood cells.

PubMed

Roth, Kevin B; Neeves, Keith B; Squier, Jeff; Marr, David W M

2016-04-01

This study describes a linear optical stretcher as a high-throughput mechanical property cytometer. Custom, inexpensive, and scalable optics image a linear diode bar source into a microfluidic channel, where cells are hydrodynamically focused into the optical stretcher. Upon entering the stretching region, antipodal optical forces generated by the refraction of tightly focused laser light at the cell membrane deform each cell in flow. Each cell relaxes as it flows out of the trap and is compared to the stretched state to determine deformation. The deformation response of untreated red blood cells and neutrophils were compared to chemically treated cells. Statistically significant differences were observed between normal, diamide-treated, and glutaraldehyde-treated red blood cells, as well as between normal and cytochalasin D-treated neutrophils. Based on the behavior of the pure, untreated populations of red cells and neutrophils, a mixed population of these cells was tested and the discrete populations were identified by deformability. © 2015 International Society for Advancement of Cytometry. © 2015 International Society for Advancement of Cytometry.

Anatomo-radiological correlation using 18-FDG-PET in abdominal sepsis model in rats. A preliminary study.

PubMed

Azevedo, Ítalo Medeiros; Carvalho, Marília Daniela Ferreira; Nascimento, Rafael Pereira; Macedo, Robson; Aquino, Mônica Raquel de Souza; Medeiros, Aldo Cunha

2017-03-01

To examine a correlation of micro-PET images with photographic images of the digestive organs in abdominal sepsis model. Male Wistar rats weighing 265±18g were used. Abdominal sepsis was induced by ligature and cecal puncture. Micro-PET Images from abdominal cavity septic foci were obtained using 18-Fluoro-deoxyglucose, looking for a correlation with photographic images of abdominal cavity organs. Pearson's correlation test was used. The mean standard uptake values (SUV) and lesion areas were 2.58±0.63SUVbwg/ml and 546.87±300.95mm2, respectively. There was a strong positive correlation between the two variables (r=0.863, p=0.137), which resulted in a coefficient of determination r2?0.75, meaning that 75% of SUV variation is explained by the lesion areas of digestive organs. Micro-PET allows high throughput assessment of lesion count and volume in pre-clinical rat model of CPL abdominal sepsis.

Helium Ion Beam Microscopy for Copper Grain Identification in BEOL Structures

NASA Astrophysics Data System (ADS)

van den Boom, Ruud J. J.; Parvaneh, Hamed; Voci, Dave; Huynh, Chuong; Stern, Lewis; Dunn, Kathleen A.; Lifshin, Eric

2009-09-01

Grain size determination in advanced metallization structures requires a technique with resolution ˜2 nm, with a high signal-to-noise ratio and high orientation-dependant contrast for unambiguous identification of grain boundaries. Ideally, such a technique would also be capable of high-throughput and rapid time-to-knowledge. The Helium Ion Microscope (HIM) offers one possibility for achieving these aims in a single platform. This article compares the performance of the HIM with Focused Ion Beam, Scanning Electron and Transmission Electron Microscopes, in terms of achievable image resolution and contrast, using plan-view and cross-sectional imaging of electroplated samples. Although the HIM is capable of sub-nanometer beam diameter, the low signal-to-noise ratio in the images necessitates signal averaging, which degrades the measured image resolution to 6-8 nm. Strategies for improving S/N are discussed in light of the trade-off between beam current and probe size, accelerating voltage, and dwell time.

All-passive pixel super-resolution of time-stretch imaging

PubMed Central

Chan, Antony C. S.; Ng, Ho-Cheung; Bogaraju, Sharat C. V.; So, Hayden K. H.; Lam, Edmund Y.; Tsia, Kevin K.

2017-01-01

Based on image encoding in a serial-temporal format, optical time-stretch imaging entails a stringent requirement of state-of-the-art fast data acquisition unit in order to preserve high image resolution at an ultrahigh frame rate — hampering the widespread utilities of such technology. Here, we propose a pixel super-resolution (pixel-SR) technique tailored for time-stretch imaging that preserves pixel resolution at a relaxed sampling rate. It harnesses the subpixel shifts between image frames inherently introduced by asynchronous digital sampling of the continuous time-stretch imaging process. Precise pixel registration is thus accomplished without any active opto-mechanical subpixel-shift control or other additional hardware. Here, we present the experimental pixel-SR image reconstruction pipeline that restores high-resolution time-stretch images of microparticles and biological cells (phytoplankton) at a relaxed sampling rate (≈2–5 GSa/s)—more than four times lower than the originally required readout rate (20 GSa/s) — is thus effective for high-throughput label-free, morphology-based cellular classification down to single-cell precision. Upon integration with the high-throughput image processing technology, this pixel-SR time-stretch imaging technique represents a cost-effective and practical solution for large scale cell-based phenotypic screening in biomedical diagnosis and machine vision for quality control in manufacturing. PMID:28303936

Quantitative detection of benzoyl peroxide in wheat flour by line-scan macro-scale Raman chemical imaging

USDA-ARS?s Scientific Manuscript database

A high-throughput Raman chemical imaging method was developed for direct inspection of benzoyl peroxide (BPO) mixed in wheat flour. A 5 W 785 nm line laser (240 mm long and 1 mm wide) was used as a Raman excitation source in a push-broom Raman imaging system. Hyperspectral Raman images were collecte...

High-throughput quantum cascade laser (QCL) spectral histopathology: a practical approach towards clinical translation.

PubMed

Pilling, Michael J; Henderson, Alex; Bird, Benjamin; Brown, Mick D; Clarke, Noel W; Gardner, Peter

2016-06-23

Infrared microscopy has become one of the key techniques in the biomedical research field for interrogating tissue. In partnership with multivariate analysis and machine learning techniques, it has become widely accepted as a method that can distinguish between normal and cancerous tissue with both high sensitivity and high specificity. While spectral histopathology (SHP) is highly promising for improved clinical diagnosis, several practical barriers currently exist, which need to be addressed before successful implementation in the clinic. Sample throughput and speed of acquisition are key barriers and have been driven by the high volume of samples awaiting histopathological examination. FTIR chemical imaging utilising FPA technology is currently state-of-the-art for infrared chemical imaging, and recent advances in its technology have dramatically reduced acquisition times. Despite this, infrared microscopy measurements on a tissue microarray (TMA), often encompassing several million spectra, takes several hours to acquire. The problem lies with the vast quantities of data that FTIR collects; each pixel in a chemical image is derived from a full infrared spectrum, itself composed of thousands of individual data points. Furthermore, data management is quickly becoming a barrier to clinical translation and poses the question of how to store these incessantly growing data sets. Recently, doubts have been raised as to whether the full spectral range is actually required for accurate disease diagnosis using SHP. These studies suggest that once spectral biomarkers have been predetermined it may be possible to diagnose disease based on a limited number of discrete spectral features. In this current study, we explore the possibility of utilising discrete frequency chemical imaging for acquiring high-throughput, high-resolution chemical images. Utilising a quantum cascade laser imaging microscope with discrete frequency collection at key diagnostic wavelengths, we demonstrate that we can diagnose prostate cancer with high sensitivity and specificity. Finally we extend the study to a large patient dataset utilising tissue microarrays, and show that high sensitivity and specificity can be achieved using high-throughput, rapid data collection, thereby paving the way for practical implementation in the clinic.

Advances in molecular labeling, high throughput imaging and machine intelligence portend powerful functional cellular biochemistry tools.

PubMed

Price, Jeffrey H; Goodacre, Angela; Hahn, Klaus; Hodgson, Louis; Hunter, Edward A; Krajewski, Stanislaw; Murphy, Robert F; Rabinovich, Andrew; Reed, John C; Heynen, Susanne

2002-01-01

Cellular behavior is complex. Successfully understanding systems at ever-increasing complexity is fundamental to advances in modern science and unraveling the functional details of cellular behavior is no exception. We present a collection of prospectives to provide a glimpse of the techniques that will aid in collecting, managing and utilizing information on complex cellular processes via molecular imaging tools. These include: 1) visualizing intracellular protein activity with fluorescent markers, 2) high throughput (and automated) imaging of multilabeled cells in statistically significant numbers, and 3) machine intelligence to analyze subcellular image localization and pattern. Although not addressed here, the importance of combining cell-image-based information with detailed molecular structure and ligand-receptor binding models cannot be overlooked. Advanced molecular imaging techniques have the potential to impact cellular diagnostics for cancer screening, clinical correlations of tissue molecular patterns for cancer biology, and cellular molecular interactions for accelerating drug discovery. The goal of finally understanding all cellular components and behaviors will be achieved by advances in both instrumentation engineering (software and hardware) and molecular biochemistry. Copyright 2002 Wiley-Liss, Inc.

Digital Biomass Accumulation Using High-Throughput Plant Phenotype Data Analysis.

PubMed

Rahaman, Md Matiur; Ahsan, Md Asif; Gillani, Zeeshan; Chen, Ming

2017-09-01

Biomass is an important phenotypic trait in functional ecology and growth analysis. The typical methods for measuring biomass are destructive, and they require numerous individuals to be cultivated for repeated measurements. With the advent of image-based high-throughput plant phenotyping facilities, non-destructive biomass measuring methods have attempted to overcome this problem. Thus, the estimation of plant biomass of individual plants from their digital images is becoming more important. In this paper, we propose an approach to biomass estimation based on image derived phenotypic traits. Several image-based biomass studies state that the estimation of plant biomass is only a linear function of the projected plant area in images. However, we modeled the plant volume as a function of plant area, plant compactness, and plant age to generalize the linear biomass model. The obtained results confirm the proposed model and can explain most of the observed variance during image-derived biomass estimation. Moreover, a small difference was observed between actual and estimated digital biomass, which indicates that our proposed approach can be used to estimate digital biomass accurately.

Automated analysis of siRNA screens of cells infected by hepatitis C and dengue viruses based on immunofluorescence microscopy images

NASA Astrophysics Data System (ADS)

Matula, Petr; Kumar, Anil; Wörz, Ilka; Harder, Nathalie; Erfle, Holger; Bartenschlager, Ralf; Eils, Roland; Rohr, Karl

2008-03-01

We present an image analysis approach as part of a high-throughput microscopy siRNA-based screening system using cell arrays for the identification of cellular genes involved in hepatitis C and dengue virus replication. Our approach comprises: cell nucleus segmentation, quantification of virus replication level in the neighborhood of segmented cell nuclei, localization of regions with transfected cells, cell classification by infection status, and quality assessment of an experiment and single images. In particular, we propose a novel approach for the localization of regions of transfected cells within cell array images, which combines model-based circle fitting and grid fitting. By this scheme we integrate information from single cell array images and knowledge from the complete cell arrays. The approach is fully automatic and has been successfully applied to a large number of cell array images from screening experiments. The experimental results show a good agreement with the expected behaviour of positive as well as negative controls and encourage the application to screens from further high-throughput experiments.

A robust, high-throughput method for computing maize ear, cob, and kernel attributes automatically from images.

PubMed

Miller, Nathan D; Haase, Nicholas J; Lee, Jonghyun; Kaeppler, Shawn M; de Leon, Natalia; Spalding, Edgar P

2017-01-01

Grain yield of the maize plant depends on the sizes, shapes, and numbers of ears and the kernels they bear. An automated pipeline that can measure these components of yield from easily-obtained digital images is needed to advance our understanding of this globally important crop. Here we present three custom algorithms designed to compute such yield components automatically from digital images acquired by a low-cost platform. One algorithm determines the average space each kernel occupies along the cob axis using a sliding-window Fourier transform analysis of image intensity features. A second counts individual kernels removed from ears, including those in clusters. A third measures each kernel's major and minor axis after a Bayesian analysis of contour points identifies the kernel tip. Dimensionless ear and kernel shape traits that may interrelate yield components are measured by principal components analysis of contour point sets. Increased objectivity and speed compared to typical manual methods are achieved without loss of accuracy as evidenced by high correlations with ground truth measurements and simulated data. Millimeter-scale differences among ear, cob, and kernel traits that ranged more than 2.5-fold across a diverse group of inbred maize lines were resolved. This system for measuring maize ear, cob, and kernel attributes is being used by multiple research groups as an automated Web service running on community high-throughput computing and distributed data storage infrastructure. Users may create their own workflow using the source code that is staged for download on a public repository. © 2016 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.

Micropillar arrays as a high-throughput screening platform for therapeutics in multiple sclerosis.

PubMed

Mei, Feng; Fancy, Stephen P J; Shen, Yun-An A; Niu, Jianqin; Zhao, Chao; Presley, Bryan; Miao, Edna; Lee, Seonok; Mayoral, Sonia R; Redmond, Stephanie A; Etxeberria, Ainhoa; Xiao, Lan; Franklin, Robin J M; Green, Ari; Hauser, Stephen L; Chan, Jonah R

2014-08-01

Functional screening for compounds that promote remyelination represents a major hurdle in the development of rational therapeutics for multiple sclerosis. Screening for remyelination is problematic, as myelination requires the presence of axons. Standard methods do not resolve cell-autonomous effects and are not suited for high-throughput formats. Here we describe a binary indicant for myelination using micropillar arrays (BIMA). Engineered with conical dimensions, micropillars permit resolution of the extent and length of membrane wrapping from a single two-dimensional image. Confocal imaging acquired from the base to the tip of the pillars allows for detection of concentric wrapping observed as 'rings' of myelin. The platform is formatted in 96-well plates, amenable to semiautomated random acquisition and automated detection and quantification. Upon screening 1,000 bioactive molecules, we identified a cluster of antimuscarinic compounds that enhance oligodendrocyte differentiation and remyelination. Our findings demonstrate a new high-throughput screening platform for potential regenerative therapeutics in multiple sclerosis.

The Open Connectome Project Data Cluster: Scalable Analysis and Vision for High-Throughput Neuroscience.

PubMed

Burns, Randal; Roncal, William Gray; Kleissas, Dean; Lillaney, Kunal; Manavalan, Priya; Perlman, Eric; Berger, Daniel R; Bock, Davi D; Chung, Kwanghun; Grosenick, Logan; Kasthuri, Narayanan; Weiler, Nicholas C; Deisseroth, Karl; Kazhdan, Michael; Lichtman, Jeff; Reid, R Clay; Smith, Stephen J; Szalay, Alexander S; Vogelstein, Joshua T; Vogelstein, R Jacob

2013-01-01

We describe a scalable database cluster for the spatial analysis and annotation of high-throughput brain imaging data, initially for 3-d electron microscopy image stacks, but for time-series and multi-channel data as well. The system was designed primarily for workloads that build connectomes - neural connectivity maps of the brain-using the parallel execution of computer vision algorithms on high-performance compute clusters. These services and open-science data sets are publicly available at openconnecto.me. The system design inherits much from NoSQL scale-out and data-intensive computing architectures. We distribute data to cluster nodes by partitioning a spatial index. We direct I/O to different systems-reads to parallel disk arrays and writes to solid-state storage-to avoid I/O interference and maximize throughput. All programming interfaces are RESTful Web services, which are simple and stateless, improving scalability and usability. We include a performance evaluation of the production system, highlighting the effec-tiveness of spatial data organization.

The Open Connectome Project Data Cluster: Scalable Analysis and Vision for High-Throughput Neuroscience

PubMed Central

Burns, Randal; Roncal, William Gray; Kleissas, Dean; Lillaney, Kunal; Manavalan, Priya; Perlman, Eric; Berger, Daniel R.; Bock, Davi D.; Chung, Kwanghun; Grosenick, Logan; Kasthuri, Narayanan; Weiler, Nicholas C.; Deisseroth, Karl; Kazhdan, Michael; Lichtman, Jeff; Reid, R. Clay; Smith, Stephen J.; Szalay, Alexander S.; Vogelstein, Joshua T.; Vogelstein, R. Jacob

2013-01-01

We describe a scalable database cluster for the spatial analysis and annotation of high-throughput brain imaging data, initially for 3-d electron microscopy image stacks, but for time-series and multi-channel data as well. The system was designed primarily for workloads that build connectomes— neural connectivity maps of the brain—using the parallel execution of computer vision algorithms on high-performance compute clusters. These services and open-science data sets are publicly available at openconnecto.me. The system design inherits much from NoSQL scale-out and data-intensive computing architectures. We distribute data to cluster nodes by partitioning a spatial index. We direct I/O to different systems—reads to parallel disk arrays and writes to solid-state storage—to avoid I/O interference and maximize throughput. All programming interfaces are RESTful Web services, which are simple and stateless, improving scalability and usability. We include a performance evaluation of the production system, highlighting the effec-tiveness of spatial data organization. PMID:24401992

Acquisition of gamma camera and physiological data by computer.

PubMed

Hack, S N; Chang, M; Line, B R; Cooper, J A; Robeson, G H

1986-11-01

We have designed, implemented, and tested a new Research Data Acquisition System (RDAS) that permits a general purpose digital computer to acquire signals from both gamma camera sources and physiological signal sources concurrently. This system overcomes the limited multi-source, high speed data acquisition capabilities found in most clinically oriented nuclear medicine computers. The RDAS can simultaneously input signals from up to four gamma camera sources with a throughput of 200 kHz per source and from up to eight physiological signal sources with an aggregate throughput of 50 kHz. Rigorous testing has found the RDAS to exhibit acceptable linearity and timing characteristics. In addition, flood images obtained by this system were compared with flood images acquired by a commercial nuclear medicine computer system. National Electrical Manufacturers Association performance standards of the flood images were found to be comparable.

Assessing Morphological and Physiological Properties of Forest Species Using High Throughput Plant Phenotyping and Imaging Techniques

NASA Astrophysics Data System (ADS)

Mazis, A.; Hiller, J.; Morgan, P.; Awada, T.; Stoerger, V.

2017-12-01

High throughput plant phenotyping is increasingly being used to assess morphological and biophysical traits of economically important crops in agriculture. In this study, the potential application of this technique in natural resources management, through the characterization of woody plants regeneration, establishment, growth, and responses to water and nutrient manipulations was assessed. Two woody species were selected for this study, Quercus prinoides and Quercus bicolor. Seeds were collected from trees growing at the edge of their natural distribution in Nebraska and Missouri, USA. Seeds were germinated in the greenhouse and transferred to the Nebraska Innovation Campus Lemnatec3D High Throughput facility at the University of Nebraska-Lincoln. Seedlings subjected to water and N manipulations, were imaged twice or three times a week using four cameras (Visible, Fluorescence, Infrared and Hyperspectral), throughout the growing season. Traditional leaf to plant levels ecophysiological measurements were concurrently acquired to assess the relationship between these two techniques. These include gas exchange (LI 6400 and LI 6800, LICOR Inc., Lincoln NE), chlorophyll content, optical characteristics (Ocean Optics USB200), water and osmotic potentials, leaf area and weight and carbon isotope ratio. In the presentation, we highlight results on the potential use of high throughput plant phenotyping techniques to assess the morphology and physiology of woody species including responses to water availability and nutrient manipulation, and its broader application under field conditions and natural resources management. Also, we explore the different capabilities imaging provides us for modeling the plant physiological and morphological growth and how it can complement the current techniques

Predicting plant biomass accumulation from image-derived parameters

PubMed Central

Chen, Dijun; Shi, Rongli; Pape, Jean-Michel; Neumann, Kerstin; Graner, Andreas; Chen, Ming; Klukas, Christian

2018-01-01

Abstract Background Image-based high-throughput phenotyping technologies have been rapidly developed in plant science recently, and they provide a great potential to gain more valuable information than traditionally destructive methods. Predicting plant biomass is regarded as a key purpose for plant breeders and ecologists. However, it is a great challenge to find a predictive biomass model across experiments. Results In the present study, we constructed 4 predictive models to examine the quantitative relationship between image-based features and plant biomass accumulation. Our methodology has been applied to 3 consecutive barley (Hordeum vulgare) experiments with control and stress treatments. The results proved that plant biomass can be accurately predicted from image-based parameters using a random forest model. The high prediction accuracy based on this model will contribute to relieving the phenotyping bottleneck in biomass measurement in breeding applications. The prediction performance is still relatively high across experiments under similar conditions. The relative contribution of individual features for predicting biomass was further quantified, revealing new insights into the phenotypic determinants of the plant biomass outcome. Furthermore, methods could also be used to determine the most important image-based features related to plant biomass accumulation, which would be promising for subsequent genetic mapping to uncover the genetic basis of biomass. Conclusions We have developed quantitative models to accurately predict plant biomass accumulation from image data. We anticipate that the analysis results will be useful to advance our views of the phenotypic determinants of plant biomass outcome, and the statistical methods can be broadly used for other plant species. PMID:29346559

Three-dimensional microCT imaging of murine embryonic development from immediate post-implantation to organogenesis: application for phenotyping analysis of early embryonic lethality in mutant animals.

PubMed

Ermakova, Olga; Orsini, Tiziana; Gambadoro, Alessia; Chiani, Francesco; Tocchini-Valentini, Glauco P

2018-04-01

In this work, we applied three-dimensional microCT imaging to study murine embryogenesis in the range from immediate post-implantation period (embryonic day 5.5) to mid-gestation (embryonic day 12.5) with the resolution up to 1.4 µm/voxel. Also, we introduce an imaging procedure for non-invasive volumetric estimation of an entire litter of embryos within the maternal uterine structures. This method allows for an accurate, detailed and systematic morphometric analysis of both embryonic and extra-embryonic components during embryogenesis. Three-dimensional imaging of unperturbed embryos was performed to visualize the egg cylinder, primitive streak, gastrulation and early organogenesis stages of murine development in the C57Bl6/N mouse reference strain. Further, we applied our microCT imaging protocol to determine the earliest point when embryonic development is arrested in a mouse line with knockout for tRNA splicing endonuclease subunit Tsen54 gene. Our analysis determined that the embryonic development in Tsen54 null embryos does not proceed beyond implantation. We demonstrated that application of microCT imaging to entire litter of non-perturbed embryos greatly facilitate studies to unravel gene function during early embryogenesis and to determine the precise point at which embryonic development is arrested in mutant animals. The described method is inexpensive, does not require lengthy embryos dissection and can be applicable for detailed analysis of mutant mice at laboratory scale as well as for high-throughput projects.

High-throughput electrical characterization for robust overlay lithography control

NASA Astrophysics Data System (ADS)

Devender, Devender; Shen, Xumin; Duggan, Mark; Singh, Sunil; Rullan, Jonathan; Choo, Jae; Mehta, Sohan; Tang, Teck Jung; Reidy, Sean; Holt, Jonathan; Kim, Hyung Woo; Fox, Robert; Sohn, D. K.

2017-03-01

Realizing sensitive, high throughput and robust overlay measurement is a challenge in current 14nm and advanced upcoming nodes with transition to 300mm and upcoming 450mm semiconductor manufacturing, where slight deviation in overlay has significant impact on reliability and yield1). Exponentially increasing number of critical masks in multi-patterning lithoetch, litho-etch (LELE) and subsequent LELELE semiconductor processes require even tighter overlay specification2). Here, we discuss limitations of current image- and diffraction- based overlay measurement techniques to meet these stringent processing requirements due to sensitivity, throughput and low contrast3). We demonstrate a new electrical measurement based technique where resistance is measured for a macro with intentional misalignment between two layers. Overlay is quantified by a parabolic fitting model to resistance where minima and inflection points are extracted to characterize overlay control and process window, respectively. Analyses using transmission electron microscopy show good correlation between actual overlay performance and overlay obtained from fitting. Additionally, excellent correlation of overlay from electrical measurements to existing image- and diffraction- based techniques is found. We also discuss challenges of integrating electrical measurement based approach in semiconductor manufacturing from Back End of Line (BEOL) perspective. Our findings open up a new pathway for accessing simultaneous overlay as well as process window and margins from a robust, high throughput and electrical measurement approach.

The impact of the condenser on cytogenetic image quality in digital microscope system.

PubMed

Ren, Liqiang; Li, Zheng; Li, Yuhua; Zheng, Bin; Li, Shibo; Chen, Xiaodong; Liu, Hong

2013-01-01

Optimizing operational parameters of the digital microscope system is an important technique to acquire high quality cytogenetic images and facilitate the process of karyotyping so that the efficiency and accuracy of diagnosis can be improved. This study investigated the impact of the condenser on cytogenetic image quality and system working performance using a prototype digital microscope image scanning system. Both theoretical analysis and experimental validations through objectively evaluating a resolution test chart and subjectively observing large numbers of specimen were conducted. The results show that the optimal image quality and large depth of field (DOF) are simultaneously obtained when the numerical aperture of condenser is set as 60%-70% of the corresponding objective. Under this condition, more analyzable chromosomes and diagnostic information are obtained. As a result, the system shows higher working stability and less restriction for the implementation of algorithms such as autofocusing especially when the system is designed to achieve high throughput continuous image scanning. Although the above quantitative results were obtained using a specific prototype system under the experimental conditions reported in this paper, the presented evaluation methodologies can provide valuable guidelines for optimizing operational parameters in cytogenetic imaging using the high throughput continuous scanning microscopes in clinical practice.

[Research on Spectral Polarization Imaging System Based on Static Modulation].

PubMed

Zhao, Hai-bo; Li, Huan; Lin, Xu-ling; Wang, Zheng

2015-04-01

The main disadvantages of traditional spectral polarization imaging system are: complex structure, with moving parts, low throughput. A novel method of spectral polarization imaging system is discussed, which is based on static polarization intensity modulation combined with Savart polariscope interference imaging. The imaging system can obtain real-time information of spectral and four Stokes polarization messages. Compared with the conventional methods, the advantages of the imaging system are compactness, low mass and no moving parts, no electrical control, no slit and big throughput. The system structure and the basic theory are introduced. The experimental system is established in the laboratory. The experimental system consists of reimaging optics, polarization intensity module, interference imaging module, and CCD data collecting and processing module. The spectral range is visible and near-infrared (480-950 nm). The white board and the plane toy are imaged by using the experimental system. The ability of obtaining spectral polarization imaging information is verified. The calibration system of static polarization modulation is set up. The statistical error of polarization degree detection is less than 5%. The validity and feasibility of the basic principle is proved by the experimental result. The spectral polarization data captured by the system can be applied to object identification, object classification and remote sensing detection.

Automated Analysis of siRNA Screens of Virus Infected Cells Based on Immunofluorescence Microscopy

NASA Astrophysics Data System (ADS)

Matula, Petr; Kumar, Anil; Wörz, Ilka; Harder, Nathalie; Erfle, Holger; Bartenschlager, Ralf; Eils, Roland; Rohr, Karl

We present an image analysis approach as part of a high-throughput microscopy screening system based on cell arrays for the identification of genes involved in Hepatitis C and Dengue virus replication. Our approach comprises: cell nucleus segmentation, quantification of virus replication level in cells, localization of regions with transfected cells, cell classification by infection status, and quality assessment of an experiment. The approach is fully automatic and has been successfully applied to a large number of cell array images from screening experiments. The experimental results show a good agreement with the expected behavior of positive as well as negative controls and encourage the application to screens from further high-throughput experiments.

High-throughput quantification of hydroxyproline for determination of collagen.

PubMed

Hofman, Kathleen; Hall, Bronwyn; Cleaver, Helen; Marshall, Susan

2011-10-15

An accurate and high-throughput assay for collagen is essential for collagen research and development of collagen products. Hydroxyproline is routinely assayed to provide a measurement for collagen quantification. The time required for sample preparation using acid hydrolysis and neutralization prior to assay is what limits the current method for determining hydroxyproline. This work describes the conditions of alkali hydrolysis that, when combined with the colorimetric assay defined by Woessner, provide a high-throughput, accurate method for the measurement of hydroxyproline. Copyright © 2011 Elsevier Inc. All rights reserved.

Extracting Fluorescent Reporter Time Courses of Cell Lineages from High-Throughput Microscopy at Low Temporal Resolution

PubMed Central

Downey, Mike J.; Jeziorska, Danuta M.; Ott, Sascha; Tamai, T. Katherine; Koentges, Georgy; Vance, Keith W.; Bretschneider, Till

2011-01-01

The extraction of fluorescence time course data is a major bottleneck in high-throughput live-cell microscopy. Here we present an extendible framework based on the open-source image analysis software ImageJ, which aims in particular at analyzing the expression of fluorescent reporters through cell divisions. The ability to track individual cell lineages is essential for the analysis of gene regulatory factors involved in the control of cell fate and identity decisions. In our approach, cell nuclei are identified using Hoechst, and a characteristic drop in Hoechst fluorescence helps to detect dividing cells. We first compare the efficiency and accuracy of different segmentation methods and then present a statistical scoring algorithm for cell tracking, which draws on the combination of various features, such as nuclear intensity, area or shape, and importantly, dynamic changes thereof. Principal component analysis is used to determine the most significant features, and a global parameter search is performed to determine the weighting of individual features. Our algorithm has been optimized to cope with large cell movements, and we were able to semi-automatically extract cell trajectories across three cell generations. Based on the MTrackJ plugin for ImageJ, we have developed tools to efficiently validate tracks and manually correct them by connecting broken trajectories and reassigning falsely connected cell positions. A gold standard consisting of two time-series with 15,000 validated positions will be released as a valuable resource for benchmarking. We demonstrate how our method can be applied to analyze fluorescence distributions generated from mouse stem cells transfected with reporter constructs containing transcriptional control elements of the Msx1 gene, a regulator of pluripotency, in mother and daughter cells. Furthermore, we show by tracking zebrafish PAC2 cells expressing FUCCI cell cycle markers, our framework can be easily adapted to different cell types and fluorescent markers. PMID:22194797

High Performance Computing Modernization Program Kerberos Throughput Test Report

DTIC Science & Technology

2017-10-26

functionality as Kerberos plugins. The pre -release production kit was used in these tests to compare against the current release kit. YubiKey support...HPCMP Kerberos Throughput Test Report 3 2. THROUGHPUT TESTING 2.1 Testing Components Throughput testing was done to determine the benefits of the pre ...both the current release kit and the pre -release production kit for a total of 378 individual tests in order to note any improvements. Based on work

Unsupervised Clustering of Subcellular Protein Expression Patterns in High-Throughput Microscopy Images Reveals Protein Complexes and Functional Relationships between Proteins

PubMed Central

Handfield, Louis-François; Chong, Yolanda T.; Simmons, Jibril; Andrews, Brenda J.; Moses, Alan M.

2013-01-01

Protein subcellular localization has been systematically characterized in budding yeast using fluorescently tagged proteins. Based on the fluorescence microscopy images, subcellular localization of many proteins can be classified automatically using supervised machine learning approaches that have been trained to recognize predefined image classes based on statistical features. Here, we present an unsupervised analysis of protein expression patterns in a set of high-resolution, high-throughput microscope images. Our analysis is based on 7 biologically interpretable features which are evaluated on automatically identified cells, and whose cell-stage dependency is captured by a continuous model for cell growth. We show that it is possible to identify most previously identified localization patterns in a cluster analysis based on these features and that similarities between the inferred expression patterns contain more information about protein function than can be explained by a previous manual categorization of subcellular localization. Furthermore, the inferred cell-stage associated to each fluorescence measurement allows us to visualize large groups of proteins entering the bud at specific stages of bud growth. These correspond to proteins localized to organelles, revealing that the organelles must be entering the bud in a stereotypical order. We also identify and organize a smaller group of proteins that show subtle differences in the way they move around the bud during growth. Our results suggest that biologically interpretable features based on explicit models of cell morphology will yield unprecedented power for pattern discovery in high-resolution, high-throughput microscopy images. PMID:23785265

Conventional and hyperspectral time-series imaging of maize lines widely used in field trials

PubMed Central

Liang, Zhikai; Pandey, Piyush; Stoerger, Vincent; Xu, Yuhang; Qiu, Yumou; Ge, Yufeng

2018-01-01

Abstract Background Maize (Zea mays ssp. mays) is 1 of 3 crops, along with rice and wheat, responsible for more than one-half of all calories consumed around the world. Increasing the yield and stress tolerance of these crops is essential to meet the growing need for food. The cost and speed of plant phenotyping are currently the largest constraints on plant breeding efforts. Datasets linking new types of high-throughput phenotyping data collected from plants to the performance of the same genotypes under agronomic conditions across a wide range of environments are essential for developing new statistical approaches and computer vision–based tools. Findings A set of maize inbreds—primarily recently off patent lines—were phenotyped using a high-throughput platform at University of Nebraska-Lincoln. These lines have been previously subjected to high-density genotyping and scored for a core set of 13 phenotypes in field trials across 13 North American states in 2 years by the Genomes 2 Fields Consortium. A total of 485 GB of image data including RGB, hyperspectral, fluorescence, and thermal infrared photos has been released. Conclusions Correlations between image-based measurements and manual measurements demonstrated the feasibility of quantifying variation in plant architecture using image data. However, naive approaches to measuring traits such as biomass can introduce nonrandom measurement errors confounded with genotype variation. Analysis of hyperspectral image data demonstrated unique signatures from stem tissue. Integrating heritable phenotypes from high-throughput phenotyping data with field data from different environments can reveal previously unknown factors that influence yield plasticity. PMID:29186425

Conventional and hyperspectral time-series imaging of maize lines widely used in field trials.

PubMed

Liang, Zhikai; Pandey, Piyush; Stoerger, Vincent; Xu, Yuhang; Qiu, Yumou; Ge, Yufeng; Schnable, James C

2018-02-01

Maize (Zea mays ssp. mays) is 1 of 3 crops, along with rice and wheat, responsible for more than one-half of all calories consumed around the world. Increasing the yield and stress tolerance of these crops is essential to meet the growing need for food. The cost and speed of plant phenotyping are currently the largest constraints on plant breeding efforts. Datasets linking new types of high-throughput phenotyping data collected from plants to the performance of the same genotypes under agronomic conditions across a wide range of environments are essential for developing new statistical approaches and computer vision-based tools. A set of maize inbreds-primarily recently off patent lines-were phenotyped using a high-throughput platform at University of Nebraska-Lincoln. These lines have been previously subjected to high-density genotyping and scored for a core set of 13 phenotypes in field trials across 13 North American states in 2 years by the Genomes 2 Fields Consortium. A total of 485 GB of image data including RGB, hyperspectral, fluorescence, and thermal infrared photos has been released. Correlations between image-based measurements and manual measurements demonstrated the feasibility of quantifying variation in plant architecture using image data. However, naive approaches to measuring traits such as biomass can introduce nonrandom measurement errors confounded with genotype variation. Analysis of hyperspectral image data demonstrated unique signatures from stem tissue. Integrating heritable phenotypes from high-throughput phenotyping data with field data from different environments can reveal previously unknown factors that influence yield plasticity. © The Authors 2017. Published by Oxford University Press.

Nile Red Detection of Bacterial Hydrocarbons and Ketones in a High-Throughput Format

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pinzon, NM; Aukema, KG; Gralnick, JA

A method for use in high-throughput screening of bacteria for the production of long-chain hydrocarbons and ketones by monitoring fluorescent light emission in the presence of Nile red is described. Nile red has previously been used to screen for polyhydroxybutyrate (PHB) and fatty acid esters, but this is the first report of screening for recombinant bacteria making hydrocarbons or ketones. The microtiter plate assay was evaluated using wild-type and recombinant strains of Shewanella oneidensis and Escherichia coli expressing the enzyme OleA, previously shown to initiate hydrocarbon biosynthesis. The strains expressing exogenous Stenotrophomonas maltophilia oleA, with increased levels of ketone productionmore » as determined by gas chromatography-mass spectrometry, were distinguished with Nile red fluorescence. Confocal microscopy images of S. oneidensis oleA-expressing strains stained with Nile red were consistent with a membrane localization of the ketones. This differed from Nile red staining of bacterial PHB or algal lipid droplets that showed intracellular inclusion bodies. These results demonstrated the applicability of Nile red in a high-throughput technique for the detection of bacterial hydrocarbons and ketones. IMPORTANCE In recent years, there has been renewed interest in advanced biofuel sources such as bacterial hydrocarbon production. Previous studies used solvent extraction of bacterial cultures followed by gas chromatography-mass spectrometry (GC-MS) to detect and quantify ketones and hydrocarbons (Beller HR, Goh EB, Keasling JD, Appl. Environ. Microbiol. 76: 1212-1223, 2010; Sukovich DJ, Seffernick JL, Richman JE, Gralnick JA, Wackett LP, Appl. Environ. Microbiol. 76: 3850-3862, 2010). While these analyses are powerful and accurate, their labor-intensive nature makes them intractable to high-throughput screening; therefore, methods for rapid identification of bacterial strains that are overproducing hydrocarbons are needed. The use of high-throughput evaluation of bacterial and algal hydrophobic molecule production via Nile red fluorescence from lipids and esters was extended in this study to include hydrocarbons and ketones. This work demonstrated accurate, high-throughput detection of high-level bacterial long-chain ketone and hydrocarbon production by screening for increased fluorescence of the hydrophobic dye Nile red.« less

RootGraph: a graphic optimization tool for automated image analysis of plant roots

PubMed Central

Cai, Jinhai; Zeng, Zhanghui; Connor, Jason N.; Huang, Chun Yuan; Melino, Vanessa; Kumar, Pankaj; Miklavcic, Stanley J.

2015-01-01

This paper outlines a numerical scheme for accurate, detailed, and high-throughput image analysis of plant roots. In contrast to existing root image analysis tools that focus on root system-average traits, a novel, fully automated and robust approach for the detailed characterization of root traits, based on a graph optimization process is presented. The scheme, firstly, distinguishes primary roots from lateral roots and, secondly, quantifies a broad spectrum of root traits for each identified primary and lateral root. Thirdly, it associates lateral roots and their properties with the specific primary root from which the laterals emerge. The performance of this approach was evaluated through comparisons with other automated and semi-automated software solutions as well as against results based on manual measurements. The comparisons and subsequent application of the algorithm to an array of experimental data demonstrate that this method outperforms existing methods in terms of accuracy, robustness, and the ability to process root images under high-throughput conditions. PMID:26224880

A High-Content Live-Cell Viability Assay and Its Validation on a Diverse 12K Compound Screen.

PubMed

Chiaravalli, Jeanne; Glickman, J Fraser

2017-08-01

We have developed a new high-content cytotoxicity assay using live cells, called "ImageTOX." We used a high-throughput fluorescence microscope system, image segmentation software, and the combination of Hoechst 33342 and SYTO 17 to simultaneously score the relative size and the intensity of the nuclei, the nuclear membrane permeability, and the cell number in a 384-well microplate format. We then performed a screen of 12,668 diverse compounds and compared the results to a standard cytotoxicity assay. The ImageTOX assay identified similar sets of compounds to the standard cytotoxicity assay, while identifying more compounds having adverse effects on cell structure, earlier in treatment time. The ImageTOX assay uses inexpensive commercially available reagents and facilitates the use of live cells in toxicity screens. Furthermore, we show that we can measure the kinetic profile of compound toxicity in a high-content, high-throughput format, following the same set of cells over an extended period of time.

Identification of Novel Compounds Inhibiting Chikungunya Virus-Induced Cell Death by High Throughput Screening of a Kinase Inhibitor Library

PubMed Central

Gomes, Rafael G. B.; da Silva, Camila T.; Taniguchi, Juliana B.; No, Joo Hwan; Lombardot, Benoit; Schwartz, Olivier; Hansen, Michael A. E.; Freitas-Junior, Lucio H.

2013-01-01

Chikungunya virus (CHIKV) is a mosquito-borne arthrogenic alphavirus that causes acute febrile illness in humans accompanied by joint pains and in many cases, persistent arthralgia lasting weeks to years. The re-emergence of CHIKV has resulted in numerous outbreaks in the eastern hemisphere, and threatens to expand in the foreseeable future. Unfortunately, no effective treatment is currently available. The present study reports the use of resazurin in a cell-based high-throughput assay, and an image-based high-content assay to identify and characterize inhibitors of CHIKV-infection in vitro. CHIKV is a highly cytopathic virus that rapidly kills infected cells. Thus, cell viability of HuH-7 cells infected with CHIKV in the presence of compounds was determined by measuring metabolic reduction of resazurin to identify inhibitors of CHIKV-associated cell death. A kinase inhibitor library of 4,000 compounds was screened against CHIKV infection of HuH-7 cells using the resazurin reduction assay, and the cell toxicity was also measured in non-infected cells. Seventy-two compounds showing ≥50% inhibition property against CHIKV at 10 µM were selected as primary hits. Four compounds having a benzofuran core scaffold (CND0335, CND0364, CND0366 and CND0415), one pyrrolopyridine (CND0545) and one thiazol-carboxamide (CND3514) inhibited CHIKV-associated cell death in a dose-dependent manner, with EC50 values between 2.2 µM and 7.1 µM. Based on image analysis, these 6 hit compounds did not inhibit CHIKV replication in the host cell. However, CHIKV-infected cells manifested less prominent apoptotic blebs typical of CHIKV cytopathic effect compared with the control infection. Moreover, treatment with these compounds reduced viral titers in the medium of CHIKV-infected cells by up to 100-fold. In conclusion, this cell-based high-throughput screening assay using resazurin, combined with the image-based high content assay approach identified compounds against CHIKV having a novel antiviral activity - inhibition of virus-induced CPE - likely by targeting kinases involved in apoptosis. PMID:24205414

Automated Interpretation of Subcellular Patterns in Fluorescence Microscope Images for Location Proteomics

PubMed Central

Chen, Xiang; Velliste, Meel; Murphy, Robert F.

2010-01-01

Proteomics, the large scale identification and characterization of many or all proteins expressed in a given cell type, has become a major area of biological research. In addition to information on protein sequence, structure and expression levels, knowledge of a protein’s subcellular location is essential to a complete understanding of its functions. Currently subcellular location patterns are routinely determined by visual inspection of fluorescence microscope images. We review here research aimed at creating systems for automated, systematic determination of location. These employ numerical feature extraction from images, feature reduction to identify the most useful features, and various supervised learning (classification) and unsupervised learning (clustering) methods. These methods have been shown to perform significantly better than human interpretation of the same images. When coupled with technologies for tagging large numbers of proteins and high-throughput microscope systems, the computational methods reviewed here enable the new subfield of location proteomics. This subfield will make critical contributions in two related areas. First, it will provide structured, high-resolution information on location to enable Systems Biology efforts to simulate cell behavior from the gene level on up. Second, it will provide tools for Cytomics projects aimed at characterizing the behaviors of all cell types before, during and after the onset of various diseases. PMID:16752421

Light-sheet microscopy for quantitative ovarian folliculometry

NASA Astrophysics Data System (ADS)

Lin, Hsiao-Chun Amy; Dutta, Rahul; Mandal, Subhamoy; Kind, Alexander; Schnieke, Angelika; Razansky, Daniel

2017-02-01

Determination of ovarian status and follicle monitoring are common methods of diagnosing female infertility. We evaluated the suitability of selective plane illumination microscopy (SPIM) for the study of ovarian follicles. Owing to the large field of view and fast acquisition speed of our newly developed SPIM system, volumetric image stacks from entire intact samples of pig ovaries have been rendered demonstrating clearly discernible follicular features like follicle diameters (70 μm - 2.5 mm), size of developing Cumulus oophorus complexes (COC ) (40 μm - 110 μm), and follicular wall thicknesses (90 μm-120 μm). The observation of clearly distinguishable COCs protruding into the follicular antrum was also shown possible, and correlation with the developmental stage of the follicles was determined. Follicles of all developmental stages were identified, and even the small primordial follicle clusters forming the egg nest could be observed. The ability of the system to non-destructively generate sub-cellular resolution 3D images of developing follicles, with excellent image contrast and high throughput capacity compared to conventional histology, suggests that it can be used to monitor follicular development and identify structural abnormalities indicative of ovarian ailments. Accurate folliculometric measurements provided by SPIM images can immensely help the understanding of ovarian physiology and provide important information for the proper management of ovarian diseases.

Label-free high-throughput detection and quantification of circulating melanoma tumor cell clusters by linear-array-based photoacoustic tomography

NASA Astrophysics Data System (ADS)

Hai, Pengfei; Zhou, Yong; Zhang, Ruiying; Ma, Jun; Li, Yang; Shao, Jin-Yu; Wang, Lihong V.

2017-04-01

Circulating tumor cell (CTC) clusters, arising from multicellular groupings in a primary tumor, greatly elevate the metastatic potential of cancer compared with single CTCs. High-throughput detection and quantification of CTC clusters are important for understanding the tumor metastatic process and improving cancer therapy. Here, we applied a linear-array-based photoacoustic tomography (LA-PAT) system and improved the image reconstruction for label-free high-throughput CTC cluster detection and quantification in vivo. The feasibility was first demonstrated by imaging CTC cluster ex vivo. The relationship between the contrast-to-noise ratios (CNRs) and the number of cells in melanoma tumor cell clusters was investigated and verified. Melanoma CTC clusters with a minimum of four cells could be detected, and the number of cells could be computed from the CNR. Finally, we demonstrated imaging of injected melanoma CTC clusters in rats in vivo. Similarly, the number of cells in the melanoma CTC clusters could be quantified. The data showed that larger CTC clusters had faster clearance rates in the bloodstream, which agreed with the literature. The results demonstrated the capability of LA-PAT to detect and quantify melanoma CTC clusters in vivo and showed its potential for tumor metastasis study and cancer therapy.

High-Throughput In Vivo Genotoxicity Testing: An Automated Readout System for the Somatic Mutation and Recombination Test (SMART)

PubMed Central

Kwak, Jihoon; Genovesio, Auguste; Kang, Myungjoo; Hansen, Michael Adsett Edberg; Han, Sung-Jun

2015-01-01

Genotoxicity testing is an important component of toxicity assessment. As illustrated by the European registration, evaluation, authorization, and restriction of chemicals (REACH) directive, it concerns all the chemicals used in industry. The commonly used in vivo mammalian tests appear to be ill adapted to tackle the large compound sets involved, due to throughput, cost, and ethical issues. The somatic mutation and recombination test (SMART) represents a more scalable alternative, since it uses Drosophila, which develops faster and requires less infrastructure. Despite these advantages, the manual scoring of the hairs on Drosophila wings required for the SMART limits its usage. To overcome this limitation, we have developed an automated SMART readout. It consists of automated imaging, followed by an image analysis pipeline that measures individual wing genotoxicity scores. Finally, we have developed a wing score-based dose-dependency approach that can provide genotoxicity profiles. We have validated our method using 6 compounds, obtaining profiles almost identical to those obtained from manual measures, even for low-genotoxicity compounds such as urethane. The automated SMART, with its faster and more reliable readout, fulfills the need for a high-throughput in vivo test. The flexible imaging strategy we describe and the analysis tools we provide should facilitate the optimization and dissemination of our methods. PMID:25830368

GPU Lossless Hyperspectral Data Compression System

NASA Technical Reports Server (NTRS)

Aranki, Nazeeh I.; Keymeulen, Didier; Kiely, Aaron B.; Klimesh, Matthew A.

2014-01-01

Hyperspectral imaging systems onboard aircraft or spacecraft can acquire large amounts of data, putting a strain on limited downlink and storage resources. Onboard data compression can mitigate this problem but may require a system capable of a high throughput. In order to achieve a high throughput with a software compressor, a graphics processing unit (GPU) implementation of a compressor was developed targeting the current state-of-the-art GPUs from NVIDIA(R). The implementation is based on the fast lossless (FL) compression algorithm reported in "Fast Lossless Compression of Multispectral-Image Data" (NPO- 42517), NASA Tech Briefs, Vol. 30, No. 8 (August 2006), page 26, which operates on hyperspectral data and achieves excellent compression performance while having low complexity. The FL compressor uses an adaptive filtering method and achieves state-of-the-art performance in both compression effectiveness and low complexity. The new Consultative Committee for Space Data Systems (CCSDS) Standard for Lossless Multispectral & Hyperspectral image compression (CCSDS 123) is based on the FL compressor. The software makes use of the highly-parallel processing capability of GPUs to achieve a throughput at least six times higher than that of a software implementation running on a single-core CPU. This implementation provides a practical real-time solution for compression of data from airborne hyperspectral instruments.

High Throughput Multispectral Image Processing with Applications in Food Science.

PubMed

Tsakanikas, Panagiotis; Pavlidis, Dimitris; Nychas, George-John

2015-01-01

Recently, machine vision is gaining attention in food science as well as in food industry concerning food quality assessment and monitoring. Into the framework of implementation of Process Analytical Technology (PAT) in the food industry, image processing can be used not only in estimation and even prediction of food quality but also in detection of adulteration. Towards these applications on food science, we present here a novel methodology for automated image analysis of several kinds of food products e.g. meat, vanilla crème and table olives, so as to increase objectivity, data reproducibility, low cost information extraction and faster quality assessment, without human intervention. Image processing's outcome will be propagated to the downstream analysis. The developed multispectral image processing method is based on unsupervised machine learning approach (Gaussian Mixture Models) and a novel unsupervised scheme of spectral band selection for segmentation process optimization. Through the evaluation we prove its efficiency and robustness against the currently available semi-manual software, showing that the developed method is a high throughput approach appropriate for massive data extraction from food samples.

High performance computing environment for multidimensional image analysis

PubMed Central

Rao, A Ravishankar; Cecchi, Guillermo A; Magnasco, Marcelo

2007-01-01

Background The processing of images acquired through microscopy is a challenging task due to the large size of datasets (several gigabytes) and the fast turnaround time required. If the throughput of the image processing stage is significantly increased, it can have a major impact in microscopy applications. Results We present a high performance computing (HPC) solution to this problem. This involves decomposing the spatial 3D image into segments that are assigned to unique processors, and matched to the 3D torus architecture of the IBM Blue Gene/L machine. Communication between segments is restricted to the nearest neighbors. When running on a 2 Ghz Intel CPU, the task of 3D median filtering on a typical 256 megabyte dataset takes two and a half hours, whereas by using 1024 nodes of Blue Gene, this task can be performed in 18.8 seconds, a 478× speedup. Conclusion Our parallel solution dramatically improves the performance of image processing, feature extraction and 3D reconstruction tasks. This increased throughput permits biologists to conduct unprecedented large scale experiments with massive datasets. PMID:17634099

High performance computing environment for multidimensional image analysis.

PubMed

Rao, A Ravishankar; Cecchi, Guillermo A; Magnasco, Marcelo

2007-07-10

The processing of images acquired through microscopy is a challenging task due to the large size of datasets (several gigabytes) and the fast turnaround time required. If the throughput of the image processing stage is significantly increased, it can have a major impact in microscopy applications. We present a high performance computing (HPC) solution to this problem. This involves decomposing the spatial 3D image into segments that are assigned to unique processors, and matched to the 3D torus architecture of the IBM Blue Gene/L machine. Communication between segments is restricted to the nearest neighbors. When running on a 2 Ghz Intel CPU, the task of 3D median filtering on a typical 256 megabyte dataset takes two and a half hours, whereas by using 1024 nodes of Blue Gene, this task can be performed in 18.8 seconds, a 478x speedup. Our parallel solution dramatically improves the performance of image processing, feature extraction and 3D reconstruction tasks. This increased throughput permits biologists to conduct unprecedented large scale experiments with massive datasets.

Integrating medical imaging analyses through a high-throughput bundled resource imaging system

NASA Astrophysics Data System (ADS)

Covington, Kelsie; Welch, E. Brian; Jeong, Ha-Kyu; Landman, Bennett A.

2011-03-01

Exploitation of advanced, PACS-centric image analysis and interpretation pipelines provides well-developed storage, retrieval, and archival capabilities along with state-of-the-art data providence, visualization, and clinical collaboration technologies. However, pursuit of integrated medical imaging analysis through a PACS environment can be limiting in terms of the overhead required to validate, evaluate and integrate emerging research technologies. Herein, we address this challenge through presentation of a high-throughput bundled resource imaging system (HUBRIS) as an extension to the Philips Research Imaging Development Environment (PRIDE). HUBRIS enables PACS-connected medical imaging equipment to invoke tools provided by the Java Imaging Science Toolkit (JIST) so that a medical imaging platform (e.g., a magnetic resonance imaging scanner) can pass images and parameters to a server, which communicates with a grid computing facility to invoke the selected algorithms. Generated images are passed back to the server and subsequently to the imaging platform from which the images can be sent to a PACS. JIST makes use of an open application program interface layer so that research technologies can be implemented in any language capable of communicating through a system shell environment (e.g., Matlab, Java, C/C++, Perl, LISP, etc.). As demonstrated in this proof-of-concept approach, HUBRIS enables evaluation and analysis of emerging technologies within well-developed PACS systems with minimal adaptation of research software, which simplifies evaluation of new technologies in clinical research and provides a more convenient use of PACS technology by imaging scientists.

Application of DNA Chip Scanning Technology for Automatic Detection of Chlamydia trachomatis and Chlamydia pneumoniae Inclusions

PubMed Central

Bogdanov, Anita; Endrész, Valeria; Urbán, Szabolcs; Lantos, Ildikó; Deák, Judit; Burián, Katalin; Önder, Kamil; Ayaydin, Ferhan; Balázs, Péter

2014-01-01

Chlamydiae are obligate intracellular bacteria that propagate in the inclusion, a specific niche inside the host cell. The standard method for counting chlamydiae is immunofluorescent staining and manual counting of chlamydial inclusions. High- or medium-throughput estimation of the reduction in chlamydial inclusions should be the basis of testing antichlamydial compounds and other drugs that positively or negatively influence chlamydial growth, yet low-throughput manual counting is the common approach. To overcome the time-consuming and subjective manual counting, we developed an automatic inclusion-counting system based on a commercially available DNA chip scanner. Fluorescently labeled inclusions are detected by the scanner, and the image is processed by ChlamyCount, a custom plug-in of the ImageJ software environment. ChlamyCount was able to measure the inclusion counts over a 1-log-unit dynamic range with a high correlation to the theoretical counts. ChlamyCount was capable of accurately determining the MICs of the novel antimicrobial compound PCC00213 and the already known antichlamydial antibiotics moxifloxacin and tetracycline. ChlamyCount was also able to measure the chlamydial growth-altering effect of drugs that influence host-bacterium interaction, such as gamma interferon, DEAE-dextran, and cycloheximide. ChlamyCount is an easily adaptable system for testing antichlamydial antimicrobials and other compounds that influence Chlamydia-host interactions. PMID:24189259

Comparison of pre-processing techniques for fluorescence microscopy images of cells labeled for actin.

PubMed

Muralidhar, Gautam S; Channappayya, Sumohana S; Slater, John H; Blinka, Ellen M; Bovik, Alan C; Frey, Wolfgang; Markey, Mia K

2008-11-06

Automated analysis of fluorescence microscopy images of endothelial cells labeled for actin is important for quantifying changes in the actin cytoskeleton. The current manual approach is laborious and inefficient. The goal of our work is to develop automated image analysis methods, thereby increasing cell analysis throughput. In this study, we present preliminary results on comparing different algorithms for cell segmentation and image denoising.

Hardware accelerator of convolution with exponential function for image processing applications

NASA Astrophysics Data System (ADS)

Panchenko, Ivan; Bucha, Victor

2015-12-01

In this paper we describe a Hardware Accelerator (HWA) for fast recursive approximation of separable convolution with exponential function. This filter can be used in many Image Processing (IP) applications, e.g. depth-dependent image blur, image enhancement and disparity estimation. We have adopted this filter RTL implementation to provide maximum throughput in constrains of required memory bandwidth and hardware resources to provide a power-efficient VLSI implementation.

A case for inherent geometric and geodetic accuracy in remotely sensed VNIR and SWIR imaging products

NASA Technical Reports Server (NTRS)

Driver, J. M.

1982-01-01

Significant aberrations can occur in acquired images which, unless compensated on board the spacecraft, can seriously impair throughput and timeliness for typical Earth observation missions. Conceptual compensations options are advanced to enable acquisition of images with inherent geometric and geodetic accuracy. Research needs are identified which, when implemented, can provide inherently accurate images. Agressive pursuit of these research needs is recommended.

High-throughput high-volume nuclear imaging for preclinical in vivo compound screening§.

PubMed

Macholl, Sven; Finucane, Ciara M; Hesterman, Jacob; Mather, Stephen J; Pauplis, Rachel; Scully, Deirdre; Sosabowski, Jane K; Jouannot, Erwan

2017-12-01

Preclinical single-photon emission computed tomography (SPECT)/CT imaging studies are hampered by low throughput, hence are found typically within small volume feasibility studies. Here, imaging and image analysis procedures are presented that allow profiling of a large volume of radiolabelled compounds within a reasonably short total study time. Particular emphasis was put on quality control (QC) and on fast and unbiased image analysis. 2-3 His-tagged proteins were simultaneously radiolabelled by 99m Tc-tricarbonyl methodology and injected intravenously (20 nmol/kg; 100 MBq; n = 3) into patient-derived xenograft (PDX) mouse models. Whole-body SPECT/CT images of 3 mice simultaneously were acquired 1, 4, and 24 h post-injection, extended to 48 h and/or by 0-2 h dynamic SPECT for pre-selected compounds. Organ uptake was quantified by automated multi-atlas and manual segmentations. Data were plotted automatically, quality controlled and stored on a collaborative image management platform. Ex vivo uptake data were collected semi-automatically and analysis performed as for imaging data. >500 single animal SPECT images were acquired for 25 proteins over 5 weeks, eventually generating >3500 ROI and >1000 items of tissue data. SPECT/CT images clearly visualized uptake in tumour and other tissues even at 48 h post-injection. Intersubject uptake variability was typically 13% (coefficient of variation, COV). Imaging results correlated well with ex vivo data. The large data set of tumour, background and systemic uptake/clearance data from 75 mice for 25 compounds allows identification of compounds of interest. The number of animals required was reduced considerably by longitudinal imaging compared to dissection experiments. All experimental work and analyses were accomplished within 3 months expected to be compatible with drug development programmes. QC along all workflow steps, blinding of the imaging contract research organization to compound properties and automation provide confidence in the data set. Additional ex vivo data were useful as a control but could be omitted from future studies in the same centre. For even larger compound libraries, radiolabelling could be expedited and the number of imaging time points adapted to increase weekly throughput. Multi-atlas segmentation could be expanded via SPECT/MRI; however, this would require an MRI-compatible mouse hotel. Finally, analysis of nuclear images of radiopharmaceuticals in clinical trials may benefit from the automated analysis procedures developed.

Optofluidic time-stretch quantitative phase microscopy.

PubMed

Guo, Baoshan; Lei, Cheng; Wu, Yi; Kobayashi, Hirofumi; Ito, Takuro; Yalikun, Yaxiaer; Lee, Sangwook; Isozaki, Akihiro; Li, Ming; Jiang, Yiyue; Yasumoto, Atsushi; Di Carlo, Dino; Tanaka, Yo; Yatomi, Yutaka; Ozeki, Yasuyuki; Goda, Keisuke

2018-03-01

Innovations in optical microscopy have opened new windows onto scientific research, industrial quality control, and medical practice over the last few decades. One of such innovations is optofluidic time-stretch quantitative phase microscopy - an emerging method for high-throughput quantitative phase imaging that builds on the interference between temporally stretched signal and reference pulses by using dispersive properties of light in both spatial and temporal domains in an interferometric configuration on a microfluidic platform. It achieves the continuous acquisition of both intensity and phase images with a high throughput of more than 10,000 particles or cells per second by overcoming speed limitations that exist in conventional quantitative phase imaging methods. Applications enabled by such capabilities are versatile and include characterization of cancer cells and microalgal cultures. In this paper, we review the principles and applications of optofluidic time-stretch quantitative phase microscopy and discuss its future perspective. Copyright © 2017 Elsevier Inc. All rights reserved.

Multiple-mouse MRI with multiple arrays of receive coils.

PubMed

Ramirez, Marc S; Esparza-Coss, Emilio; Bankson, James A

2010-03-01

Compared to traditional single-animal imaging methods, multiple-mouse MRI has been shown to dramatically improve imaging throughput and reduce the potentially prohibitive cost for instrument access. To date, up to a single radiofrequency coil has been dedicated to each animal being simultaneously scanned, thus limiting the sensitivity, flexibility, and ultimate throughput. The purpose of this study was to investigate the feasibility of multiple-mouse MRI with a phased-array coil dedicated to each animal. A dual-mouse imaging system, consisting of a pair of two-element phased-array coils, was developed and used to achieve acceleration factors greater than the number of animals scanned at once. By simultaneously scanning two mice with a retrospectively gated cardiac cine MRI sequence, a 3-fold acceleration was achieved with signal-to-noise ratio in the heart that is equivalent to that achieved with an unaccelerated scan using a commercial mouse birdcage coil. (c) 2010 Wiley-Liss, Inc.

High-Throughput Image Analysis of Fibrillar Materials: A Case Study on Polymer Nanofiber Packing, Alignment, and Defects in Organic Field Effect Transistors.

PubMed

Persson, Nils E; Rafshoon, Joshua; Naghshpour, Kaylie; Fast, Tony; Chu, Ping-Hsun; McBride, Michael; Risteen, Bailey; Grover, Martha; Reichmanis, Elsa

2017-10-18

High-throughput discovery of process-structure-property relationships in materials through an informatics-enabled empirical approach is an increasingly utilized technique in materials research due to the rapidly expanding availability of data. Here, process-structure-property relationships are extracted for the nucleation, growth, and deposition of semiconducting poly(3-hexylthiophene) (P3HT) nanofibers used in organic field effect transistors, via high-throughput image analysis. This study is performed using an automated image analysis pipeline combining existing open-source software and new algorithms, enabling the rapid evaluation of structural metrics for images of fibrillar materials, including local orientational order, fiber length density, and fiber length distributions. We observe that microfluidic processing leads to fibers that pack with unusually high density, while sonication yields fibers that pack sparsely with low alignment. This is attributed to differences in their crystallization mechanisms. P3HT nanofiber packing during thin film deposition exhibits behavior suggesting that fibers are confined to packing in two-dimensional layers. We find that fiber alignment, a feature correlated with charge carrier mobility, is driven by increasing fiber length, and that shorter fibers tend to segregate to the buried dielectric interface during deposition, creating potentially performance-limiting defects in alignment. Another barrier to perfect alignment is the curvature of P3HT fibers; we propose a mechanistic simulation of fiber growth that reconciles both this curvature and the log-normal distribution of fiber lengths inherent to the fiber populations under consideration.

Automated vector selection of SIVQ and parallel computing integration MATLAB™: Innovations supporting large-scale and high-throughput image analysis studies.

PubMed

Cheng, Jerome; Hipp, Jason; Monaco, James; Lucas, David R; Madabhushi, Anant; Balis, Ulysses J

2011-01-01

Spatially invariant vector quantization (SIVQ) is a texture and color-based image matching algorithm that queries the image space through the use of ring vectors. In prior studies, the selection of one or more optimal vectors for a particular feature of interest required a manual process, with the user initially stochastically selecting candidate vectors and subsequently testing them upon other regions of the image to verify the vector's sensitivity and specificity properties (typically by reviewing a resultant heat map). In carrying out the prior efforts, the SIVQ algorithm was noted to exhibit highly scalable computational properties, where each region of analysis can take place independently of others, making a compelling case for the exploration of its deployment on high-throughput computing platforms, with the hypothesis that such an exercise will result in performance gains that scale linearly with increasing processor count. An automated process was developed for the selection of optimal ring vectors to serve as the predicate matching operator in defining histopathological features of interest. Briefly, candidate vectors were generated from every possible coordinate origin within a user-defined vector selection area (VSA) and subsequently compared against user-identified positive and negative "ground truth" regions on the same image. Each vector from the VSA was assessed for its goodness-of-fit to both the positive and negative areas via the use of the receiver operating characteristic (ROC) transfer function, with each assessment resulting in an associated area-under-the-curve (AUC) figure of merit. Use of the above-mentioned automated vector selection process was demonstrated in two cases of use: First, to identify malignant colonic epithelium, and second, to identify soft tissue sarcoma. For both examples, a very satisfactory optimized vector was identified, as defined by the AUC metric. Finally, as an additional effort directed towards attaining high-throughput capability for the SIVQ algorithm, we demonstrated the successful incorporation of it with the MATrix LABoratory (MATLAB™) application interface. The SIVQ algorithm is suitable for automated vector selection settings and high throughput computation.

Fully-automated, high-throughput micro-computed tomography analysis of body composition enables therapeutic efficacy monitoring in preclinical models.

PubMed

Wyatt, S K; Barck, K H; Kates, L; Zavala-Solorio, J; Ross, J; Kolumam, G; Sonoda, J; Carano, R A D

2015-11-01

The ability to non-invasively measure body composition in mouse models of obesity and obesity-related disorders is essential for elucidating mechanisms of metabolic regulation and monitoring the effects of novel treatments. These studies aimed to develop a fully automated, high-throughput micro-computed tomography (micro-CT)-based image analysis technique for longitudinal quantitation of adipose, non-adipose and lean tissue as well as bone and demonstrate utility for assessing the effects of two distinct treatments. An initial validation study was performed in diet-induced obesity (DIO) and control mice on a vivaCT 75 micro-CT system. Subsequently, four groups of DIO mice were imaged pre- and post-treatment with an experimental agonistic antibody specific for anti-fibroblast growth factor receptor 1 (anti-FGFR1, R1MAb1), control immunoglobulin G antibody, a known anorectic antiobesity drug (rimonabant, SR141716), or solvent control. The body composition analysis technique was then ported to a faster micro-CT system (CT120) to markedly increase throughput as well as to evaluate the use of micro-CT image intensity for hepatic lipid content in DIO and control mice. Ex vivo chemical analysis and colorimetric analysis of the liver triglycerides were performed as the standard metrics for correlation with body composition and hepatic lipid status, respectively. Micro-CT-based body composition measures correlate with ex vivo chemical analysis metrics and enable distinction between DIO and control mice. R1MAb1 and rimonabant have differing effects on body composition as assessed by micro-CT. High-throughput body composition imaging is possible using a modified CT120 system. Micro-CT also provides a non-invasive assessment of hepatic lipid content. This work describes, validates and demonstrates utility of a fully automated image analysis technique to quantify in vivo micro-CT-derived measures of adipose, non-adipose and lean tissue, as well as bone. These body composition metrics highly correlate with standard ex vivo chemical analysis and enable longitudinal evaluation of body composition and therapeutic efficacy monitoring.

Multiscale Porosity and Mechanical Properties of Mancos Shale: Evaluation of REV and Scale Separation

NASA Astrophysics Data System (ADS)

Heath, J. E.; Dewers, T. A.; Yoon, H.; Mozley, P.

2016-12-01

Heterogeneity from the nanometer to core and larger length scales is a major challenge to understanding coupled processes in shale. To develop methods to address this challenge, we present application of high throughput multi-beam scanning electron microscopy (mSEM) and nano-to-micro-scale mechanics to the Mancos Shale. We use a 61-beam mSEM to collect 6 nm resolution SEM images at the scale of several square millimeters. These images are analyzed for pore size and shape characteristics including spatial correlation and structure. Nano-indentation, micropillar compression, and axisymmetric testing at multiple length scales allows for examining the influence of sampling size on mechanical response. The combined data set is used to: investigate representative elementary volumes (and areas for the 2D images) for the Mancos Shale; determine if scale separation occurs; and determine if transport and mechanical properties at a given length scale can be statistically defined. Sandia National Laboratories is a multi-program laboratory managed and operated by Sandia Corporation, a wholly owned subsidiary of Lockheed Martin Corporation, for the U.S. Department of Energy's National Nuclear Security Administration under contract DE-AC04-94AL85000.

Real-Time Aggressive Image Data Compression

DTIC Science & Technology

1990-03-31

implemented with higher degrees of modularity, concurrency, and higher levels of machine intelligence , thereby providing higher data -throughput rates...Project Summary Project Title: Real-Time Aggressive Image Data Compression Principal Investigators: Dr. Yih-Fang Huang and Dr. Ruey-wen Liu Institution...Summary The objective of the proposed research is to develop reliable algorithms !.hat can achieve aggressive image data compression (with a compression

High-throughput live-imaging of embryos in microwell arrays using a modular specimen mounting system.

PubMed

Donoughe, Seth; Kim, Chiyoung; Extavour, Cassandra G

2018-04-30

High-throughput live-imaging of embryos is an essential technique in developmental biology, but it is difficult and costly to mount and image embryos in consistent conditions. Here, we present OMMAwell, a simple, reusable device to easily mount dozens of embryos in arrays of agarose microwells with customizable dimensions and spacing. OMMAwell can be configured to mount specimens for upright or inverted microscopes, and includes a reservoir to hold live-imaging medium to maintain constant moisture and osmolarity of specimens during time-lapse imaging. All device components can be fabricated by cutting pieces from a sheet of acrylic using a laser cutter or by making them with a 3D printer. We demonstrate how to design a custom mold and use it to live-image dozens of embryos at a time. We include descriptions, schematics, and design files for 13 additional molds for nine animal species, including most major traditional laboratory models and a number of emerging model systems. Finally, we provide instructions for researchers to customize OMMAwell inserts for embryos or tissues not described herein. © 2018. Published by The Company of Biologists Ltd.

Prediction-based association control scheme in dense femtocell networks.

PubMed

Sung, Nak Woon; Pham, Ngoc-Thai; Huynh, Thong; Hwang, Won-Joo; You, Ilsun; Choo, Kim-Kwang Raymond

2017-01-01

The deployment of large number of femtocell base stations allows us to extend the coverage and efficiently utilize resources in a low cost manner. However, the small cell size of femtocell networks can result in frequent handovers to the mobile user, and consequently throughput degradation. Thus, in this paper, we propose predictive association control schemes to improve the system's effective throughput. Our design focuses on reducing handover frequency without impacting on throughput. The proposed schemes determine handover decisions that contribute most to the network throughput and are proper for distributed implementations. The simulation results show significant gains compared with existing methods in terms of handover frequency and network throughput perspective.

Development and Validation of an Automated High-Throughput System for Zebrafish In Vivo Screenings

PubMed Central

Virto, Juan M.; Holgado, Olaia; Diez, Maria; Izpisua Belmonte, Juan Carlos; Callol-Massot, Carles

2012-01-01

The zebrafish is a vertebrate model compatible with the paradigms of drug discovery. The small size and transparency of zebrafish embryos make them amenable for the automation necessary in high-throughput screenings. We have developed an automated high-throughput platform for in vivo chemical screenings on zebrafish embryos that includes automated methods for embryo dispensation, compound delivery, incubation, imaging and analysis of the results. At present, two different assays to detect cardiotoxic compounds and angiogenesis inhibitors can be automatically run in the platform, showing the versatility of the system. A validation of these two assays with known positive and negative compounds, as well as a screening for the detection of unknown anti-angiogenic compounds, have been successfully carried out in the system developed. We present a totally automated platform that allows for high-throughput screenings in a vertebrate organism. PMID:22615792

Mixing and transport during pharmaceutical twin-screw wet granulation: experimental analysis via chemical imaging.

PubMed

Kumar, Ashish; Vercruysse, Jurgen; Toiviainen, Maunu; Panouillot, Pierre-Emmanuel; Juuti, Mikko; Vanhoorne, Valérie; Vervaet, Chris; Remon, Jean Paul; Gernaey, Krist V; De Beer, Thomas; Nopens, Ingmar

2014-07-01

Twin-screw granulation is a promising continuous alternative for traditional batch high shear wet granulation (HSWG). The extent of HSWG in a twin screw granulator (TSG) is greatly governed by the residence time of the granulation materials in the TSG and degree of mixing. In order to determine the residence time distribution (RTD) and mixing in TSG, mostly visual observation and particle tracking methods are used, which are either inaccurate and difficult for short RTD, or provide an RTD only for a finite number of preferential tracer paths. In this study, near infrared chemical imaging, which is more accurate and provides a complete RTD, was used. The impact of changes in material throughput (10-17 kg/h), screw speed (500-900 rpm), number of kneading discs (2-12) and stagger angle (30-90°) on the RTD and axial mixing of the material was characterised. The experimental RTD curves were used to calculate the mean residence time, mean centred variance and the Péclet number to determine the axial mixing and predominance of convective over dispersive transport. The results showed that screw speed is the most influential parameter in terms of RTD and axial mixing in the TSG and established a significant interaction between screw design parameters (number and stagger angle of kneading discs) and the process parameters (material throughput and number of kneading discs). The results of the study will allow the development and validation of a transport model capable of predicting the RTD and macro-mixing in the TSG. These can later be coupled with a population balance model in order to predict granulation yields in a TSG more accurately. Copyright © 2014 Elsevier B.V. All rights reserved.

Laser-Induced Fluorescence Detection in High-Throughput Screening of Heterogeneous Catalysts and Single Cells Analysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Su, Hui

2001-01-01

Laser-induced fluorescence detection is one of the most sensitive detection techniques and it has found enormous applications in various areas. The purpose of this research was to develop detection approaches based on laser-induced fluorescence detection in two different areas, heterogeneous catalysts screening and single cell study. First, the author introduced laser-induced imaging (LIFI) as a high-throughput screening technique for heterogeneous catalysts to explore the use of this high-throughput screening technique in discovery and study of various heterogeneous catalyst systems. This scheme is based on the fact that the creation or the destruction of chemical bonds alters the fluorescence properties ofmore » suitably designed molecules. By irradiating the region immediately above the catalytic surface with a laser, the fluorescence intensity of a selected product or reactant can be imaged by a charge-coupled device (CCD) camera to follow the catalytic activity as a function of time and space. By screening the catalytic activity of vanadium pentoxide catalysts in oxidation of naphthalene, they demonstrated LIFI has good detection performance and the spatial and temporal resolution needed for high-throughput screening of heterogeneous catalysts. The sample packing density can reach up to 250 x 250 subunits/cm 2 for 40-μm wells. This experimental set-up also can screen solid catalysts via near infrared thermography detection. In the second part of this dissertation, the author used laser-induced native fluorescence coupled with capillary electrophoresis (LINF-CE) and microscope imaging to study the single cell degranulation. On the basis of good temporal correlation with events observed through an optical microscope, they have identified individual peaks in the fluorescence electropherograms as serotonin released from the granular core on contact with the surrounding fluid.« less

High-throughput analysis of yeast replicative aging using a microfluidic system

PubMed Central

Jo, Myeong Chan; Liu, Wei; Gu, Liang; Dang, Weiwei; Qin, Lidong

2015-01-01

Saccharomyces cerevisiae has been an important model for studying the molecular mechanisms of aging in eukaryotic cells. However, the laborious and low-throughput methods of current yeast replicative lifespan assays limit their usefulness as a broad genetic screening platform for research on aging. We address this limitation by developing an efficient, high-throughput microfluidic single-cell analysis chip in combination with high-resolution time-lapse microscopy. This innovative design enables, to our knowledge for the first time, the determination of the yeast replicative lifespan in a high-throughput manner. Morphological and phenotypical changes during aging can also be monitored automatically with a much higher throughput than previous microfluidic designs. We demonstrate highly efficient trapping and retention of mother cells, determination of the replicative lifespan, and tracking of yeast cells throughout their entire lifespan. Using the high-resolution and large-scale data generated from the high-throughput yeast aging analysis (HYAA) chips, we investigated particular longevity-related changes in cell morphology and characteristics, including critical cell size, terminal morphology, and protein subcellular localization. In addition, because of the significantly improved retention rate of yeast mother cell, the HYAA-Chip was capable of demonstrating replicative lifespan extension by calorie restriction. PMID:26170317

IR Imager Exposure Calculator

Science.gov Websites

CTIO Infrared Imager Exposure Time Calculator Note: ISPI throughput values updated 12 March 2005 S/N ratio 10 Exposure Time 1 (seconds) Calculate S/N for specified Total Integration Time Calculate Total Integration Time to reach Desired S/N Submit Exposure Calculation Request [CTIO Home] [CTIO IR

Rapid identification of salmonella serotypes with stereo and hyperspectral microscope imaging Methods

USDA-ARS?s Scientific Manuscript database

The hyperspectral microscope imaging (HMI) method can reduce detection time within 8 hours including incubation process. The early and rapid detection with this method in conjunction with the high throughput capabilities makes HMI method a prime candidate for implementation for the food industry. Th...

Rapid Identification of Salmonella Serotypes with Stereo and Hyperspectral Microscope Imaging Methods

USDA-ARS?s Scientific Manuscript database

The hyperspectral microscope imaging (HMI) method can reduce detection time within 8 hours including incubation process. The early and rapid detection with this method in conjunction with the high throughput capabilities makes HMI method a prime candidate for implementation for the food industry. Th...

A high-throughput two channel discrete wavelet transform architecture for the JPEG2000 standard

NASA Astrophysics Data System (ADS)

Badakhshannoory, Hossein; Hashemi, Mahmoud R.; Aminlou, Alireza; Fatemi, Omid

2005-07-01

The Discrete Wavelet Transform (DWT) is increasingly recognized in image and video compression standards, as indicated by its use in JPEG2000. The lifting scheme algorithm is an alternative DWT implementation that has a lower computational complexity and reduced resource requirement. In the JPEG2000 standard two lifting scheme based filter banks are introduced: the 5/3 and 9/7. In this paper a high throughput, two channel DWT architecture for both of the JPEG2000 DWT filters is presented. The proposed pipelined architecture has two separate input channels that process the incoming samples simultaneously with minimum memory requirement for each channel. The architecture had been implemented in VHDL and synthesized on a Xilinx Virtex2 XCV1000. The proposed architecture applies DWT on a 2K by 1K image at 33 fps with a 75 MHZ clock frequency. This performance is achieved with 70% less resources than two independent single channel modules. The high throughput and reduced resource requirement has made this architecture the proper choice for real time applications such as Digital Cinema.

Laser-Induced Fluorescence Detection in High-Throughput Screening of Heterogeneous Catalysts and Single Cells Analysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Su, Hui

2001-01-01

Laser-induced fluorescence detection is one of the most sensitive detection techniques and it has found enormous applications in various areas. The purpose of this research was to develop detection approaches based on laser-induced fluorescence detection in two different areas, heterogeneous catalysts screening and single cell study. First, we introduced laser-induced imaging (LIFI) as a high-throughput screening technique for heterogeneous catalysts to explore the use of this high-throughput screening technique in discovery and study of various heterogeneous catalyst systems. This scheme is based on the fact that the creation or the destruction of chemical bonds alters the fluorescence properties of suitablymore » designed molecules. By irradiating the region immediately above the catalytic surface with a laser, the fluorescence intensity of a selected product or reactant can be imaged by a charge-coupled device (CCD) camera to follow the catalytic activity as a function of time and space. By screening the catalytic activity of vanadium pentoxide catalysts in oxidation of naphthalene, we demonstrated LIFI has good detection performance and the spatial and temporal resolution needed for high-throughput screening of heterogeneous catalysts. The sample packing density can reach up to 250 x 250 subunits/cm 2 for 40-μm wells. This experimental set-up also can screen solid catalysts via near infrared thermography detection.« less

High-resolution, high-throughput imaging with a multibeam scanning electron microscope.

PubMed

Eberle, A L; Mikula, S; Schalek, R; Lichtman, J; Knothe Tate, M L; Zeidler, D

2015-08-01

Electron-electron interactions and detector bandwidth limit the maximal imaging speed of single-beam scanning electron microscopes. We use multiple electron beams in a single column and detect secondary electrons in parallel to increase the imaging speed by close to two orders of magnitude and demonstrate imaging for a variety of samples ranging from biological brain tissue to semiconductor wafers. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

High-throughput screening of high Monascus pigment-producing strain based on digital image processing.

PubMed

Xia, Meng-lei; Wang, Lan; Yang, Zhi-xia; Chen, Hong-zhang

2016-04-01

This work proposed a new method which applied image processing and support vector machine (SVM) for screening of mold strains. Taking Monascus as example, morphological characteristics of Monascus colony were quantified by image processing. And the association between the characteristics and pigment production capability was determined by SVM. On this basis, a highly automated screening strategy was achieved. The accuracy of the proposed strategy is 80.6 %, which is compatible with the existing methods (81.1 % for microplate and 85.4 % for flask). Meanwhile, the screening of 500 colonies only takes 20-30 min, which is the highest rate among all published results. By applying this automated method, 13 strains with high-predicted production were obtained and the best one produced as 2.8-fold (226 U/mL) of pigment and 1.9-fold (51 mg/L) of lovastatin compared with the parent strain. The current study provides us with an effective and promising method for strain improvement.

Industrial-scale separation of high-purity single-chirality single-wall carbon nanotubes for biological imaging

PubMed Central

Yomogida, Yohei; Tanaka, Takeshi; Zhang, Minfang; Yudasaka, Masako; Wei, Xiaojun; Kataura, Hiromichi

2016-01-01

Single-chirality, single-wall carbon nanotubes are desired due to their inherent physical properties and performance characteristics. Here, we demonstrate a chromatographic separation method based on a newly discovered chirality-selective affinity between carbon nanotubes and a gel containing a mixture of the surfactants. In this system, two different selectivities are found: chiral-angle selectivity and diameter selectivity. Since the chirality of nanotubes is determined by the chiral angle and diameter, combining these independent selectivities leads to high-resolution single-chirality separation with milligram-scale throughput and high purity. Furthermore, we present efficient vascular imaging of mice using separated single-chirality (9,4) nanotubes. Due to efficient absorption and emission, blood vessels can be recognized even with the use of ∼100-fold lower injected dose than the reported value for pristine nanotubes. Thus, 1 day of separation provides material for up to 15,000 imaging experiments, which is acceptable for industrial use. PMID:27350127

REgolith X-Ray Imaging Spectrometer (REXIS) Aboard NASA’s OSIRIS-REx Mission

NASA Astrophysics Data System (ADS)

Hong, JaeSub; Allen, Branden; Grindlay, Jonathan E.; Binzel, Richard P.; Masterson, Rebecca; Inamdar, Niraj K; Chodas, Mark; Smith, Matthew W; Bautz, Mark W.; Kissel, Steven E; Villasenor, Jesus Noel; Oprescu, Antonia

2014-06-01

The REgolith X-Ray Imaging Spectrometer (REXIS) is a student-led instrument being designed, built, and operated as a collaborative effort involving MIT and Harvard. It is a part of NASA's OSIRIS-REx mission, which is scheduled for launch in September of 2016 for a rendezvous with, and collection of a sample from the surface of the primitive carbonaceous chondrite-like asteroid 101955 Bennu in 2019. REXIS will determine spatial variations in elemental composition of Bennu's surface through solar-induced X-ray fluorescence. REXIS consists of four X-ray CCDs in the detector plane and an X-ray mask. It is the first coded-aperture X-ray telescope in a planetary mission, which combines the benefit of high X-ray throughput of wide-field collimation with imaging capability of a coded-mask, enabling detection of elemental surface distributions at approximately 50-200 m scales. We present an overview of the REXIS instrument and the expected performance.

Indexing amyloid peptide diffraction from serial femtosecond crystallography: New algorithms for sparse patterns

DOE PAGES

Brewster, Aaron S.; Sawaya, Michael R.; Rodriguez, Jose; ...

2015-01-23

Still diffraction patterns from peptide nanocrystals with small unit cells are challenging to index using conventional methods owing to the limited number of spots and the lack of crystal orientation information for individual images. New indexing algorithms have been developed as part of the Computational Crystallography Toolbox( cctbx) to overcome these challenges. Accurate unit-cell information derived from an aggregate data set from thousands of diffraction patterns can be used to determine a crystal orientation matrix for individual images with as few as five reflections. These algorithms are potentially applicable not only to amyloid peptides but also to any set ofmore » diffraction patterns with sparse properties, such as low-resolution virus structures or high-throughput screening of still images captured by raster-scanning at synchrotron sources. As a proof of concept for this technique, successful integration of X-ray free-electron laser (XFEL) data to 2.5 Å resolution for the amyloid segment GNNQQNY from the Sup35 yeast prion is presented.« less

MODIS Aqua Optical Throughput Degradation Impact on Relative Spectral Response and Calibration on Ocean Color Products

NASA Technical Reports Server (NTRS)

Lee, Shihyan; Meister, Gerhard

2017-01-01

Since Moderate Resolution Imaging Spectroradiometer Aqua's launch in 2002, the radiometric system gains of the reflective solar bands have been degrading, indicating changes in the systems optical throughput. To estimate the optical throughput degradation, the electronic gain changes were estimated and removed from the measured system gain. The derived optical throughput degradation shows a rate that is much faster in the shorter wavelengths than the longer wavelengths. The wavelength-dependent optical throughput degradation modulated the relative spectral response (RSR) of the bands. In addition, the optical degradation is also scan angle-dependent due to large changes in response versus the scan angle over time. We estimated the modulated RSR as a function of time and scan angles and its impacts on sensor radiometric calibration for the ocean science. Our results show that the calibration bias could be up to 1.8 % for band 8 (412 nm) due to its larger out-of-band response. For the other ocean bands, the calibration biases are much smaller with magnitudes at least one order smaller.

The Impact of the Condenser on Cytogenetic Image Quality in Digital Microscope System

PubMed Central

Ren, Liqiang; Li, Zheng; Li, Yuhua; Zheng, Bin; Li, Shibo; Chen, Xiaodong; Liu, Hong

2013-01-01

Background: Optimizing operational parameters of the digital microscope system is an important technique to acquire high quality cytogenetic images and facilitate the process of karyotyping so that the efficiency and accuracy of diagnosis can be improved. OBJECTIVE: This study investigated the impact of the condenser on cytogenetic image quality and system working performance using a prototype digital microscope image scanning system. Methods: Both theoretical analysis and experimental validations through objectively evaluating a resolution test chart and subjectively observing large numbers of specimen were conducted. Results: The results show that the optimal image quality and large depth of field (DOF) are simultaneously obtained when the numerical aperture of condenser is set as 60%–70% of the corresponding objective. Under this condition, more analyzable chromosomes and diagnostic information are obtained. As a result, the system shows higher working stability and less restriction for the implementation of algorithms such as autofocusing especially when the system is designed to achieve high throughput continuous image scanning. Conclusions: Although the above quantitative results were obtained using a specific prototype system under the experimental conditions reported in this paper, the presented evaluation methodologies can provide valuable guidelines for optimizing operational parameters in cytogenetic imaging using the high throughput continuous scanning microscopes in clinical practice. PMID:23676284

Lens-free computational imaging of capillary morphogenesis within three-dimensional substrates

NASA Astrophysics Data System (ADS)

Weidling, John; Isikman, Serhan O.; Greenbaum, Alon; Ozcan, Aydogan; Botvinick, Elliot

2012-12-01

Endothelial cells cultured in three-dimensional (3-D) extracellular matrices spontaneously form microvessels in response to soluble and matrix-bound factors. Such cultures are common for the study of angiogenesis and may find widespread use in drug discovery. Vascular networks are imaged over weeks to measure the distribution of vessel morphogenic parameters. Measurements require micron-scale spatial resolution, which for light microscopy comes at the cost of limited field-of-view (FOV) and shallow depth-of-focus (DOF). Small FOVs and DOFs necessitate lateral and axial mechanical scanning, thus limiting imaging throughput. We present a lens-free holographic on-chip microscopy technique to rapidly image microvessels within a Petri dish over a large volume without any mechanical scanning. This on-chip method uses partially coherent illumination and a CMOS sensor to record in-line holographic images of the sample. For digital reconstruction of the measured holograms, we implement a multiheight phase recovery method to obtain phase images of capillary morphogenesis over a large FOV (24 mm2) with ˜1.5 μm spatial resolution. On average, measured capillary length in our method was within approximately 2% of lengths measured using a 10× microscope objective. These results suggest lens-free on-chip imaging is a useful toolset for high-throughput monitoring and quantitative analysis of microvascular 3-D networks.

iScreen: Image-Based High-Content RNAi Screening Analysis Tools.

PubMed

Zhong, Rui; Dong, Xiaonan; Levine, Beth; Xie, Yang; Xiao, Guanghua

2015-09-01

High-throughput RNA interference (RNAi) screening has opened up a path to investigating functional genomics in a genome-wide pattern. However, such studies are often restricted to assays that have a single readout format. Recently, advanced image technologies have been coupled with high-throughput RNAi screening to develop high-content screening, in which one or more cell image(s), instead of a single readout, were generated from each well. This image-based high-content screening technology has led to genome-wide functional annotation in a wider spectrum of biological research studies, as well as in drug and target discovery, so that complex cellular phenotypes can be measured in a multiparametric format. Despite these advances, data analysis and visualization tools are still largely lacking for these types of experiments. Therefore, we developed iScreen (image-Based High-content RNAi Screening Analysis Tool), an R package for the statistical modeling and visualization of image-based high-content RNAi screening. Two case studies were used to demonstrate the capability and efficiency of the iScreen package. iScreen is available for download on CRAN (http://cran.cnr.berkeley.edu/web/packages/iScreen/index.html). The user manual is also available as a supplementary document. © 2014 Society for Laboratory Automation and Screening.

Automated processing of zebrafish imaging data: a survey.

PubMed

Mikut, Ralf; Dickmeis, Thomas; Driever, Wolfgang; Geurts, Pierre; Hamprecht, Fred A; Kausler, Bernhard X; Ledesma-Carbayo, María J; Marée, Raphaël; Mikula, Karol; Pantazis, Periklis; Ronneberger, Olaf; Santos, Andres; Stotzka, Rainer; Strähle, Uwe; Peyriéras, Nadine

2013-09-01

Due to the relative transparency of its embryos and larvae, the zebrafish is an ideal model organism for bioimaging approaches in vertebrates. Novel microscope technologies allow the imaging of developmental processes in unprecedented detail, and they enable the use of complex image-based read-outs for high-throughput/high-content screening. Such applications can easily generate Terabytes of image data, the handling and analysis of which becomes a major bottleneck in extracting the targeted information. Here, we describe the current state of the art in computational image analysis in the zebrafish system. We discuss the challenges encountered when handling high-content image data, especially with regard to data quality, annotation, and storage. We survey methods for preprocessing image data for further analysis, and describe selected examples of automated image analysis, including the tracking of cells during embryogenesis, heartbeat detection, identification of dead embryos, recognition of tissues and anatomical landmarks, and quantification of behavioral patterns of adult fish. We review recent examples for applications using such methods, such as the comprehensive analysis of cell lineages during early development, the generation of a three-dimensional brain atlas of zebrafish larvae, and high-throughput drug screens based on movement patterns. Finally, we identify future challenges for the zebrafish image analysis community, notably those concerning the compatibility of algorithms and data formats for the assembly of modular analysis pipelines.

Automated Processing of Zebrafish Imaging Data: A Survey

PubMed Central

Dickmeis, Thomas; Driever, Wolfgang; Geurts, Pierre; Hamprecht, Fred A.; Kausler, Bernhard X.; Ledesma-Carbayo, María J.; Marée, Raphaël; Mikula, Karol; Pantazis, Periklis; Ronneberger, Olaf; Santos, Andres; Stotzka, Rainer; Strähle, Uwe; Peyriéras, Nadine

2013-01-01

Abstract Due to the relative transparency of its embryos and larvae, the zebrafish is an ideal model organism for bioimaging approaches in vertebrates. Novel microscope technologies allow the imaging of developmental processes in unprecedented detail, and they enable the use of complex image-based read-outs for high-throughput/high-content screening. Such applications can easily generate Terabytes of image data, the handling and analysis of which becomes a major bottleneck in extracting the targeted information. Here, we describe the current state of the art in computational image analysis in the zebrafish system. We discuss the challenges encountered when handling high-content image data, especially with regard to data quality, annotation, and storage. We survey methods for preprocessing image data for further analysis, and describe selected examples of automated image analysis, including the tracking of cells during embryogenesis, heartbeat detection, identification of dead embryos, recognition of tissues and anatomical landmarks, and quantification of behavioral patterns of adult fish. We review recent examples for applications using such methods, such as the comprehensive analysis of cell lineages during early development, the generation of a three-dimensional brain atlas of zebrafish larvae, and high-throughput drug screens based on movement patterns. Finally, we identify future challenges for the zebrafish image analysis community, notably those concerning the compatibility of algorithms and data formats for the assembly of modular analysis pipelines. PMID:23758125

Ruby-Helix: an implementation of helical image processing based on object-oriented scripting language.

PubMed

Metlagel, Zoltan; Kikkawa, Yayoi S; Kikkawa, Masahide

2007-01-01

Helical image analysis in combination with electron microscopy has been used to study three-dimensional structures of various biological filaments or tubes, such as microtubules, actin filaments, and bacterial flagella. A number of packages have been developed to carry out helical image analysis. Some biological specimens, however, have a symmetry break (seam) in their three-dimensional structure, even though their subunits are mostly arranged in a helical manner. We refer to these objects as "asymmetric helices". All the existing packages are designed for helically symmetric specimens, and do not allow analysis of asymmetric helical objects, such as microtubules with seams. Here, we describe Ruby-Helix, a new set of programs for the analysis of "helical" objects with or without a seam. Ruby-Helix is built on top of the Ruby programming language and is the first implementation of asymmetric helical reconstruction for practical image analysis. It also allows easier and semi-automated analysis, performing iterative unbending and accurate determination of the repeat length. As a result, Ruby-Helix enables us to analyze motor-microtubule complexes with higher throughput to higher resolution.

High throughput analysis of samples in flowing liquid

DOEpatents

Ambrose, W. Patrick; Grace, W. Kevin; Goodwin, Peter M.; Jett, James H.; Orden, Alan Van; Keller, Richard A.

2001-01-01

Apparatus and method enable imaging multiple fluorescent sample particles in a single flow channel. A flow channel defines a flow direction for samples in a flow stream and has a viewing plane perpendicular to the flow direction. A laser beam is formed as a ribbon having a width effective to cover the viewing plane. Imaging optics are arranged to view the viewing plane to form an image of the fluorescent sample particles in the flow stream, and a camera records the image formed by the imaging optics.

Coherent imaging at the diffraction limit

PubMed Central

Thibault, Pierre; Guizar-Sicairos, Manuel; Menzel, Andreas

2014-01-01

X-ray ptychography, a scanning coherent diffractive imaging technique, holds promise for imaging with dose-limited resolution and sensitivity. If the foreseen increase of coherent flux by orders of magnitude can be matched by additional technological and analytical advances, ptychography may approach imaging speeds familiar from full-field methods while retaining its inherently quantitative nature and metrological versatility. Beyond promises of high throughput, spectroscopic applications in three dimensions become feasible, as do measurements of sample dynamics through time-resolved imaging or careful characterization of decoherence effects. PMID:25177990

Coherent imaging at the diffraction limit.

PubMed

Thibault, Pierre; Guizar-Sicairos, Manuel; Menzel, Andreas

2014-09-01

X-ray ptychography, a scanning coherent diffractive imaging technique, holds promise for imaging with dose-limited resolution and sensitivity. If the foreseen increase of coherent flux by orders of magnitude can be matched by additional technological and analytical advances, ptychography may approach imaging speeds familiar from full-field methods while retaining its inherently quantitative nature and metrological versatility. Beyond promises of high throughput, spectroscopic applications in three dimensions become feasible, as do measurements of sample dynamics through time-resolved imaging or careful characterization of decoherence effects.

Prediction-based association control scheme in dense femtocell networks

PubMed Central

Pham, Ngoc-Thai; Huynh, Thong; Hwang, Won-Joo; You, Ilsun; Choo, Kim-Kwang Raymond

2017-01-01

The deployment of large number of femtocell base stations allows us to extend the coverage and efficiently utilize resources in a low cost manner. However, the small cell size of femtocell networks can result in frequent handovers to the mobile user, and consequently throughput degradation. Thus, in this paper, we propose predictive association control schemes to improve the system’s effective throughput. Our design focuses on reducing handover frequency without impacting on throughput. The proposed schemes determine handover decisions that contribute most to the network throughput and are proper for distributed implementations. The simulation results show significant gains compared with existing methods in terms of handover frequency and network throughput perspective. PMID:28328992

On-chip ultraviolet holography for high-throughput nanoparticle and biomolecule detection

NASA Astrophysics Data System (ADS)

Daloglu, Mustafa Ugur; Ray, Aniruddha; Gorocs, Zoltán.; Xiong, Matthew; Malik, Ravinder; Bitan, Gal; McLeod, Euan; Ozcan, Aydogan

2018-02-01

Nanoparticle and biomolecule imaging has become an important need for various applications. In an effort to find a higher throughput alternative to existing devices, we have designed a lensfree on-chip holographic imaging platform operating at an ultraviolet (UV) wavelength of 266 nm. With a custom-designed free-space light delivery system to illuminate the sample that is placed very close (<0.5 mm) to an opto-electronic image sensor chip, without any imaging lenses in between, the full active area of the imager chip (>16 mm2 ) was utilized as the imaging field-of-view (FOV) capturing holographic signatures of target objects on a chip. These holograms were then digitally back propagated to extract both the amplitude and phase information of the sample. The increased forward scattering from nanoparticles due to this shorter illumination wavelength has enabled us to image individual particles that are smaller than 30 nm over an FOV of >16 mm2 . Our platform was further utilized in high-contrast imaging of nanoscopic biomolecule aggregates since 266 nm illumination light is strongly absorbed by biomolecules including proteins and nucleic acids. Aggregates of Cu/Zn-superoxide dismutase (SOD1), which has been linked to a fatal neurodegenerative disease, ALS (amyotrophic lateral sclerosis), have been imaged with significantly improved contrast compared to imaging at visible wavelengths. This unique UV imaging modality could be valuable for biomedical applications (e.g., viral load measurements) and environmental monitoring including air and water quality monitoring.

High-throughput analysis of sub-visible mAb aggregate particles using automated fluorescence microscopy imaging.

PubMed

Paul, Albert Jesuran; Bickel, Fabian; Röhm, Martina; Hospach, Lisa; Halder, Bettina; Rettich, Nina; Handrick, René; Herold, Eva Maria; Kiefer, Hans; Hesse, Friedemann

2017-07-01

Aggregation of therapeutic proteins is a major concern as aggregates lower the yield and can impact the efficacy of the drug as well as the patient's safety. It can occur in all production stages; thus, it is essential to perform a detailed analysis for protein aggregates. Several methods such as size exclusion high-performance liquid chromatography (SE-HPLC), light scattering, turbidity, light obscuration, and microscopy-based approaches are used to analyze aggregates. None of these methods allows determination of all types of higher molecular weight (HMW) species due to a limited size range. Furthermore, quantification and specification of different HMW species are often not possible. Moreover, automation is a perspective challenge coming up with automated robotic laboratory systems. Hence, there is a need for a fast, high-throughput-compatible method, which can detect a broad size range and enable quantification and classification. We describe a novel approach for the detection of aggregates in the size range 1 to 1000 μm combining fluorescent dyes for protein aggregate labelling and automated fluorescence microscope imaging (aFMI). After appropriate selection of the dye and method optimization, our method enabled us to detect various types of HMW species of monoclonal antibodies (mAbs). Using 10 μmol L -1 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonate (Bis-ANS) in combination with aFMI allowed the analysis of mAb aggregates induced by different stresses occurring during downstream processing, storage, and administration. Validation of our results was performed by SE-HPLC, UV-Vis spectroscopy, and dynamic light scattering. With this new approach, we could not only reliably detect different HMW species but also quantify and classify them in an automated approach. Our method achieves high-throughput requirements and the selection of various fluorescent dyes enables a broad range of applications.

Uninterrupted monitoring of drug effects in human-induced pluripotent stem cell-derived cardiomyocytes with bioluminescence Ca2+ microscopy.

PubMed

Suzuki, Kazushi; Onishi, Takahito; Nakada, Chieko; Takei, Shunsuke; Daniels, Matthew J; Nakano, Masahiro; Matsuda, Tomoki; Nagai, Takeharu

2018-05-18

Cardiomyocytes derived from human-induced pluripotent stem cells are a powerful platform for high-throughput drug screening in vitro. However, current modalities for drug testing, such as electrophysiology and fluorescence imaging have inherent drawbacks. To circumvent these problems, we report the development of a bioluminescent Ca 2+ indicator GmNL(Ca 2+ ), and its application in a customized microscope for high-throughput drug screening. GmNL(Ca 2+ ) gives a 140% signal change with Ca 2+ , and can image drug-induced changes of Ca 2+ dynamics in cultured cells. Since bioluminescence requires application of a chemical substrate, which is consumed over ~ 30 min we made a dedicated microscope with automated drug dispensing inside a light-tight box, to control drug addition. To overcome thermal instability of the luminescent substrate, or small molecule, dual climate control enables distinct temperature settings in the drug reservoir and the biological sample. By combining GmNL(Ca 2+ ) with this adaptation, we could image spontaneous Ca 2+ transients in cultured cardiomyocytes and phenotype their response to well-known drugs without accessing the sample directly. In addition, the bioluminescent strategy demonstrates minimal perturbation of contractile parameters and long-term observation attributable to lack of phototoxicity and photobleaching. Overall, bioluminescence may enable more accurate drug screening in a high-throughput manner.

Vortex coronagraphs for the Habitable Exoplanet Imaging Mission concept: theoretical performance and telescope requirements

NASA Astrophysics Data System (ADS)

Ruane, Garreth; Mawet, Dimitri; Mennesson, Bertrand; Jewell, Jeffrey; Shaklan, Stuart

2018-01-01

The Habitable Exoplanet Imaging Mission concept requires an optical coronagraph that provides deep starlight suppression over a broad spectral bandwidth, high throughput for point sources at small angular separation, and insensitivity to temporally varying, low-order aberrations. Vortex coronagraphs are a promising solution that performs optimally on off-axis, monolithic telescopes and may also be designed for segmented telescopes with minor losses in performance. We describe the key advantages of vortex coronagraphs on off-axis telescopes such as (1) unwanted diffraction due to aberrations is passively rejected in several low-order Zernike modes relaxing the wavefront stability requirements for imaging Earth-like planets from <10 to >100 pm rms, (2) stars with angular diameters >0.1 λ / D may be sufficiently suppressed, (3) the absolute planet throughput is >10 % , even for unfavorable telescope architectures, and (4) broadband solutions (Δλ / λ > 0.1) are readily available for both monolithic and segmented apertures. The latter make use of grayscale apodizers in an upstream pupil plane to provide suppression of diffracted light from amplitude discontinuities in the telescope pupil without inducing additional stroke on the deformable mirrors. We set wavefront stability requirements on the telescope, based on a stellar irradiance threshold set at an angular separation of 3 ± 0.5λ / D from the star, and discuss how some requirements may be relaxed by trading robustness to aberrations for planet throughput.

A High-Throughput Automated Microfluidic Platform for Calcium Imaging of Taste Sensing.

PubMed

Hsiao, Yi-Hsing; Hsu, Chia-Hsien; Chen, Chihchen

2016-07-08

The human enteroendocrine L cell line NCI-H716, expressing taste receptors and taste signaling elements, constitutes a unique model for the studies of cellular responses to glucose, appetite regulation, gastrointestinal motility, and insulin secretion. Targeting these gut taste receptors may provide novel treatments for diabetes and obesity. However, NCI-H716 cells are cultured in suspension and tend to form multicellular aggregates, preventing high-throughput calcium imaging due to interferences caused by laborious immobilization and stimulus delivery procedures. Here, we have developed an automated microfluidic platform that is capable of trapping more than 500 single cells into microwells with a loading efficiency of 77% within two minutes, delivering multiple chemical stimuli and performing calcium imaging with enhanced spatial and temporal resolutions when compared to bath perfusion systems. Results revealed the presence of heterogeneity in cellular responses to the type, concentration, and order of applied sweet and bitter stimuli. Sucralose and denatonium benzoate elicited robust increases in the intracellular Ca(2+) concentration. However, glucose evoked a rapid elevation of intracellular Ca(2+) followed by reduced responses to subsequent glucose stimulation. Using Gymnema sylvestre as a blocking agent for the sweet taste receptor confirmed that different taste receptors were utilized for sweet and bitter tastes. This automated microfluidic platform is cost-effective, easy to fabricate and operate, and may be generally applicable for high-throughput and high-content single-cell analysis and drug screening.

Data for automated, high-throughput microscopy analysis of intracellular bacterial colonies using spot detection.

PubMed

Ernstsen, Christina L; Login, Frédéric H; Jensen, Helene H; Nørregaard, Rikke; Møller-Jensen, Jakob; Nejsum, Lene N

2017-10-01

Quantification of intracellular bacterial colonies is useful in strategies directed against bacterial attachment, subsequent cellular invasion and intracellular proliferation. An automated, high-throughput microscopy-method was established to quantify the number and size of intracellular bacterial colonies in infected host cells (Detection and quantification of intracellular bacterial colonies by automated, high-throughput microscopy, Ernstsen et al., 2017 [1]). The infected cells were imaged with a 10× objective and number of intracellular bacterial colonies, their size distribution and the number of cell nuclei were automatically quantified using a spot detection-tool. The spot detection-output was exported to Excel, where data analysis was performed. In this article, micrographs and spot detection data are made available to facilitate implementation of the method.

IFDOTMETER: A New Software Application for Automated Immunofluorescence Analysis.

PubMed

Rodríguez-Arribas, Mario; Pizarro-Estrella, Elisa; Gómez-Sánchez, Rubén; Yakhine-Diop, S M S; Gragera-Hidalgo, Antonio; Cristo, Alejandro; Bravo-San Pedro, Jose M; González-Polo, Rosa A; Fuentes, José M

2016-04-01

Most laboratories interested in autophagy use different imaging software for managing and analyzing heterogeneous parameters in immunofluorescence experiments (e.g., LC3-puncta quantification and determination of the number and size of lysosomes). One solution would be software that works on a user's laptop or workstation that can access all image settings and provide quick and easy-to-use analysis of data. Thus, we have designed and implemented an application called IFDOTMETER, which can run on all major operating systems because it has been programmed using JAVA (Sun Microsystems). Briefly, IFDOTMETER software has been created to quantify a variety of biological hallmarks, including mitochondrial morphology and nuclear condensation. The program interface is intuitive and user-friendly, making it useful for users not familiar with computer handling. By setting previously defined parameters, the software can automatically analyze a large number of images without the supervision of the researcher. Once analysis is complete, the results are stored in a spreadsheet. Using software for high-throughput cell image analysis offers researchers the possibility of performing comprehensive and precise analysis of a high number of images in an automated manner, making this routine task easier. © 2015 Society for Laboratory Automation and Screening.

Quantitative Image Informatics for Cancer Research (QIICR) | Informatics Technology for Cancer Research (ITCR)

Cancer.gov

Imaging has enormous untapped potential to improve cancer research through software to extract and process morphometric and functional biomarkers. In the era of non-cytotoxic treatment agents, multi- modality image-guided ablative therapies and rapidly evolving computational resources, quantitative imaging software can be transformative in enabling minimally invasive, objective and reproducible evaluation of cancer treatment response. Post-processing algorithms are integral to high-throughput analysis and fine- grained differentiation of multiple molecular targets.

High throughput phenotyping of tomato spotted wilt disease in peanuts using unmanned aerial systems and multispectral imaging

USDA-ARS?s Scientific Manuscript database

The amount of visible and near infrared light reflected by plants varies depending on their health. In this study, multispectral images were acquired by quadcopter for detecting tomato spot wilt virus amongst twenty genetic varieties of peanuts. The plants were visually assessed to acquire ground ...

Using high-content imaging data from ToxCast to analyze toxicological tipping points (TDS)

EPA Science Inventory

Translating results obtained from high-throughput screening to risk assessment is vital for reducing dependence on animal testing. We studied the effects of 976 chemicals (ToxCast Phase I and II) in HepG2 cells using high-content imaging (HCI) to measure dose and time-depende...

Raman microspectrometer combined with scattering microscopy and lensless imaging for bacteria identification

NASA Astrophysics Data System (ADS)

Strola, S. A.; Schultz, E.; Allier, C. P.; DesRoches, B.; Lemmonier, J.; Dinten, J.-M.

2013-03-01

In this paper, we report on a compact prototype capable both of lensfree imaging, Raman spectrometry and scattering microscopy from bacteria samples. This instrument allows high-throughput real-time characterization without the need of markers, making it potentially suitable to field label-free biomedical and environmental applications. Samples are illuminated from above with a focused-collimated 532nm laser beam and can be x-y-z scanned. The bacteria detection is based on emerging lensfree imaging technology able to localize cells of interest over a large field-of-view of 24mm2. Raman signal and scattered light are then collected by separate measurement arms simultaneously. In the first arm the emission light is fed by a fiber into a prototype spectrometer, developed by Tornado Spectral System based on Tornado's High Throughput Virtual Slit (HTVS) novel technology. The enhanced light throughput in the spectral region of interest (500-1800 cm-1) reduces Raman acquisition time down to few seconds, thus facilitating experimental protocols and avoiding the bacteria deterioration induced by laser thermal heating. Scattered light impinging in the second arm is collected onto a charge-coupled-device. The reconstructed image allows studying the single bacteria diffraction pattern and their specific structural features. The characterization and identification of different bacteria have been performed to validate and optimize the acquisition system and the component setup. The results obtained demonstrate the benefits of these three techniques combination by providing the precise bacteria localization, their chemical composition and a morphology description. The procedure for a rapid identification of particular pathogen bacteria in a sample is illustrated.

Lessons From Paired Data From exPVP Maize Lines in Agronomic Field Trials and RGB And Hyperspectral Time-Series Imaging In Controlled Environments

NASA Astrophysics Data System (ADS)

Schnable, J. C.; Pandey, P.; Ge, Y.; Xu, Y.; Qiu, Y.; Liang, Z.

2017-12-01

Maize Zea mays ssp. mays is one of three crops, along with rice and wheat, responsible for more than 1/2 of all calories consumed around the world. Increasing the yield and stress tolerance of these crops is essential to meet the growing need for food. The cost and speed of plant phenotyping is currently the largest constraint on plant breeding efforts. Datasets linking new types of high throughput phenotyping data collected from plants to the performance of the same genotypes under agronomic conditions across a wide range of environments are essential for developing new statistical approaches and computer vision based tools. A set of maize inbreds and hybrids - primarily recently off patent lines - were phenotyped using a high throughput platform at University of Nebraska-Lincoln. These lines have been previously subjected to high density genotyping, and scored for a core set of 13 phenotypes in field trials across 13 North American states in 2014, 2015, 2016, and 2017. Correlations between image-based measurements and manual measurements demonstrated the feasibility of quantifying variation in plant architecture using image data. However, we demonstrate that naive approaches to measuring traits such as biomass where are developed without integrating genotypic information can introduce nonrandom measurement errors which are confounded with variation between plant accessions. Analysis of hyperspectral image data demonstrated unique signatures from stem tissue which were not identified using aerial imagry. Integrating heritable phenotypes from high-throughput phenotyping data with field data from different environments can reveal previously unknown factors influencing yield plasticity.

High-throughput multiple-mouse imaging with micro-PET/CT for whole-skeleton assessment.

PubMed

Yagi, Masashi; Arentsen, Luke; Shanley, Ryan M; Hui, Susanta K

2014-11-01

Recent studies have proven that skeleton-wide functional assessment is essential to comprehensively understand physiological aspects of the skeletal system. Therefore, in contrast to regional imaging studies utilizing a multiple-animal holder (mouse hotel), we attempted to develop and characterize a multiple-mouse imaging system with micro-PET/CT for high-throughput whole-skeleton assessment. Using items found in a laboratory, a simple mouse hotel that houses four mice linked with gas anesthesia was constructed. A mouse-simulating phantom was used to measure uniformity in a cross sectional area and flatness (Amax/Amin*100) along the axial, radial and tangential directions, where Amax and Amin are maximum and minimum activity concentration in the profile, respectively. Fourteen mice were used for single- or multiple-micro-PET/CT scans. NaF uptake was measured at eight skeletal sites (skull to tibia). Skeletal (18)F activities measured with mice in the mouse hotel were within 1.6 ± 4% (mean ± standard deviation) of those measured with mice in the single-mouse holder. Single-holder scanning yields slightly better uniformity and flatness over the hotel. Compared to use of the single-mouse holder, scanning with the mouse hotel reduced study time (by 65%), decreased the number of scans (four-fold), reduced cost, required less computer storage space (40%), and maximized (18)F usage. The mouse hotel allows high-throughput, quantitatively equivalent scanning compared to the single-mouse holder for micro-PET/CT imaging for whole-skeleton assessment of mice. Copyright © 2014 Associazione Italiana di Fisica Medica. Published by Elsevier Ltd. All rights reserved.

FBI Fingerprint Image Capture System High-Speed-Front-End throughput modeling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rathke, P.M.

1993-09-01

The Federal Bureau of Investigation (FBI) has undertaken a major modernization effort called the Integrated Automated Fingerprint Identification System (IAFISS). This system will provide centralized identification services using automated fingerprint, subject descriptor, mugshot, and document processing. A high-speed Fingerprint Image Capture System (FICS) is under development as part of the IAFIS program. The FICS will capture digital and microfilm images of FBI fingerprint cards for input into a central database. One FICS design supports two front-end scanning subsystems, known as the High-Speed-Front-End (HSFE) and Low-Speed-Front-End, to supply image data to a common data processing subsystem. The production rate of themore » HSFE is critical to meeting the FBI`s fingerprint card processing schedule. A model of the HSFE has been developed to help identify the issues driving the production rate, assist in the development of component specifications, and guide the evolution of an operations plan. A description of the model development is given, the assumptions are presented, and some HSFE throughput analysis is performed.« less

Advances in single-cell experimental design made possible by automated imaging platforms with feedback through segmentation.

PubMed

Crick, Alex J; Cammarota, Eugenia; Moulang, Katie; Kotar, Jurij; Cicuta, Pietro

2015-01-01

Live optical microscopy has become an essential tool for studying the dynamical behaviors and variability of single cells, and cell-cell interactions. However, experiments and data analysis in this area are often extremely labor intensive, and it has often not been achievable or practical to perform properly standardized experiments on a statistically viable scale. We have addressed this challenge by developing automated live imaging platforms, to help standardize experiments, increasing throughput, and unlocking previously impossible ones. Our real-time cell tracking programs communicate in feedback with microscope and camera control software, and they are highly customizable, flexible, and efficient. As examples of our current research which utilize these automated platforms, we describe two quite different applications: egress-invasion interactions of malaria parasites and red blood cells, and imaging of immune cells which possess high motility and internal dynamics. The automated imaging platforms are able to track a large number of motile cells simultaneously, over hours or even days at a time, greatly increasing data throughput and opening up new experimental possibilities. Copyright © 2015 Elsevier Inc. All rights reserved.

Intellicount: High-Throughput Quantification of Fluorescent Synaptic Protein Puncta by Machine Learning

PubMed Central

Fantuzzo, J. A.; Mirabella, V. R.; Zahn, J. D.

2017-01-01

Abstract Synapse formation analyses can be performed by imaging and quantifying fluorescent signals of synaptic markers. Traditionally, these analyses are done using simple or multiple thresholding and segmentation approaches or by labor-intensive manual analysis by a human observer. Here, we describe Intellicount, a high-throughput, fully-automated synapse quantification program which applies a novel machine learning (ML)-based image processing algorithm to systematically improve region of interest (ROI) identification over simple thresholding techniques. Through processing large datasets from both human and mouse neurons, we demonstrate that this approach allows image processing to proceed independently of carefully set thresholds, thus reducing the need for human intervention. As a result, this method can efficiently and accurately process large image datasets with minimal interaction by the experimenter, making it less prone to bias and less liable to human error. Furthermore, Intellicount is integrated into an intuitive graphical user interface (GUI) that provides a set of valuable features, including automated and multifunctional figure generation, routine statistical analyses, and the ability to run full datasets through nested folders, greatly expediting the data analysis process. PMID:29218324

Transfer, Imaging, and Analysis Plate for Facile Handling of 384 Hanging Drop 3D Tissue Spheroids

PubMed Central

Cavnar, Stephen P.; Salomonsson, Emma; Luker, Kathryn E.; Luker, Gary D.; Takayama, Shuichi

2014-01-01

Three-dimensional culture systems bridge the experimental gap between in vivo and in vitro physiology. However, nonstandardized formation and limited downstream adaptability of 3D cultures have hindered mainstream adoption of these systems for biological applications, especially for low- and moderate-throughput assays commonly used in biomedical research. Here we build on our recent development of a 384-well hanging drop plate for spheroid culture to design a complementary spheroid transfer and imaging (TRIM) plate. The low-aspect ratio wells of the TRIM plate facilitated highfidelity, user-independent, contact-based collection of hanging drop spheroids. Using the TRIM plate, we demonstrated several downstream analyses, including bulk tissue collection for flow cytometry, high-resolution low working-distance immersion imaging, and timely reagent delivery for enzymatic studies. Low working-distance multiphoton imaging revealed a cell type–dependent, macroscopic spheroid structure. Unlike ovarian cancer spheroids, which formed loose, disk-shaped spheroids, human mammary fibroblasts formed tight, spherical, and nutrient-limited spheroids. Beyond the applications we describe here, we expect the hanging drop spheroid plate and complementary TRIM plate to facilitate analyses of spheroids across the spectrum of throughput, particularly for bulk collection of spheroids and high-content imaging. PMID:24051516

Transfer, imaging, and analysis plate for facile handling of 384 hanging drop 3D tissue spheroids.

PubMed

Cavnar, Stephen P; Salomonsson, Emma; Luker, Kathryn E; Luker, Gary D; Takayama, Shuichi

2014-04-01

Three-dimensional culture systems bridge the experimental gap between in vivo and in vitro physiology. However, nonstandardized formation and limited downstream adaptability of 3D cultures have hindered mainstream adoption of these systems for biological applications, especially for low- and moderate-throughput assays commonly used in biomedical research. Here we build on our recent development of a 384-well hanging drop plate for spheroid culture to design a complementary spheroid transfer and imaging (TRIM) plate. The low-aspect ratio wells of the TRIM plate facilitated high-fidelity, user-independent, contact-based collection of hanging drop spheroids. Using the TRIM plate, we demonstrated several downstream analyses, including bulk tissue collection for flow cytometry, high-resolution low working-distance immersion imaging, and timely reagent delivery for enzymatic studies. Low working-distance multiphoton imaging revealed a cell type-dependent, macroscopic spheroid structure. Unlike ovarian cancer spheroids, which formed loose, disk-shaped spheroids, human mammary fibroblasts formed tight, spherical, and nutrient-limited spheroids. Beyond the applications we describe here, we expect the hanging drop spheroid plate and complementary TRIM plate to facilitate analyses of spheroids across the spectrum of throughput, particularly for bulk collection of spheroids and high-content imaging.

Lensless digital holographic microscopy and its applications in biomedicine and environmental monitoring.

PubMed

Wu, Yichen; Ozcan, Aydogan

2018-03-01

Optical compound microscope has been a major tool in biomedical imaging for centuries. Its performance relies on relatively complicated, bulky and expensive lenses and alignment mechanics. In contrast, the lensless microscope digitally reconstructs microscopic images of specimens without using any lenses, as a result of which it can be made much smaller, lighter and lower-cost. Furthermore, the limited space-bandwidth product of objective lenses in a conventional microscope can be significantly surpassed by a lensless microscope. Such lensless imaging designs have enabled high-resolution and high-throughput imaging of specimens using compact, portable and cost-effective devices to potentially address various point-of-care, global-health and telemedicine related challenges. In this review, we discuss the operation principles and the methods behind lensless digital holographic on-chip microscopy. We also go over various applications that are enabled by cost-effective and compact implementations of lensless microscopy, including some recent work on air quality monitoring, which utilized machine learning for high-throughput and accurate quantification of particulate matter in air. Finally, we conclude with a brief future outlook of this computational imaging technology. Copyright © 2017 Elsevier Inc. All rights reserved.

Microscale Laminar Vortices for High-Purity Extraction and Release of Circulating Tumor Cells.

PubMed

Hur, Soojung Claire; Che, James; Di Carlo, Dino

2017-01-01

Circulating tumor cells (CTCs) are disseminated tumor cells that reflect the tumors of origin and can provide a liquid biopsy that would potentially enable noninvasive tumor profiling, treatment monitoring, and identification of targeted treatments. Accurate and rapid purification of CTCs holds great potential to improve cancer care but the task remains technically challenging. Microfluidic isolation of CTCs within microscale vortices enables high-throughput and size-based purification of rare CTCs from bodily fluids. Collected cells are highly pure, viable, and easily accessible, allowing seamless integration with various downstream applications. Here, we describe how to fabricate the High-Throughput Vortex Chip (Vortex-HT) and to process diluted whole blood for CTC collection. Lastly, immunostaining and imaging protocols for CTC classification and corresponding CTC image galleries are reported.

Electron beam throughput from raster to imaging

NASA Astrophysics Data System (ADS)

Zywno, Marek

2016-12-01

Two architectures of electron beam tools are presented: single beam MEBES Exara designed and built by Etec Systems for mask writing, and the Reflected E-Beam Lithography tool (REBL), designed and built by KLA-Tencor under a DARPA Agreement No. HR0011-07-9-0007. Both tools have implemented technologies not used before to achieve their goals. The MEBES X, renamed Exara for marketing purposes, used an air bearing stage running in vacuum to achieve smooth continuous scanning. The REBL used 2 dimensional imaging to distribute charge to a 4k pixel swath to achieve writing times on the order of 1 wafer per hour, scalable to throughput approaching optical projection tools. Three stage architectures were designed for continuous scanning of wafers: linear maglev, rotary maglev, and dual linear maglev.

A rapid enzymatic assay for high-throughput screening of adenosine-producing strains

PubMed Central

Dong, Huina; Zu, Xin; Zheng, Ping; Zhang, Dawei

2015-01-01

Adenosine is a major local regulator of tissue function and industrially useful as precursor for the production of medicinal nucleoside substances. High-throughput screening of adenosine overproducers is important for industrial microorganism breeding. An enzymatic assay of adenosine was developed by combined adenosine deaminase (ADA) with indophenol method. The ADA catalyzes the cleavage of adenosine to inosine and NH3, the latter can be accurately determined by indophenol method. The assay system was optimized to deliver a good performance and could tolerate the addition of inorganic salts and many nutrition components to the assay mixtures. Adenosine could be accurately determined by this assay using 96-well microplates. Spike and recovery tests showed that this assay can accurately and reproducibly determine increases in adenosine in fermentation broth without any pretreatment to remove proteins and potentially interfering low-molecular-weight molecules. This assay was also applied to high-throughput screening for high adenosine-producing strains. The high selectivity and accuracy of the ADA assay provides rapid and high-throughput analysis of adenosine in large numbers of samples. PMID:25580842

Reflective optical imaging system

DOEpatents

Shafer, David R.

2000-01-01

An optical system compatible with short wavelength (extreme ultraviolet) radiation comprising four reflective elements for projecting a mask image onto a substrate. The four optical elements are characterized in order from object to image as convex, concave, convex and concave mirrors. The optical system is particularly suited for step and scan lithography methods. The invention increases the slit dimensions associated with ringfield scanning optics, improves wafer throughput and allows higher semiconductor device density.

Reflective optical imaging method and circuit

DOEpatents

Shafer, David R.

2001-01-01

An optical system compatible with short wavelength (extreme ultraviolet) radiation comprising four reflective elements for projecting a mask image onto a substrate. The four optical elements are characterized in order from object to image as convex, concave, convex and concave mirrors. The optical system is particularly suited for step and scan lithography methods. The invention increases the slit dimensions associated with ringfield scanning optics, improves wafer throughput and allows higher semiconductor device density.

Fluorescent and Lanthanide Labeling for Ligand Screens, Assays, and Imaging

PubMed Central

Josan, Jatinder S.; De Silva, Channa R.; Yoo, Byunghee; Lynch, Ronald M.; Pagel, Mark D.; Vagner, Josef; Hruby, Victor J.

2012-01-01

The use of fluorescent (or luminescent) and metal contrast agents in high-throughput screens, in vitro assays, and molecular imaging procedures has rapidly expanded in recent years. Here we describe the development and utility of high-affinity ligands for cancer theranostics and other in vitro screening studies. In this context, we also illustrate the syntheses and use of heteromultivalent ligands as targeted imaging agents. PMID:21318902

Digital image processing for the earth resources technology satellite data.

NASA Technical Reports Server (NTRS)

Will, P. M.; Bakis, R.; Wesley, M. A.

1972-01-01

This paper discusses the problems of digital processing of the large volumes of multispectral image data that are expected to be received from the ERTS program. Correction of geometric and radiometric distortions are discussed and a byte oriented implementation is proposed. CPU timing estimates are given for a System/360 Model 67, and show that a processing throughput of 1000 image sets per week is feasible.

Interleaved EPI based fMRI improved by multiplexed sensitivity encoding (MUSE) and simultaneous multi-band imaging.

PubMed

Chang, Hing-Chiu; Gaur, Pooja; Chou, Ying-hui; Chu, Mei-Lan; Chen, Nan-kuei

2014-01-01

Functional magnetic resonance imaging (fMRI) is a non-invasive and powerful imaging tool for detecting brain activities. The majority of fMRI studies are performed with single-shot echo-planar imaging (EPI) due to its high temporal resolution. Recent studies have demonstrated that, by increasing the spatial-resolution of fMRI, previously unidentified neuronal networks can be measured. However, it is challenging to improve the spatial resolution of conventional single-shot EPI based fMRI. Although multi-shot interleaved EPI is superior to single-shot EPI in terms of the improved spatial-resolution, reduced geometric distortions, and sharper point spread function (PSF), interleaved EPI based fMRI has two main limitations: 1) the imaging throughput is lower in interleaved EPI; 2) the magnitude and phase signal variations among EPI segments (due to physiological noise, subject motion, and B0 drift) are translated to significant in-plane aliasing artifact across the field of view (FOV). Here we report a method that integrates multiple approaches to address the technical limitations of interleaved EPI-based fMRI. Firstly, the multiplexed sensitivity-encoding (MUSE) post-processing algorithm is used to suppress in-plane aliasing artifacts resulting from time-domain signal instabilities during dynamic scans. Secondly, a simultaneous multi-band interleaved EPI pulse sequence, with a controlled aliasing scheme incorporated, is implemented to increase the imaging throughput. Thirdly, the MUSE algorithm is then generalized to accommodate fMRI data obtained with our multi-band interleaved EPI pulse sequence, suppressing both in-plane and through-plane aliasing artifacts. The blood-oxygenation-level-dependent (BOLD) signal detectability and the scan throughput can be significantly improved for interleaved EPI-based fMRI. Our human fMRI data obtained from 3 Tesla systems demonstrate the effectiveness of the developed methods. It is expected that future fMRI studies requiring high spatial-resolvability and fidelity will largely benefit from the reported techniques.

Dual-modality NIRF-MRI cubosomes and hexosomes: High throughput formulation and in vivo biodistribution.

PubMed

Tran, Nhiem; Bye, Nicole; Moffat, Bradford A; Wright, David K; Cuddihy, Andrew; Hinton, Tracey M; Hawley, Adrian M; Reynolds, Nicholas P; Waddington, Lynne J; Mulet, Xavier; Turnley, Ann M; Morganti-Kossmann, M Cristina; Muir, Benjamin W

2017-02-01

Engineered nanoparticles with multiple complementary imaging modalities are of great benefit to the rapid treatment and diagnosis of disease in various organs. Herein, we report the formulation of cubosomes and hexosomes that carry multiple amphiphilic imaging contrast agents in their self-assembled lipid bilayers. This is the first report of the use of both near infrared fluorescent (NIRF) imaging and gadolinium lipid based magnetic resonance (MR) imaging modalities in cubosomes and hexosomes. High-throughput screening was used to rapidly optimize formulations with desirable nano-architectures and low in vitro cytotoxicity. The dual-modal imaging nanoparticles in vivo biodistribution and organ specific contrast enhancement were then studied. The NIRF in vivo imaging results indicated accumulation of both cubosomes and hexosomes in the liver and spleen of mice up to 20h post-injection. Remarkably, the biodistribution of the nanoparticle formulations was affected by the mesophase (i.e. cubic or hexagonal), a finding of significant importance for the future use of these compounds, with hexosomes showing higher accumulation in the spleen than the liver compared to cubosomes. Furthermore, in vivo MRI data of animals injected with either type of lyotropic liquid crystal nanoparticle displayed enhanced contrast in the liver and spleen. Copyright © 2016 Elsevier B.V. All rights reserved.

Microfluidics platform for single-shot dose-response analysis of chloride channel-modulating compounds.

PubMed

Jin, Byung-Ju; Ko, Eun-A; Namkung, Wan; Verkman, A S

2013-10-07

We previously developed cell-based kinetics assays of chloride channel modulators utilizing genetically encoded yellow fluorescent proteins. Fluorescence platereader-based high-throughput screens yielded small-molecule activators and inhibitors of the cAMP-activated chloride channel CFTR and calcium-activated chloride channels, including TMEM16A. Here, we report a microfluidics platform for single-shot determination of concentration-activity relations in which a 1.5 × 1.5 mm square area of adherent cultured cells is exposed for 5-10 min to a pseudo-logarithmic gradient of test compound generated by iterative, two-component channel mixing. Cell fluorescence is imaged following perfusion with an iodide-containing solution to give iodide influx rate at each location in the image field, thus quantifying modulator effects over a wide range of concentrations in a single measurement. IC50 determined for CFTR and TMEM16A activators and inhibitors by single-shot microfluidics were in agreement with conventional plate reader measurements. The microfluidics approach developed here may accelerate the discovery and characterization of chloride channel-targeted drugs.

Tracking C. elegans and its neuromuscular activity using NemaFlex II

NASA Astrophysics Data System (ADS)

van Bussel, Frank; Rahman, Mizanur; Blawzdziewicz, Jerzy; Vanapalli, Siva

NemaFlex is a recently developed experimental platform designed to analyze the movement and muscular strength of crawling C. elegans. Physically it is a microfluidic device consisting of an array of deformable PDMS pillars, with which the C. elegans interacts in the course of moving through the system; image data is then acquired through a transparent top plate. The software component uses this image data to track the worm's movements and measure pillar deflections and thereby the forces exerted by the worm, in a fully automated, high-throughput manner. In order to correlate the force results with muscle activations the pillar deflections need to be precisely associated with mechanical contact on the worm's body, which requires accurate determination and representation of the body's position within the complex background. Here we discuss issues encountered in extracting this position data from the surrounding environment.

Malic Acid Carbon Dots: From Super-resolution Live-Cell Imaging to Highly Efficient Separation.

PubMed

Zhi, Bo; Cui, Yi; Wang, Shengyang; Frank, Benjamin P; Williams, Denise N; Brown, Richard P; Melby, Eric S; Hamers, Robert J; Rosenzweig, Zeev; Fairbrother, D Howard; Orr, Galya; Haynes, Christy L

2018-06-15

As-synthesized malic acid carbon dots are found to possess photoblinking properties that are outstanding and superior compared to those of conventional dyes. Considering their excellent biocompatibility, malic acid carbon dots are suitable for super-resolution fluorescence localization microscopy under a variety of conditions, as we demonstrate in fixed and live trout gill epithelial cells. In addition, during imaging experiments, the so-called "excitation wavelength-dependent" emission was not observed for individual as-made malic acid carbon dots, which motivated us to develop a time-saving and high-throughput separation technique to isolate malic acid carbon dots into fractions of different particle size distributions using C 18 reversed-phase silica gel column chromatography. This post-treatment allowed us to determine how particle size distribution influences the optical properties of malic acid carbon dot fractions, that is, optical band gap energies and photoluminescence behaviors.

Automated High-Throughput Quantification of Mitotic Spindle Positioning from DIC Movies of Caenorhabditis Embryos

PubMed Central

Cluet, David; Spichty, Martin; Delattre, Marie

2014-01-01

The mitotic spindle is a microtubule-based structure that elongates to accurately segregate chromosomes during anaphase. Its position within the cell also dictates the future cell cleavage plan, thereby determining daughter cell orientation within a tissue or cell fate adoption for polarized cells. Therefore, the mitotic spindle ensures at the same time proper cell division and developmental precision. Consequently, spindle dynamics is the matter of intensive research. Among the different cellular models that have been explored, the one-cell stage C. elegans embryo has been an essential and powerful system to dissect the molecular and biophysical basis of spindle elongation and positioning. Indeed, in this large and transparent cell, spindle poles (or centrosomes) can be easily detected from simple DIC microscopy by human eyes. To perform quantitative and high-throughput analysis of spindle motion, we developed a computer program ACT for Automated-Centrosome-Tracking from DIC movies of C. elegans embryos. We therefore offer an alternative to the image acquisition and processing of transgenic lines expressing fluorescent spindle markers. Consequently, experiments on large sets of cells can be performed with a simple setup using inexpensive microscopes. Moreover, analysis of any mutant or wild-type backgrounds is accessible because laborious rounds of crosses with transgenic lines become unnecessary. Last, our program allows spindle detection in other nematode species, offering the same quality of DIC images but for which techniques of transgenesis are not accessible. Thus, our program also opens the way towards a quantitative evolutionary approach of spindle dynamics. Overall, our computer program is a unique macro for the image- and movie-processing platform ImageJ. It is user-friendly and freely available under an open-source licence. ACT allows batch-wise analysis of large sets of mitosis events. Within 2 minutes, a single movie is processed and the accuracy of the automated tracking matches the precision of the human eye. PMID:24763198

Higher Throughput Toxicokinetics to Allow Extrapolation (EPA-Japan Bilateral EDSP meeting)

EPA Science Inventory

As part of "Ongoing EDSP Directions & Activities" I will present CSS research on high throughput toxicokinetics, including in vitro data and models to allow rapid determination of the real world doses that may cause endocrine disruption.

Dynamic Environmental Photosynthetic Imaging Reveals Emergent Phenotypes

DOE PAGES

Cruz, Jeffrey A.; Savage, Linda J.; Zegarac, Robert; ...

2016-06-22

Understanding and improving the productivity and robustness of plant photosynthesis requires high-throughput phenotyping under environmental conditions that are relevant to the field. Here we demonstrate the dynamic environmental photosynthesis imager (DEPI), an experimental platform for integrated, continuous, and high-throughput measurements of photosynthetic parameters during plant growth under reproducible yet dynamic environmental conditions. Using parallel imagers obviates the need to move plants or sensors, reducing artifacts and allowing simultaneous measurement on large numbers of plants. As a result, DEPI can reveal phenotypes that are not evident under standard laboratory conditions but emerge under progressively more dynamic illumination. We show examples inmore » mutants of Arabidopsis of such “emergent phenotypes” that are highly transient and heterogeneous, appearing in different leaves under different conditions and depending in complex ways on both environmental conditions and plant developmental age. Finally, these emergent phenotypes appear to be caused by a range of phenomena, suggesting that such previously unseen processes are critical for plant responses to dynamic environments.« less

Automation of Technology for Cancer Research.

PubMed

van der Ent, Wietske; Veneman, Wouter J; Groenewoud, Arwin; Chen, Lanpeng; Tulotta, Claudia; Hogendoorn, Pancras C W; Spaink, Herman P; Snaar-Jagalska, B Ewa

2016-01-01

Zebrafish embryos can be obtained for research purposes in large numbers at low cost and embryos develop externally in limited space, making them highly suitable for high-throughput cancer studies and drug screens. Non-invasive live imaging of various processes within the larvae is possible due to their transparency during development, and a multitude of available fluorescent transgenic reporter lines.To perform high-throughput studies, handling large amounts of embryos and larvae is required. With such high number of individuals, even minute tasks may become time-consuming and arduous. In this chapter, an overview is given of the developments in the automation of various steps of large scale zebrafish cancer research for discovering important cancer pathways and drugs for the treatment of human disease. The focus lies on various tools developed for cancer cell implantation, embryo handling and sorting, microfluidic systems for imaging and drug treatment, and image acquisition and analysis. Examples will be given of employment of these technologies within the fields of toxicology research and cancer research.

Automated evaluation of AIMS images: an approach to minimize evaluation variability

NASA Astrophysics Data System (ADS)

Dürr, Arndt C.; Arndt, Martin; Fiebig, Jan; Weiss, Samuel

2006-05-01

Defect disposition and qualification with stepper simulating AIMS tools on advanced masks of the 90nm node and below is key to match the customer's expectations for "defect free" masks, i.e. masks containing only non-printing design variations. The recently available AIMS tools allow for a large degree of automated measurements enhancing the throughput of masks and hence reducing cycle time - up to 50 images can be recorded per hour. However, this amount of data still has to be evaluated by hand which is not only time-consuming but also error prone and exhibits a variability depending on the person doing the evaluation which adds to the tool intrinsic variability and decreases the reliability of the evaluation. In this paper we present the results of an MatLAB based algorithm which automatically evaluates AIMS images. We investigate its capabilities regarding throughput, reliability and matching with handmade evaluation for a large variety of dark and clear defects and discuss the limitations of an automated AIMS evaluation algorithm.

High-throughput microscopy must re-invent the microscope rather than speed up its functions

PubMed Central

Oheim, M

2007-01-01

Knowledge gained from the revolutions in genomics and proteomics has helped to identify many of the key molecules involved in cellular signalling. Researchers, both in academia and in the pharmaceutical industry, now screen, at a sub-cellular level, where and when these proteins interact. Fluorescence imaging and molecular labelling combine to provide a powerful tool for real-time functional biochemistry with molecular resolution. However, they traditionally have been work-intensive, required trained personnel, and suffered from low through-put due to sample preparation, loading and handling. The need for speeding up microscopy is apparent from the tremendous complexity of cellular signalling pathways, the inherent biological variability, as well as the possibility that the same molecule plays different roles in different sub-cellular compartments. Research institutes and companies have teamed up to develop imaging cytometers of ever-increasing complexity. However, to truly go high-speed, sub-cellular imaging must free itself from the rigid framework of current microscopes. PMID:17603553

Vivaldi: A Domain-Specific Language for Volume Processing and Visualization on Distributed Heterogeneous Systems.

PubMed

Choi, Hyungsuk; Choi, Woohyuk; Quan, Tran Minh; Hildebrand, David G C; Pfister, Hanspeter; Jeong, Won-Ki

2014-12-01

As the size of image data from microscopes and telescopes increases, the need for high-throughput processing and visualization of large volumetric data has become more pressing. At the same time, many-core processors and GPU accelerators are commonplace, making high-performance distributed heterogeneous computing systems affordable. However, effectively utilizing GPU clusters is difficult for novice programmers, and even experienced programmers often fail to fully leverage the computing power of new parallel architectures due to their steep learning curve and programming complexity. In this paper, we propose Vivaldi, a new domain-specific language for volume processing and visualization on distributed heterogeneous computing systems. Vivaldi's Python-like grammar and parallel processing abstractions provide flexible programming tools for non-experts to easily write high-performance parallel computing code. Vivaldi provides commonly used functions and numerical operators for customized visualization and high-throughput image processing applications. We demonstrate the performance and usability of Vivaldi on several examples ranging from volume rendering to image segmentation.

Oufti: An integrated software package for high-accuracy, high-throughput quantitative microscopy analysis

PubMed Central

Paintdakhi, Ahmad; Parry, Bradley; Campos, Manuel; Irnov, Irnov; Elf, Johan; Surovtsev, Ivan; Jacobs-Wagner, Christine

2016-01-01

Summary With the realization that bacteria display phenotypic variability among cells and exhibit complex subcellular organization critical for cellular function and behavior, microscopy has re-emerged as a primary tool in bacterial research during the last decade. However, the bottleneck in today’s single-cell studies is quantitative image analysis of cells and fluorescent signals. Here, we address current limitations through the development of Oufti, a stand-alone, open-source software package for automated measurements of microbial cells and fluorescence signals from microscopy images. Oufti provides computational solutions for tracking touching cells in confluent samples, handles various cell morphologies, offers algorithms for quantitative analysis of both diffraction and non-diffraction-limited fluorescence signals, and is scalable for high-throughput analysis of massive datasets, all with subpixel precision. All functionalities are integrated in a single package. The graphical user interface, which includes interactive modules for segmentation, image analysis, and post-processing analysis, makes the software broadly accessible to users irrespective of their computational skills. PMID:26538279

A computational image analysis glossary for biologists.

PubMed

Roeder, Adrienne H K; Cunha, Alexandre; Burl, Michael C; Meyerowitz, Elliot M

2012-09-01

Recent advances in biological imaging have resulted in an explosion in the quality and quantity of images obtained in a digital format. Developmental biologists are increasingly acquiring beautiful and complex images, thus creating vast image datasets. In the past, patterns in image data have been detected by the human eye. Larger datasets, however, necessitate high-throughput objective analysis tools to computationally extract quantitative information from the images. These tools have been developed in collaborations between biologists, computer scientists, mathematicians and physicists. In this Primer we present a glossary of image analysis terms to aid biologists and briefly discuss the importance of robust image analysis in developmental studies.

Whole-animal imaging with high spatio-temporal resolution

NASA Astrophysics Data System (ADS)

Chhetri, Raghav; Amat, Fernando; Wan, Yinan; Höckendorf, Burkhard; Lemon, William C.; Keller, Philipp J.

2016-03-01

We developed isotropic multiview (IsoView) light-sheet microscopy in order to image fast cellular dynamics, such as cell movements in an entire developing embryo or neuronal activity throughput an entire brain or nervous system, with high resolution in all dimensions, high imaging speeds, good physical coverage and low photo-damage. To achieve high temporal resolution and high spatial resolution at the same time, IsoView microscopy rapidly images large specimens via simultaneous light-sheet illumination and fluorescence detection along four orthogonal directions. In a post-processing step, these four views are then combined by means of high-throughput multiview deconvolution to yield images with a system resolution of ≤ 450 nm in all three dimensions. Using IsoView microscopy, we performed whole-animal functional imaging of Drosophila embryos and larvae at a spatial resolution of 1.1-2.5 μm and at a temporal resolution of 2 Hz for up to 9 hours. We also performed whole-brain functional imaging in larval zebrafish and multicolor imaging of fast cellular dynamics across entire, gastrulating Drosophila embryos with isotropic, sub-cellular resolution. Compared with conventional (spatially anisotropic) light-sheet microscopy, IsoView microscopy improves spatial resolution at least sevenfold and decreases resolution anisotropy at least threefold. Compared with existing high-resolution light-sheet techniques, such as lattice lightsheet microscopy or diSPIM, IsoView microscopy effectively doubles the penetration depth and provides subsecond temporal resolution for specimens 400-fold larger than could previously be imaged.

Netest: A Tool to Measure the Maximum Burst Size, Available Bandwidth and Achievable Throughput

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jin, Guojun; Tierney, Brian

2003-01-31

Distinguishing available bandwidth and achievable throughput is essential for improving network applications' performance. Achievable throughput is the throughput considering a number of factors such as network protocol, host speed, network path, and TCP buffer space, where as available bandwidth only considers the network path. Without understanding this difference, trying to improve network applications' performance is like ''blind men feeling the elephant'' [4]. In this paper, we define and distinguish bandwidth and throughput, and debate which part of each is achievable and which is available. Also, we introduce and discuss a new concept - Maximum Burst Size that is crucial tomore » the network performance and bandwidth sharing. A tool, netest, is introduced to help users to determine the available bandwidth, and provides information to achieve better throughput with fairness of sharing the available bandwidth, thus reducing misuse of the network.« less

MicroFilament Analyzer, an image analysis tool for quantifying fibrillar orientation, reveals changes in microtubule organization during gravitropism.

PubMed

Jacques, Eveline; Buytaert, Jan; Wells, Darren M; Lewandowski, Michal; Bennett, Malcolm J; Dirckx, Joris; Verbelen, Jean-Pierre; Vissenberg, Kris

2013-06-01

Image acquisition is an important step in the study of cytoskeleton organization. As visual interpretations and manual measurements of digital images are prone to errors and require a great amount of time, a freely available software package named MicroFilament Analyzer (MFA) was developed. The goal was to provide a tool that facilitates high-throughput analysis to determine the orientation of filamentous structures on digital images in a more standardized, objective and repeatable way. Here, the rationale and applicability of the program is demonstrated by analyzing the microtubule patterns in epidermal cells of control and gravi-stimulated Arabidopsis thaliana roots. Differential expansion of cells on either side of the root results in downward bending of the root tip. As cell expansion depends on the properties of the cell wall, this may imply a differential orientation of cellulose microfibrils. As cellulose deposition is orchestrated by cortical microtubules, the microtubule patterns were analyzed. The MFA program detects the filamentous structures on the image and identifies the main orientation(s) within individual cells. This revealed four distinguishable microtubule patterns in root epidermal cells. The analysis indicated that gravitropic stimulation and developmental age are both significant factors that determine microtubule orientation. Moreover, the data show that an altered microtubule pattern does not precede differential expansion. Other possible applications are also illustrated, including field emission scanning electron micrographs of cellulose microfibrils in plant cell walls and images of fluorescent actin. © 2013 The Authors The Plant Journal © 2013 John Wiley & Sons Ltd.

Sequencing the Connectome

PubMed Central

Zador, Anthony M.; Dubnau, Joshua; Oyibo, Hassana K.; Zhan, Huiqing; Cao, Gang; Peikon, Ian D.

2012-01-01

Connectivity determines the function of neural circuits. Historically, circuit mapping has usually been viewed as a problem of microscopy, but no current method can achieve high-throughput mapping of entire circuits with single neuron precision. Here we describe a novel approach to determining connectivity. We propose BOINC (“barcoding of individual neuronal connections”), a method for converting the problem of connectivity into a form that can be read out by high-throughput DNA sequencing. The appeal of using sequencing is that its scale—sequencing billions of nucleotides per day is now routine—is a natural match to the complexity of neural circuits. An inexpensive high-throughput technique for establishing circuit connectivity at single neuron resolution could transform neuroscience research. PMID:23109909

A Fully Automated High-Throughput Zebrafish Behavioral Ototoxicity Assay.

PubMed

Todd, Douglas W; Philip, Rohit C; Niihori, Maki; Ringle, Ryan A; Coyle, Kelsey R; Zehri, Sobia F; Zabala, Leanne; Mudery, Jordan A; Francis, Ross H; Rodriguez, Jeffrey J; Jacob, Abraham

2017-08-01

Zebrafish animal models lend themselves to behavioral assays that can facilitate rapid screening of ototoxic, otoprotective, and otoregenerative drugs. Structurally similar to human inner ear hair cells, the mechanosensory hair cells on their lateral line allow the zebrafish to sense water flow and orient head-to-current in a behavior called rheotaxis. This rheotaxis behavior deteriorates in a dose-dependent manner with increased exposure to the ototoxin cisplatin, thereby establishing itself as an excellent biomarker for anatomic damage to lateral line hair cells. Building on work by our group and others, we have built a new, fully automated high-throughput behavioral assay system that uses automated image analysis techniques to quantify rheotaxis behavior. This novel system consists of a custom-designed swimming apparatus and imaging system consisting of network-controlled Raspberry Pi microcomputers capturing infrared video. Automated analysis techniques detect individual zebrafish, compute their orientation, and quantify the rheotaxis behavior of a zebrafish test population, producing a powerful, high-throughput behavioral assay. Using our fully automated biological assay to test a standardized ototoxic dose of cisplatin against varying doses of compounds that protect or regenerate hair cells may facilitate rapid translation of candidate drugs into preclinical mammalian models of hearing loss.

Application Of Empirical Phase Diagrams For Multidimensional Data Visualization Of High Throughput Microbatch Crystallization Experiments.

PubMed

Klijn, Marieke E; Hubbuch, Jürgen

2018-04-27

Protein phase diagrams are a tool to investigate cause and consequence of solution conditions on protein phase behavior. The effects are scored according to aggregation morphologies such as crystals or amorphous precipitates. Solution conditions affect morphological features, such as crystal size, as well as kinetic features, such as crystal growth time. Common used data visualization techniques include individual line graphs or symbols-based phase diagrams. These techniques have limitations in terms of handling large datasets, comprehensiveness or completeness. To eliminate these limitations, morphological and kinetic features obtained from crystallization images generated with high throughput microbatch experiments have been visualized with radar charts in combination with the empirical phase diagram (EPD) method. Morphological features (crystal size, shape, and number, as well as precipitate size) and kinetic features (crystal and precipitate onset and growth time) are extracted for 768 solutions with varying chicken egg white lysozyme concentration, salt type, ionic strength and pH. Image-based aggregation morphology and kinetic features were compiled into a single and easily interpretable figure, thereby showing that the EPD method can support high throughput crystallization experiments in its data amount as well as its data complexity. Copyright © 2018. Published by Elsevier Inc.

A compact high-resolution 3-D imaging spectrometer for discovering Oases on Mars

USGS Publications Warehouse

Ge, J.; Ren, D.; Lunine, J.I.; Brown, R.H.; Yelle, R.V.; Soderblom, L.A.; ,

2002-01-01

A new design for a very lightweight, very high throughput reflectance sectrometer enabled by two new technologies being developed is presented. These new technologies include integral field unit optics to enable simultaneous imaging and spectroscopy at high spatial resolution with an infrared (IR) array, and silicon grisms to enable compact and high-resolution spectroscopy.

Characterizing nanoscale scanning probes using electron microscopy: A novel fixture and a practical guide

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jacobs, Tevis D. B., E-mail: tjacobs@pitt.edu; Wabiszewski, Graham E.; Goodman, Alexander J.

2016-01-15

The nanoscale geometry of probe tips used for atomic force microscopy (AFM) measurements determines the lateral resolution, contributes to the strength of the tip-surface interaction, and can be a significant source of uncertainty in the quantitative analysis of results. While inverse imaging of the probe tip has been used successfully to determine probe tip geometry, direct observation of the tip profile using electron microscopy (EM) confers several advantages: it provides direct (rather than indirect) imaging, requires fewer algorithmic parameters, and does not require bringing the tip into contact with a sample. In the past, EM-based observation of the probe tipmore » has been achieved using ad hoc mounting methods that are constrained by low throughput, the risk of contamination, and repeatability issues. We report on a probe fixture designed for use in a commercial transmission electron microscope that enables repeatable mounting of multiple AFM probes as well as a reference grid for beam alignment. This communication describes the design, fabrication, and advantages of this probe fixture, including full technical drawings for machining. Further, best practices are discussed for repeatable, non-destructive probe imaging. Finally, examples of the fixture’s use are described, including characterization of common commercial AFM probes in their out-of-the-box condition.« less

Characterizing nanoscale scanning probes using electron microscopy: A novel fixture and a practical guide

NASA Astrophysics Data System (ADS)

Jacobs, Tevis D. B.; Wabiszewski, Graham E.; Goodman, Alexander J.; Carpick, Robert W.

2016-01-01

The nanoscale geometry of probe tips used for atomic force microscopy (AFM) measurements determines the lateral resolution, contributes to the strength of the tip-surface interaction, and can be a significant source of uncertainty in the quantitative analysis of results. While inverse imaging of the probe tip has been used successfully to determine probe tip geometry, direct observation of the tip profile using electron microscopy (EM) confers several advantages: it provides direct (rather than indirect) imaging, requires fewer algorithmic parameters, and does not require bringing the tip into contact with a sample. In the past, EM-based observation of the probe tip has been achieved using ad hoc mounting methods that are constrained by low throughput, the risk of contamination, and repeatability issues. We report on a probe fixture designed for use in a commercial transmission electron microscope that enables repeatable mounting of multiple AFM probes as well as a reference grid for beam alignment. This communication describes the design, fabrication, and advantages of this probe fixture, including full technical drawings for machining. Further, best practices are discussed for repeatable, non-destructive probe imaging. Finally, examples of the fixture's use are described, including characterization of common commercial AFM probes in their out-of-the-box condition.

High Throughput Determination of Tetramine in Drinking ...

EPA Pesticide Factsheets

Report The sampling and analytical procedure (SAP) presented herein, describes a method for the high throughput determination of tetramethylene disulfotetramine in drinking water by solid phase extraction and isotope dilution gas chromatography/mass spectrometry. This method, which will be included in the SAM, is expected to provide the Water Laboratory Alliance, as part of EPA’s Environmental Response Laboratory Network, with a more reliable and faster means of analyte collection and measurement.

A high-throughput lab-on-a-chip interface for zebrafish embryo tests in drug discovery and ecotoxicology

NASA Astrophysics Data System (ADS)

Zhu, Feng; Akagi, Jin; Hall, Chris J.; Crosier, Kathryn E.; Crosier, Philip S.; Delaage, Pierre; Wlodkowic, Donald

2013-12-01

Drug discovery screenings performed on zebrafish embryos mirror with a high level of accuracy. The tests usually performed on mammalian animal models, and the fish embryo toxicity assay (FET) is one of the most promising alternative approaches to acute ecotoxicity testing with adult fish. Notwithstanding this, conventional methods utilising 96-well microtiter plates and manual dispensing of fish embryos are very time-consuming. They rely on laborious and iterative manual pipetting that is a main source of analytical errors and low throughput. In this work, we present development of a miniaturised and high-throughput Lab-on-a-Chip (LOC) platform for automation of FET assays. The 3D high-density LOC array was fabricated in poly-methyl methacrylate (PMMA) transparent thermoplastic using infrared laser micromachining while the off-chip interfaces were fabricated using additive manufacturing processes (FDM and SLA). The system's design facilitates rapid loading and immobilization of a large number of embryos in predefined clusters of traps during continuous microperfusion of drugs/toxins. It has been conceptually designed to seamlessly interface with both upright and inverted fluorescent imaging systems and also to directly interface with conventional microtiter plate readers that accept 96-well plates. We also present proof-of-concept interfacing with a high-speed imaging cytometer Plate RUNNER HD® capable of multispectral image acquisition with resolution of up to 8192 x 8192 pixels and depth of field of about 40 μm. Furthermore, we developed a miniaturized and self-contained analytical device interfaced with a miniaturized USB microscope. This system modification is capable of performing rapid imaging of multiple embryos at a low resolution for drug toxicity analysis.

Dissecting the Phenotypic Components of Crop Plant Growth and Drought Responses Based on High-Throughput Image Analysis[W][OPEN

PubMed Central

Chen, Dijun; Neumann, Kerstin; Friedel, Swetlana; Kilian, Benjamin; Chen, Ming; Altmann, Thomas; Klukas, Christian

2014-01-01

Significantly improved crop varieties are urgently needed to feed the rapidly growing human population under changing climates. While genome sequence information and excellent genomic tools are in place for major crop species, the systematic quantification of phenotypic traits or components thereof in a high-throughput fashion remains an enormous challenge. In order to help bridge the genotype to phenotype gap, we developed a comprehensive framework for high-throughput phenotype data analysis in plants, which enables the extraction of an extensive list of phenotypic traits from nondestructive plant imaging over time. As a proof of concept, we investigated the phenotypic components of the drought responses of 18 different barley (Hordeum vulgare) cultivars during vegetative growth. We analyzed dynamic properties of trait expression over growth time based on 54 representative phenotypic features. The data are highly valuable to understand plant development and to further quantify growth and crop performance features. We tested various growth models to predict plant biomass accumulation and identified several relevant parameters that support biological interpretation of plant growth and stress tolerance. These image-based traits and model-derived parameters are promising for subsequent genetic mapping to uncover the genetic basis of complex agronomic traits. Taken together, we anticipate that the analytical framework and analysis results presented here will be useful to advance our views of phenotypic trait components underlying plant development and their responses to environmental cues. PMID:25501589

High-throughput microfluidic line scan imaging for cytological characterization

NASA Astrophysics Data System (ADS)

Hutcheson, Joshua A.; Powless, Amy J.; Majid, Aneeka A.; Claycomb, Adair; Fritsch, Ingrid; Balachandran, Kartik; Muldoon, Timothy J.

2015-03-01

Imaging cells in a microfluidic chamber with an area scan camera is difficult due to motion blur and data loss during frame readout causing discontinuity of data acquisition as cells move at relatively high speeds through the chamber. We have developed a method to continuously acquire high-resolution images of cells in motion through a microfluidics chamber using a high-speed line scan camera. The sensor acquires images in a line-by-line fashion in order to continuously image moving objects without motion blur. The optical setup comprises an epi-illuminated microscope with a 40X oil immersion, 1.4 NA objective and a 150 mm tube lens focused on a microfluidic channel. Samples containing suspended cells fluorescently stained with 0.01% (w/v) proflavine in saline are introduced into the microfluidics chamber via a syringe pump; illumination is provided by a blue LED (455 nm). Images were taken of samples at the focal plane using an ELiiXA+ 8k/4k monochrome line-scan camera at a line rate of up to 40 kHz. The system's line rate and fluid velocity are tightly controlled to reduce image distortion and are validated using fluorescent microspheres. Image acquisition was controlled via MATLAB's Image Acquisition toolbox. Data sets comprise discrete images of every detectable cell which may be subsequently mined for morphological statistics and definable features by a custom texture analysis algorithm. This high-throughput screening method, comparable to cell counting by flow cytometry, provided efficient examination including counting, classification, and differentiation of saliva, blood, and cultured human cancer cells.

Advanced imaging techniques for the study of plant growth and development.

PubMed

Sozzani, Rosangela; Busch, Wolfgang; Spalding, Edgar P; Benfey, Philip N

2014-05-01

A variety of imaging methodologies are being used to collect data for quantitative studies of plant growth and development from living plants. Multi-level data, from macroscopic to molecular, and from weeks to seconds, can be acquired. Furthermore, advances in parallelized and automated image acquisition enable the throughput to capture images from large populations of plants under specific growth conditions. Image-processing capabilities allow for 3D or 4D reconstruction of image data and automated quantification of biological features. These advances facilitate the integration of imaging data with genome-wide molecular data to enable systems-level modeling. Copyright © 2013 Elsevier Ltd. All rights reserved.

High throughput integrated thermal characterization with non-contact optical calorimetry

NASA Astrophysics Data System (ADS)

Hou, Sichao; Huo, Ruiqing; Su, Ming

2017-10-01

Commonly used thermal analysis tools such as calorimeter and thermal conductivity meter are separated instruments and limited by low throughput, where only one sample is examined each time. This work reports an infrared based optical calorimetry with its theoretical foundation, which is able to provide an integrated solution to characterize thermal properties of materials with high throughput. By taking time domain temperature information of spatially distributed samples, this method allows a single device (infrared camera) to determine the thermal properties of both phase change systems (melting temperature and latent heat of fusion) and non-phase change systems (thermal conductivity and heat capacity). This method further allows these thermal properties of multiple samples to be determined rapidly, remotely, and simultaneously. In this proof-of-concept experiment, the thermal properties of a panel of 16 samples including melting temperatures, latent heats of fusion, heat capacities, and thermal conductivities have been determined in 2 min with high accuracy. Given the high thermal, spatial, and temporal resolutions of the advanced infrared camera, this method has the potential to revolutionize the thermal characterization of materials by providing an integrated solution with high throughput, high sensitivity, and short analysis time.

An Image-Based Algorithm for Precise and Accurate High Throughput Assessment of Drug Activity against the Human Parasite Trypanosoma cruzi

PubMed Central

Moraes, Carolina Borsoi; Yang, Gyongseon; Kang, Myungjoo; Freitas-Junior, Lucio H.; Hansen, Michael A. E.

2014-01-01

We present a customized high content (image-based) and high throughput screening algorithm for the quantification of Trypanosoma cruzi infection in host cells. Based solely on DNA staining and single-channel images, the algorithm precisely segments and identifies the nuclei and cytoplasm of mammalian host cells as well as the intracellular parasites infecting the cells. The algorithm outputs statistical parameters including the total number of cells, number of infected cells and the total number of parasites per image, the average number of parasites per infected cell, and the infection ratio (defined as the number of infected cells divided by the total number of cells). Accurate and precise estimation of these parameters allow for both quantification of compound activity against parasites, as well as the compound cytotoxicity, thus eliminating the need for an additional toxicity-assay, hereby reducing screening costs significantly. We validate the performance of the algorithm using two known drugs against T.cruzi: Benznidazole and Nifurtimox. Also, we have checked the performance of the cell detection with manual inspection of the images. Finally, from the titration of the two compounds, we confirm that the algorithm provides the expected half maximal effective concentration (EC50) of the anti-T. cruzi activity. PMID:24503652

An Unsupervised kNN Method to Systematically Detect Changes in Protein Localization in High-Throughput Microscopy Images.

PubMed

Lu, Alex Xijie; Moses, Alan M

2016-01-01

Despite the importance of characterizing genes that exhibit subcellular localization changes between conditions in proteome-wide imaging experiments, many recent studies still rely upon manual evaluation to assess the results of high-throughput imaging experiments. We describe and demonstrate an unsupervised k-nearest neighbours method for the detection of localization changes. Compared to previous classification-based supervised change detection methods, our method is much simpler and faster, and operates directly on the feature space to overcome limitations in needing to manually curate training sets that may not generalize well between screens. In addition, the output of our method is flexible in its utility, generating both a quantitatively ranked list of localization changes that permit user-defined cut-offs, and a vector for each gene describing feature-wise direction and magnitude of localization changes. We demonstrate that our method is effective at the detection of localization changes using the Δrpd3 perturbation in Saccharomyces cerevisiae, where we capture 71.4% of previously known changes within the top 10% of ranked genes, and find at least four new localization changes within the top 1% of ranked genes. The results of our analysis indicate that simple unsupervised methods may be able to identify localization changes in images without laborious manual image labelling steps.

High throughput operando studies using Fourier transform infrared imaging and Raman spectroscopy.

PubMed

Li, Guosheng; Hu, Dehong; Xia, Guanguang; White, J M; Zhang, Conrad

2008-07-01

A prototype high throughput operando (HTO) reactor designed and built for catalyst screening and characterization combines Fourier transform infrared (FT-IR) imaging and Raman spectroscopy in operando conditions. Using a focal plane array detector (HgCdTe focal plane array, 128x128 pixels, and 1610 Hz frame rate) for the FT-IR imaging system, the catalyst activity and selectivity of all parallel reaction channels can be simultaneously followed. Each image data set possesses 16 384 IR spectra with a spectral range of 800-4000 cm(-1) and with an 8 cm(-1) resolution. Depending on the signal-to-noise ratio, 2-20 s are needed to generate a full image of all reaction channels for a data set. Results on reactant conversion and product selectivity are obtained from FT-IR spectral analysis. Six novel Raman probes, one for each reaction channel, were specially designed and house built at Pacific Northwest National Laboratory, to simultaneously collect Raman spectra of the catalysts and possible reaction intermediates on the catalyst surface under operando conditions. As a model system, methanol partial oxidation reaction on silica-supported molybdenum oxide (MoO3SiO2) catalysts has been studied under different reaction conditions to demonstrate the performance of the HTO reactor.

Interleaved diffusion-weighted EPI improved by adaptive partial-Fourier and multi-band multiplexed sensitivity-encoding reconstruction

PubMed Central

Chang, Hing-Chiu; Guhaniyogi, Shayan; Chen, Nan-kuei

2014-01-01

Purpose We report a series of techniques to reliably eliminate artifacts in interleaved echo-planar imaging (EPI) based diffusion weighted imaging (DWI). Methods First, we integrate the previously reported multiplexed sensitivity encoding (MUSE) algorithm with a new adaptive Homodyne partial-Fourier reconstruction algorithm, so that images reconstructed from interleaved partial-Fourier DWI data are free from artifacts even in the presence of either a) motion-induced k-space energy peak displacement, or b) susceptibility field gradient induced fast phase changes. Second, we generalize the previously reported single-band MUSE framework to multi-band MUSE, so that both through-plane and in-plane aliasing artifacts in multi-band multi-shot interleaved DWI data can be effectively eliminated. Results The new adaptive Homodyne-MUSE reconstruction algorithm reliably produces high-quality and high-resolution DWI, eliminating residual artifacts in images reconstructed with previously reported methods. Furthermore, the generalized MUSE algorithm is compatible with multi-band and high-throughput DWI. Conclusion The integration of the multi-band and adaptive Homodyne-MUSE algorithms significantly improves the spatial-resolution, image quality, and scan throughput of interleaved DWI. We expect that the reported reconstruction framework will play an important role in enabling high-resolution DWI for both neuroscience research and clinical uses. PMID:24925000

High-Throughput Toxicokinetics (HTTK) R package (CompTox CoP presentation)

EPA Science Inventory

Toxicokinetics (TK) provides a bridge between HTS and HTE by predicting tissue concentrations due to exposure, but traditional TK methods are resource intensive. Relatively high throughput TK (HTTK) methods have been used by the pharmaceutical industry to determine range of effic...

Development and application of Fourier-transform infrared chemical imaging of tumour in human tissue.

PubMed

Petter, C H; Heigl, N; Rainer, M; Bakry, R; Pallua, J; Bonn, G K; Huck, C W

2009-01-01

Fourier-transform infrared (FT-IR) based mapping and imaging is a fast emerging technology which is being increasingly applied to investigate tissues in the high-throughput mode. The high resolution close to the cellular level, the possibility to determine the bio-distribution of molecules of interest (proteins, peptides, lipids, carbohydrates) without any pre-treatment and the offer to yield molecular structure information have brought evidence that this technique allows to gain new insights in cancer pathology. Thus, several individual mainly protein and peptide cancer markers ("biomarkers") can be identified from FT-IR tissue images, enabling accurate discrimination between healthy and tumour areas. Optimal data acquisition (spatial resolution, spectral resolution, signal to noise ratio), classification, and validation are necessary to establish practical protocols that can be translated to the qualitative and quantitative clinical routine analysis. Thereby, the development of modern fast infrared imaging systems has strongly supported its acceptance in clinical histopathology. In this review, the necessity of analysis based on global cancer statistics, instrumental setups and developments, experimental state of the art are summarised and applications to investigate different kinds of cancer (e.g., prostate, breast, cervical, colon, oral cavity) are shown and discussed in detail.

Investigating the Use of Cloudbursts for High-Throughput Medical Image Registration

PubMed Central

Kim, Hyunjoo; Parashar, Manish; Foran, David J.; Yang, Lin

2010-01-01

This paper investigates the use of clouds and autonomic cloudbursting to support a medical image registration. The goal is to enable a virtual computational cloud that integrates local computational environments and public cloud services on-the-fly, and support image registration requests from different distributed researcher groups with varied computational requirements and QoS constraints. The virtual cloud essentially implements shared and coordinated task-spaces, which coordinates the scheduling of jobs submitted by a dynamic set of research groups to their local job queues. A policy-driven scheduling agent uses the QoS constraints along with performance history and the state of the resources to determine the appropriate size and mix of the public and private cloud resource that should be allocated to a specific request. The virtual computational cloud and the medical image registration service have been developed using the CometCloud engine and have been deployed on a combination of private clouds at Rutgers University and the Cancer Institute of New Jersey and Amazon EC2. An experimental evaluation is presented and demonstrates the effectiveness of autonomic cloudbursts and policy-based autonomic scheduling for this application. PMID:20640235

Optimization of High-Throughput Sequencing Kinetics for determining enzymatic rate constants of thousands of RNA substrates

PubMed Central

Niland, Courtney N.; Jankowsky, Eckhard; Harris, Michael E.

2016-01-01

Quantification of the specificity of RNA binding proteins and RNA processing enzymes is essential to understanding their fundamental roles in biological processes. High Throughput Sequencing Kinetics (HTS-Kin) uses high throughput sequencing and internal competition kinetics to simultaneously monitor the processing rate constants of thousands of substrates by RNA processing enzymes. This technique has provided unprecedented insight into the substrate specificity of the tRNA processing endonuclease ribonuclease P. Here, we investigate the accuracy and robustness of measurements associated with each step of the HTS-Kin procedure. We examine the effect of substrate concentration on the observed rate constant, determine the optimal kinetic parameters, and provide guidelines for reducing error in amplification of the substrate population. Importantly, we find that high-throughput sequencing, and experimental reproducibility contribute their own sources of error, and these are the main sources of imprecision in the quantified results when otherwise optimized guidelines are followed. PMID:27296633

High-throughput determination of structural phase diagram and constituent phases using GRENDEL

NASA Astrophysics Data System (ADS)

Kusne, A. G.; Keller, D.; Anderson, A.; Zaban, A.; Takeuchi, I.

2015-11-01

Advances in high-throughput materials fabrication and characterization techniques have resulted in faster rates of data collection and rapidly growing volumes of experimental data. To convert this mass of information into actionable knowledge of material process-structure-property relationships requires high-throughput data analysis techniques. This work explores the use of the Graph-based endmember extraction and labeling (GRENDEL) algorithm as a high-throughput method for analyzing structural data from combinatorial libraries, specifically, to determine phase diagrams and constituent phases from both x-ray diffraction and Raman spectral data. The GRENDEL algorithm utilizes a set of physical constraints to optimize results and provides a framework by which additional physics-based constraints can be easily incorporated. GRENDEL also permits the integration of database data as shown by the use of critically evaluated data from the Inorganic Crystal Structure Database in the x-ray diffraction data analysis. Also the Sunburst radial tree map is demonstrated as a tool to visualize material structure-property relationships found through graph based analysis.

Two-dimensional Mathematical Model of Oil-bearing Materials in Extrusion-type Transportation over Rectangular Screw Core

NASA Astrophysics Data System (ADS)

Gukasyan, A. V.; Koshevoy, E. P.; Kosachev, V. S.

2018-05-01

A comparative analysis of alternative models for plastic flow in extrusive transportation of oil-bearing materials was conducted; the research was directed at determining the function describing the screw core throughput capacity of the press (extruder). Transition from a one-dimensional model to a two-dimensional model significantly improves the mathematical model and allows using two-dimensional rheological models determining the throughput of the screw core.

Machine vision for digital microfluidics

NASA Astrophysics Data System (ADS)

Shin, Yong-Jun; Lee, Jeong-Bong

2010-01-01

Machine vision is widely used in an industrial environment today. It can perform various tasks, such as inspecting and controlling production processes, that may require humanlike intelligence. The importance of imaging technology for biological research or medical diagnosis is greater than ever. For example, fluorescent reporter imaging enables scientists to study the dynamics of gene networks with high spatial and temporal resolution. Such high-throughput imaging is increasingly demanding the use of machine vision for real-time analysis and control. Digital microfluidics is a relatively new technology with expectations of becoming a true lab-on-a-chip platform. Utilizing digital microfluidics, only small amounts of biological samples are required and the experimental procedures can be automatically controlled. There is a strong need for the development of a digital microfluidics system integrated with machine vision for innovative biological research today. In this paper, we show how machine vision can be applied to digital microfluidics by demonstrating two applications: machine vision-based measurement of the kinetics of biomolecular interactions and machine vision-based droplet motion control. It is expected that digital microfluidics-based machine vision system will add intelligence and automation to high-throughput biological imaging in the future.

Multiplex Staining by Sequential Immunostaining and Antibody Removal on Routine Tissue Sections.

PubMed

Bolognesi, Maddalena Maria; Manzoni, Marco; Scalia, Carla Rossana; Zannella, Stefano; Bosisio, Francesca Maria; Faretta, Mario; Cattoretti, Giorgio

2017-08-01

Multiplexing, labeling for multiple immunostains in the very same cell or tissue section in situ, has raised considerable interest. The methods proposed include the use of labeled primary antibodies, spectral separation of fluorochromes, bleaching of the fluorophores or chromogens, blocking of previous antibody layers, all in various combinations. The major obstacles to the diffusion of this technique are high costs in custom antibodies and instruments, low throughput, and scarcity of specialized skills or facilities. We have validated a method based on common primary and secondary antibodies and diffusely available fluorescent image scanners. It entails rounds of four-color indirect immunofluorescence, image acquisition, and removal (stripping) of the antibodies, before another stain is applied. The images are digitally registered and the autofluorescence is subtracted. Removal of antibodies is accomplished by disulfide cleavage and a detergent or by a chaotropic salt treatment, this latter followed by antigen refolding. More than 30 different antibody stains can be applied to one single section from routinely fixed and embedded tissue. This method requires a modest investment in hardware and materials and uses freeware image analysis software. Multiplexing on routine tissue sections is a high throughput tool for in situ characterization of neoplastic, reactive, inflammatory, and normal cells.

Massively Parallel Rogue Cell Detection Using Serial Time-Encoded Amplified Microscopy of Inertially Ordered Cells in High-Throughput Flow

DTIC Science & Technology

2012-08-01

techniques and STEAM imager. It couples the high-speed capability of the STEAM imager and differential phase contrast imaging of DIC / Nomarski microscopy...On 10 TPE chips, we obtained 9 homogenous and strong bonds, the failed bond being due to operator error and presence of air bubbles in the TPE...instruments, structural dynamics, and microelectromechanical systems (MEMS) via laser-scanning surface vibrometry , and observation of biomechanical motility

Advances in Automated Plankton Imaging: Enhanced Throughput, Automated Staining, and Extended Deployment Modes for Imaging FlowCytobot

NASA Astrophysics Data System (ADS)

Sosik, H. M.; Olson, R. J.; Brownlee, E.; Brosnahan, M.; Crockford, E. T.; Peacock, E.; Shalapyonok, A.

2016-12-01

Imaging FlowCytobot (IFCB) was developed to fill a need for automated identification and monitoring of nano- and microplankton, especially phytoplankton in the size range 10 200 micrometer, which are important in coastal blooms (including harmful algal blooms). IFCB uses a combination of flow cytometric and video technology to capture high resolution (1 micrometer) images of suspended particles. This proven, now commercially available, submersible instrument technology has been deployed in fixed time series locations for extended periods (months to years) and in shipboard laboratories where underway water is automatically analyzed during surveys. Building from these successes, we have now constructed and evaluated three new prototype IFCB designs that extend measurement and deployment capabilities. To improve cell counting statistics without degrading image quality, a high throughput version (IFCB-HT) incorporates in-flow acoustic focusing to non-disruptively pre-concentrate cells before the measurement area of the flow cell. To extend imaging to all heterotrophic cells (even those that do not exhibit chlorophyll fluorescence), Staining IFCB (IFCB-S) incorporates automated addition of a live-cell fluorescent stain (fluorescein diacetate) to samples before analysis. A horizontally-oriented IFCB-AV design addresses the need for spatial surveying from surface autonomous vehicles, including design features that reliably eliminate air bubbles and mitigate wave motion impacts. Laboratory evaluation and test deployments in waters near Woods Hole show the efficacy of each of these enhanced IFCB designs.

Improved spatial resolution of luminescence images acquired with a silicon line scanning camera

NASA Astrophysics Data System (ADS)

Teal, Anthony; Mitchell, Bernhard; Juhl, Mattias K.

2018-04-01

Luminescence imaging is currently being used to provide spatially resolved defect in high volume silicon solar cell production. One option to obtain the high throughput required for on the fly detection is the use a silicon line scan cameras. However, when using a silicon based camera, the spatial resolution is reduced as a result of the weakly absorbed light scattering within the camera's chip. This paper address this issue by applying deconvolution from a measured point spread function. This paper extends the methods for determining the point spread function of a silicon area camera to a line scan camera with charge transfer. The improvement in resolution is quantified in the Fourier domain and in spatial domain on an image of a multicrystalline silicon brick. It is found that light spreading beyond the active sensor area is significant in line scan sensors, but can be corrected for through normalization of the point spread function. The application of this method improves the raw data, allowing effective detection of the spatial resolution of defects in manufacturing.

Falling-incident detection and throughput enhancement in a multi-camera video-surveillance system.

PubMed

Shieh, Wann-Yun; Huang, Ju-Chin

2012-09-01

For most elderly, unpredictable falling incidents may occur at the corner of stairs or a long corridor due to body frailty. If we delay to rescue a falling elder who is likely fainting, more serious consequent injury may occur. Traditional secure or video surveillance systems need caregivers to monitor a centralized screen continuously, or need an elder to wear sensors to detect falling incidents, which explicitly waste much human power or cause inconvenience for elders. In this paper, we propose an automatic falling-detection algorithm and implement this algorithm in a multi-camera video surveillance system. The algorithm uses each camera to fetch the images from the regions required to be monitored. It then uses a falling-pattern recognition algorithm to determine if a falling incident has occurred. If yes, system will send short messages to someone needs to be noticed. The algorithm has been implemented in a DSP-based hardware acceleration board for functionality proof. Simulation results show that the accuracy of falling detection can achieve at least 90% and the throughput of a four-camera surveillance system can be improved by about 2.1 times. Copyright © 2011 IPEM. Published by Elsevier Ltd. All rights reserved.

Resolution and throughput optimized intraoperative spectrally encoded coherence tomography and reflectometry (iSECTR) for multimodal imaging during ophthalmic microsurgery

NASA Astrophysics Data System (ADS)

Malone, Joseph D.; El-Haddad, Mohamed T.; Leeburg, Kelsey C.; Terrones, Benjamin D.; Tao, Yuankai K.

2018-02-01

Limited visualization of semi-transparent structures in the eye remains a critical barrier to improving clinical outcomes and developing novel surgical techniques. While increases in imaging speed has enabled intraoperative optical coherence tomography (iOCT) imaging of surgical dynamics, several critical barriers to clinical adoption remain. Specifically, these include (1) static field-of-views (FOVs) requiring manual instrument-tracking; (2) high frame-rates require sparse sampling, which limits FOV; and (3) small iOCT FOV also limits the ability to co-register data with surgical microscopy. We previously addressed these limitations in image-guided ophthalmic microsurgery by developing microscope-integrated multimodal intraoperative swept-source spectrally encoded scanning laser ophthalmoscopy and optical coherence tomography. Complementary en face images enabled orientation and coregistration with the widefield surgical microscope view while OCT imaging enabled depth-resolved visualization of surgical instrument positions relative to anatomic structures-of-interest. In addition, we demonstrated novel integrated segmentation overlays for augmented-reality surgical guidance. Unfortunately, our previous system lacked the resolution and optical throughput for in vivo retinal imaging and necessitated removal of cornea and lens. These limitations were predominately a result of optical aberrations from imaging through a shared surgical microscope objective lens, which was modeled as a paraxial surface. Here, we present an optimized intraoperative spectrally encoded coherence tomography and reflectometry (iSECTR) system. We use a novel lens characterization method to develop an accurate model of surgical microscope objective performance and balance out inherent aberrations using iSECTR relay optics. Using this system, we demonstrate in vivo multimodal ophthalmic imaging through a surgical microscope

Developing a gene biomarker at the tipping point of adaptive and adverse responses in human bronchial epithelial cells

EPA Science Inventory

Determining mechanism-based biomarkers that distinguish adaptive and adverse cellular processes is critical to understanding the health effects of environmental exposures. Shifting from in vivo, low-throughput toxicity studies to high-throughput screening (HTS) paradigms and risk...

Improved real-time imaging spectrometer

NASA Technical Reports Server (NTRS)

Lambert, James L. (Inventor); Chao, Tien-Hsin (Inventor); Yu, Jeffrey W. (Inventor); Cheng, Li-Jen (Inventor)

1993-01-01

An improved AOTF-based imaging spectrometer that offers several advantages over prior art AOTF imaging spectrometers is presented. The ability to electronically set the bandpass wavelength provides observational flexibility. Various improvements in optical architecture provide simplified magnification variability, improved image resolution and light throughput efficiency and reduced sensitivity to ambient light. Two embodiments of the invention are: (1) operation in the visible/near-infrared domain of wavelength range 0.48 to 0.76 microns; and (2) infrared configuration which operates in the wavelength range of 1.2 to 2.5 microns.

Reducing Speckle In One-Look SAR Images

NASA Technical Reports Server (NTRS)

Nathan, K. S.; Curlander, J. C.

1990-01-01

Local-adaptive-filter algorithm incorporated into digital processing of synthetic-aperture-radar (SAR) echo data to reduce speckle in resulting imagery. Involves use of image statistics in vicinity of each picture element, in conjunction with original intensity of element, to estimate brightness more nearly proportional to true radar reflectance of corresponding target. Increases ratio of signal to speckle noise without substantial degradation of resolution common to multilook SAR images. Adapts to local variations of statistics within scene, preserving subtle details. Computationally simple. Lends itself to parallel processing of different segments of image, making possible increased throughput.

BioImageXD: an open, general-purpose and high-throughput image-processing platform.

PubMed

Kankaanpää, Pasi; Paavolainen, Lassi; Tiitta, Silja; Karjalainen, Mikko; Päivärinne, Joacim; Nieminen, Jonna; Marjomäki, Varpu; Heino, Jyrki; White, Daniel J

2012-06-28

BioImageXD puts open-source computer science tools for three-dimensional visualization and analysis into the hands of all researchers, through a user-friendly graphical interface tuned to the needs of biologists. BioImageXD has no restrictive licenses or undisclosed algorithms and enables publication of precise, reproducible and modifiable workflows. It allows simple construction of processing pipelines and should enable biologists to perform challenging analyses of complex processes. We demonstrate its performance in a study of integrin clustering in response to selected inhibitors.

Performance of the SIR-B digital image processing subsystem

NASA Technical Reports Server (NTRS)

Curlander, J. C.

1986-01-01

A ground-based system to generate digital SAR image products has been developed and implemented in support of the SIR-B mission. This system is designed to achieve the maximum throughput while meeting strict image fidelity criteria. Its capabilities include: automated radiometric and geometric correction of the output imagery; high-precision absolute location without tiepoint registration; filtering of the raw data to remove spurious signals from alien radars; and automated catologing to maintain a full set of radar and image production facility in support of the SIR-B science investigators routinely produces over 80 image frames per week.

Theoretical characterization of annular array as a volumetric optoacoustic ultrasound handheld probe

NASA Astrophysics Data System (ADS)

Kalkhoran, Mohammad Azizian; Vray, Didier

2018-02-01

Optoacoustic ultrasound (OPUS) is a promising hybridized technique for simultaneous acquisition of functional and morphological data. The optical specificity of optoacoustic leverages the diagnostic aptitude of ultrasonography beyond anatomy. However, this integration has been rarely practiced for volumetric imaging. The challenge lies in the effective imaging probes that preserve the functionality of both modalities. The potentials of a sparse annular array for volumetric OPUS imaging are theoretically investigated. In order to evaluate and optimize the performance characteristics of the probe, series of analysis in the framework of system model matrix was carried out. The two criteria of voxel crosstalk and eigenanalysis have been employed to unveil information about the spatial sensitivity, aliasing, and number of definable spatial frequency components. Based on these benchmarks, the optimal parameters for volumetric handheld probe are determined. In particular, the number, size, and the arrangement of the elements and overall aperture dimension were investigated. The result of the numerical simulation suggests that the segmented-annular array of 128 negatively focused elements with 1λ × 20λ size, operating at 5-MHz central frequency showcases a good agreement with the physical requirement of both imaging systems. We hypothesize that these features enable a high-throughput volumetric passive/active ultrasonic imaging system with great potential for clinical applications.

Use of Optical Imaging Technology in the Validation of a New, Rapid, Cost-Effective Drug Screen as Part of a Tiered In Vivo Screening Paradigm for Development of Drugs To Treat Cutaneous Leishmaniasis

PubMed Central

Parriot, Sandi; Hudson, Thomas H.; Lang, Thierry; Ngundam, Franklyn; Leed, Susan; Sena, Jenell; Harris, Michael; O'Neil, Michael; Sciotti, Richard; Read, Lisa; Lecoeur, Herve; Grogl, Max

2017-01-01

ABSTRACT In any drug discovery and development effort, a reduction in the time of the lead optimization cycle is critical to decrease the time to license and reduce costs. In addition, ethical guidelines call for the more ethical use of animals to minimize the number of animals used and decrease their suffering. Therefore, any effort to develop drugs to treat cutaneous leishmaniasis requires multiple tiers of in vivo testing that start with higher-throughput efficacy assessments and progress to lower-throughput models with the most clinical relevance. Here, we describe the validation of a high-throughput, first-tier, noninvasive model of lesion suppression that uses an in vivo optical imaging technology for the initial screening of compounds. A strong correlation between luciferase activity and the parasite load at up to 18 days postinfection was found. This correlation allows the direct assessment of the effects of drug treatment on parasite burden. We demonstrate that there is a strong correlation between drug efficacy measured on day 18 postinfection and the suppression of lesion size by day 60 postinfection, which allows us to reach an accurate conclusion on drug efficacy in only 18 days. Compounds demonstrating a significant reduction in the bioluminescence signal compared to that in control animals can be tested in lower-throughput, more definitive tests of lesion cure in BALB/c mice and Golden Syrian hamsters (GSH) using Old World and New World parasites. PMID:28137819

Geospatial analytics to evaluate point-of-dispensing sites for mass immunizations in Allegheny County, Pennsylvania.

PubMed

Everett, Kibri H; Potter, Margaret A; Wheaton, William D; Gleason, Sherrianne M; Brown, Shawn T; Lee, Bruce Y

2013-01-01

Public health agencies use mass immunization locations to quickly administer vaccines to protect a population against an epidemic. The selection of such locations is frequently determined by available staffing levels and in some places, not all potential sites can be opened, often because of a lack of resources. Public health agencies need assistance in determining which n sites are the prime ones to open given available staff to minimize travel time and travel distance for those in the population who need to get to a site to receive treatment. Employ geospatial analytical methods to identify the prime n locations from a predetermined set of potential locations (eg, schools) and determine which locations may not be able to achieve the throughput necessary to reach the herd immunity threshold based on varying R0 values. Spatial location-allocation algorithms were used to select the ideal n mass vaccination locations. Allegheny County, Pennsylvania, served as the study area. The most favorable sites were selected and the number of individuals required to be vaccinated to achieve the herd immunity threshold for a given R0, ranging from 1.5 to 7, was determined. Locations that did not meet the Centers for Disease Control and Prevention throughput recommendation for smallpox were identified. At R0 = 1.5, all mass immunization locations met the required throughput to achieve the herd immunity threshold within 5 days. As R0s increased from 2 to 7, an increasing number of sites were inadequate to meet throughput requirements. Identifying the top n sites and categorizing those with throughput challenges allows health departments to adjust staffing, shift length, or the number of sites. This method has the potential to be expanded to select immunization locations under a number of additional scenarios.

Computational imaging of sperm locomotion.

PubMed

Daloglu, Mustafa Ugur; Ozcan, Aydogan

2017-08-01

Not only essential for scientific research, but also in the analysis of male fertility and for animal husbandry, sperm tracking and characterization techniques have been greatly benefiting from computational imaging. Digital image sensors, in combination with optical microscopy tools and powerful computers, have enabled the use of advanced detection and tracking algorithms that automatically map sperm trajectories and calculate various motility parameters across large data sets. Computational techniques are driving the field even further, facilitating the development of unconventional sperm imaging and tracking methods that do not rely on standard optical microscopes and objective lenses, which limit the field of view and volume of the semen sample that can be imaged. As an example, a holographic on-chip sperm imaging platform, only composed of a light-emitting diode and an opto-electronic image sensor, has emerged as a high-throughput, low-cost and portable alternative to lens-based traditional sperm imaging and tracking methods. In this approach, the sample is placed very close to the image sensor chip, which captures lensfree holograms generated by the interference of the background illumination with the light scattered from sperm cells. These holographic patterns are then digitally processed to extract both the amplitude and phase information of the spermatozoa, effectively replacing the microscope objective lens with computation. This platform has further enabled high-throughput 3D imaging of spermatozoa with submicron 3D positioning accuracy in large sample volumes, revealing various rare locomotion patterns. We believe that computational chip-scale sperm imaging and 3D tracking techniques will find numerous opportunities in both sperm related research and commercial applications. © The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Rapid analysis and exploration of fluorescence microscopy images.

PubMed

Pavie, Benjamin; Rajaram, Satwik; Ouyang, Austin; Altschuler, Jason M; Steininger, Robert J; Wu, Lani F; Altschuler, Steven J

2014-03-19

Despite rapid advances in high-throughput microscopy, quantitative image-based assays still pose significant challenges. While a variety of specialized image analysis tools are available, most traditional image-analysis-based workflows have steep learning curves (for fine tuning of analysis parameters) and result in long turnaround times between imaging and analysis. In particular, cell segmentation, the process of identifying individual cells in an image, is a major bottleneck in this regard. Here we present an alternate, cell-segmentation-free workflow based on PhenoRipper, an open-source software platform designed for the rapid analysis and exploration of microscopy images. The pipeline presented here is optimized for immunofluorescence microscopy images of cell cultures and requires minimal user intervention. Within half an hour, PhenoRipper can analyze data from a typical 96-well experiment and generate image profiles. Users can then visually explore their data, perform quality control on their experiment, ensure response to perturbations and check reproducibility of replicates. This facilitates a rapid feedback cycle between analysis and experiment, which is crucial during assay optimization. This protocol is useful not just as a first pass analysis for quality control, but also may be used as an end-to-end solution, especially for screening. The workflow described here scales to large data sets such as those generated by high-throughput screens, and has been shown to group experimental conditions by phenotype accurately over a wide range of biological systems. The PhenoBrowser interface provides an intuitive framework to explore the phenotypic space and relate image properties to biological annotations. Taken together, the protocol described here will lower the barriers to adopting quantitative analysis of image based screens.

Plant phenomics: an overview of image acquisition technologies and image data analysis algorithms

PubMed Central

Perez-Sanz, Fernando; Navarro, Pedro J

2017-01-01

Abstract The study of phenomes or phenomics has been a central part of biology. The field of automatic phenotype acquisition technologies based on images has seen an important advance in the last years. As with other high-throughput technologies, it addresses a common set of problems, including data acquisition and analysis. In this review, we give an overview of the main systems developed to acquire images. We give an in-depth analysis of image processing with its major issues and the algorithms that are being used or emerging as useful to obtain data out of images in an automatic fashion. PMID:29048559

Development of a diffraction imaging flow cytometer

PubMed Central

Jacobs, Kenneth M.; Lu, Jun Q.

2013-01-01

Diffraction images record angle-resolved distribution of scattered light from a particle excited by coherent light and can correlate highly with the 3D morphology of a particle. We present a jet-in-fluid design of flow chamber for acquisition of clear diffraction images in a laminar flow. Diffraction images of polystyrene spheres of different diameters were acquired and found to correlate highly with the calculated ones based on the Mie theory. Fast Fourier transform analysis indicated that the measured images can be used to extract sphere diameter values. These results demonstrate the significant potentials of high-throughput diffraction imaging flow cytometry for extracting 3D morphological features of cells. PMID:19794790

Adverse outcome pathway (AOP) development II: Best practices

EPA Science Inventory

Organization of existing and emerging toxicological knowledge into adverse outcome pathway (AOP) descriptions can facilitate greater application of mechanistic data, including high throughput in vitro, high content omics and imaging, and biomarkers, in risk-based decision-making....

Integrated, multi-scale, spatial-temporal cell biology--A next step in the post genomic era.

PubMed

Horwitz, Rick

2016-03-01

New microscopic approaches, high-throughput imaging, and gene editing promise major new insights into cellular behaviors. When coupled with genomic and other 'omic information and "mined" for correlations and associations, a new breed of powerful and useful cellular models should emerge. These top down, coarse-grained, and statistical models, in turn, can be used to form hypotheses merging with fine-grained, bottom up mechanistic studies and models that are the back bone of cell biology. The goal of the Allen Institute for Cell Science is to develop the top down approach by developing a high throughput microscopy pipeline that is integrated with modeling, using gene edited hiPS cell lines in various physiological and pathological contexts. The output of these experiments and models will be an "animated" cell, capable of integrating and analyzing image data generated from experiments and models. Copyright © 2015 Elsevier Inc. All rights reserved.

Hazardous waste incinerators under waste uncertainty: balancing and throughput maximization via heat recuperation.

PubMed

Tsiliyannis, Christos Aristeides

2013-09-01

Hazardous waste incinerators (HWIs) differ substantially from thermal power facilities, since instead of maximizing energy production with the minimum amount of fuel, they aim at maximizing throughput. Variations in quantity or composition of received waste loads may significantly diminish HWI throughput (the decisive profit factor), from its nominal design value. A novel formulation of combustion balance is presented, based on linear operators, which isolates the wastefeed vector from the invariant combustion stoichiometry kernel. Explicit expressions for the throughput are obtained, in terms of incinerator temperature, fluegas heat recuperation ratio and design parameters, for an arbitrary number of wastes, based on fundamental principles (mass and enthalpy balances). The impact of waste variations, of recuperation ratio and of furnace temperature is explicitly determined. It is shown that in the presence of waste uncertainty, the throughput may be a decreasing or increasing function of incinerator temperature and recuperation ratio, depending on the sign of a dimensionless parameter related only to the uncertain wastes. The dimensionless parameter is proposed as a sharp a' priori waste 'fingerprint', determining the necessary increase or decrease of manipulated variables (recuperation ratio, excess air, auxiliary fuel feed rate, auxiliary air flow) in order to balance the HWI and maximize throughput under uncertainty in received wastes. A 10-step procedure is proposed for direct application subject to process capacity constraints. The results may be useful for efficient HWI operation and for preparing hazardous waste blends. Copyright © 2013 Elsevier Ltd. All rights reserved.

FIM, a Novel FTIR-Based Imaging Method for High Throughput Locomotion Analysis

PubMed Central

Otto, Nils; Löpmeier, Tim; Valkov, Dimitar; Jiang, Xiaoyi; Klämbt, Christian

2013-01-01

We designed a novel imaging technique based on frustrated total internal reflection (FTIR) to obtain high resolution and high contrast movies. This FTIR-based Imaging Method (FIM) is suitable for a wide range of biological applications and a wide range of organisms. It operates at all wavelengths permitting the in vivo detection of fluorescent proteins. To demonstrate the benefits of FIM, we analyzed large groups of crawling Drosophila larvae. The number of analyzable locomotion tracks was increased by implementing a new software module capable of preserving larval identity during most collision events. This module is integrated in our new tracking program named FIMTrack which subsequently extracts a number of features required for the analysis of complex locomotion phenotypes. FIM enables high throughput screening for even subtle behavioral phenotypes. We tested this newly developed setup by analyzing locomotion deficits caused by the glial knockdown of several genes. Suppression of kinesin heavy chain (khc) or rab30 function led to contraction pattern or head sweeping defects, which escaped in previous analysis. Thus, FIM permits forward genetic screens aimed to unravel the neural basis of behavior. PMID:23349775

CellProfiler Tracer: exploring and validating high-throughput, time-lapse microscopy image data.

PubMed

Bray, Mark-Anthony; Carpenter, Anne E

2015-11-04

Time-lapse analysis of cellular images is an important and growing need in biology. Algorithms for cell tracking are widely available; what researchers have been missing is a single open-source software package to visualize standard tracking output (from software like CellProfiler) in a way that allows convenient assessment of track quality, especially for researchers tuning tracking parameters for high-content time-lapse experiments. This makes quality assessment and algorithm adjustment a substantial challenge, particularly when dealing with hundreds of time-lapse movies collected in a high-throughput manner. We present CellProfiler Tracer, a free and open-source tool that complements the object tracking functionality of the CellProfiler biological image analysis package. Tracer allows multi-parametric morphological data to be visualized on object tracks, providing visualizations that have already been validated within the scientific community for time-lapse experiments, and combining them with simple graph-based measures for highlighting possible tracking artifacts. CellProfiler Tracer is a useful, free tool for inspection and quality control of object tracking data, available from http://www.cellprofiler.org/tracer/.

Automated image-based assay for evaluation of HIV neutralization and cell-to-cell fusion inhibition.

PubMed

Sheik-Khalil, Enas; Bray, Mark-Anthony; Özkaya Şahin, Gülsen; Scarlatti, Gabriella; Jansson, Marianne; Carpenter, Anne E; Fenyö, Eva Maria

2014-08-30

Standardized techniques to detect HIV-neutralizing antibody responses are of great importance in the search for an HIV vaccine. Here, we present a high-throughput, high-content automated plaque reduction (APR) assay based on automated microscopy and image analysis that allows evaluation of neutralization and inhibition of cell-cell fusion within the same assay. Neutralization of virus particles is measured as a reduction in the number of fluorescent plaques, and inhibition of cell-cell fusion as a reduction in plaque area. We found neutralization strength to be a significant factor in the ability of virus to form syncytia. Further, we introduce the inhibitory concentration of plaque area reduction (ICpar) as an additional measure of antiviral activity, i.e. fusion inhibition. We present an automated image based high-throughput, high-content HIV plaque reduction assay. This allows, for the first time, simultaneous evaluation of neutralization and inhibition of cell-cell fusion within the same assay, by quantifying the reduction in number of plaques and mean plaque area, respectively. Inhibition of cell-to-cell fusion requires higher quantities of inhibitory reagent than inhibition of virus neutralization.

Phase imaging of mechanical properties of live cells (Conference Presentation)

NASA Astrophysics Data System (ADS)

Wax, Adam

2017-02-01

The mechanisms by which cells respond to mechanical stimuli are essential for cell function yet not well understood. Many rheological tools have been developed to characterize cellular viscoelastic properties but these typically require direct mechanical contact, limiting their throughput. We have developed a new approach for characterizing the organization of subcellular structures using a label free, noncontact, single-shot phase imaging method that correlates to measured cellular mechanical stiffness. The new analysis approach measures refractive index variance and relates it to disorder strength. These measurements are compared to cellular stiffness, measured using the same imaging tool to visualize nanoscale responses to flow shear stimulus. The utility of the technique is shown by comparing shear stiffness and phase disorder strength across five cellular populations with varying mechanical properties. An inverse relationship between disorder strength and shear stiffness is shown, suggesting that cell mechanical properties can be assessed in a format amenable to high throughput studies using this novel, non-contact technique. Further studies will be presented which include examination of mechanical stiffness in early carcinogenic events and investigation of the role of specific cellular structural proteins in mechanotransduction.

Imaging flow cytometry for the screening of compounds that disrupt the Plasmodium falciparum digestive vacuole.

PubMed

Chia, Wan Ni; Lee, Yan Quan; Tan, Kevin Shyong-Wei

2017-01-01

Malaria, despite being one of the world's oldest infectious diseases, remains difficult to eradicate because the parasite is rapidly developing resistance to frontline chemotherapies. Previous studies have shown that the parasite exhibits features resembling programmed cell death upon treatment with drugs that disrupt its digestive vacuole (DV), providing a phenotypic readout that can be detected using the imaging flow cytometer. Large compound collections can thus be screened to identify drugs that are able to disrupt the DV of the malaria parasite using this high-content high-throughput screening platform. As a proof-of-concept, 4440 compounds were screened using this platform in 4months and 254 hits (5.7% hit rate) were obtained. Additionally, 25 compounds (0.6% top hit rate) were observed to retain potent DV disruption activity that was comparable to the canonical DV disruptive drug chloroquine when tested at a ten-fold lower concentration from the original screen. This pilot study demonstrates the robustness and high-throughput capability of the imaging flow cytometer and we report herein the methodology of this screening assay. Copyright © 2016 Elsevier Inc. All rights reserved.

Real-time image processing for label-free enrichment of Actinobacteria cultivated in picolitre droplets.

PubMed

Zang, Emerson; Brandes, Susanne; Tovar, Miguel; Martin, Karin; Mech, Franziska; Horbert, Peter; Henkel, Thomas; Figge, Marc Thilo; Roth, Martin

2013-09-21

The majority of today's antimicrobial therapeutics is derived from secondary metabolites produced by Actinobacteria. While it is generally assumed that less than 1% of Actinobacteria species from soil habitats have been cultivated so far, classic screening approaches fail to supply new substances, often due to limited throughput and frequent rediscovery of already known strains. To overcome these restrictions, we implement high-throughput cultivation of soil-derived Actinobacteria in microfluidic pL-droplets by generating more than 600,000 pure cultures per hour from a spore suspension that can subsequently be incubated for days to weeks. Moreover, we introduce triggered imaging with real-time image-based droplet classification as a novel universal method for pL-droplet sorting. Growth-dependent droplet sorting at frequencies above 100 Hz is performed for label-free enrichment and extraction of microcultures. The combination of both cultivation of Actinobacteria in pL-droplets and real-time detection of growing Actinobacteria has great potential in screening for yet unknown species as well as their undiscovered natural products.

A probabilistic approach to joint cell tracking and segmentation in high-throughput microscopy videos.

PubMed

Arbelle, Assaf; Reyes, Jose; Chen, Jia-Yun; Lahav, Galit; Riklin Raviv, Tammy

2018-04-22

We present a novel computational framework for the analysis of high-throughput microscopy videos of living cells. The proposed framework is generally useful and can be applied to different datasets acquired in a variety of laboratory settings. This is accomplished by tying together two fundamental aspects of cell lineage construction, namely cell segmentation and tracking, via a Bayesian inference of dynamic models. In contrast to most existing approaches, which aim to be general, no assumption of cell shape is made. Spatial, temporal, and cross-sectional variation of the analysed data are accommodated by two key contributions. First, time series analysis is exploited to estimate the temporal cell shape uncertainty in addition to cell trajectory. Second, a fast marching (FM) algorithm is used to integrate the inferred cell properties with the observed image measurements in order to obtain image likelihood for cell segmentation, and association. The proposed approach has been tested on eight different time-lapse microscopy data sets, some of which are high-throughput, demonstrating promising results for the detection, segmentation and association of planar cells. Our results surpass the state of the art for the Fluo-C2DL-MSC data set of the Cell Tracking Challenge (Maška et al., 2014). Copyright © 2018 Elsevier B.V. All rights reserved.

Development and evaluation of a novel high-throughput image-based fluorescent neutralization test for detection of Zika virus infection.

PubMed

Koishi, Andrea Cristine; Suzukawa, Andréia Akemi; Zanluca, Camila; Camacho, Daria Elena; Comach, Guillermo; Duarte Dos Santos, Claudia Nunes

2018-03-01

Zika virus (ZIKV) is an emerging arbovirus belonging to the genus flavivirus that comprises other important public health viruses, such as dengue (DENV) and yellow fever (YFV). In general, ZIKV infection is a self-limiting disease, however cases of Guillain-Barré syndrome and congenital brain abnormalities in newborn infants have been reported. Diagnosing ZIKV infection remains a challenge, as viral RNA detection is only applicable until a few days after the onset of symptoms. After that, serological tests must be applied, and, as expected, high cross-reactivity between ZIKV and other flavivirus serology is observed. Plaque reduction neutralization test (PRNT) is indicated to confirm positive samples for being more specific, however it is laborious intensive and time consuming, representing a major bottleneck for patient diagnosis. To overcome this limitation, we developed a high-throughput image-based fluorescent neutralization test for ZIKV infection by serological detection. Using 226 human specimens, we showed that the new test presented higher throughput than traditional PRNT, maintaining the correlation between results. Furthermore, when tested with dengue virus samples, it showed 50.53% less cross reactivity than MAC-ELISA. This fluorescent neutralization test could be used for clinical diagnosis confirmation of ZIKV infection, as well as for vaccine clinical trials and seroprevalence studies.

CCR researchers identify pathway critical for preventing premature aging | Center for Cancer Research

Cancer.gov

Hutchinson-Gilford progeria syndrome (HGPS) is a rare, fatal disease in which patients age prematurely. To identify primary HGPS driver mechanisms, Nard Kubben, Ph.D., a Research Fellow in the laboratory of Tom Misteli, Ph.D., in CCR’s Laboratory of Receptor Biology and Gene Expression, and colleagues in the NCI High-throughput Imaging Facility developed an imaging-based

High-Resolution X-Ray Telescopes

NASA Technical Reports Server (NTRS)

ODell, Stephen L.; Brissenden, Roger J.; Davis, William; Elsner, Ronald F.; Elvis, Martin; Freeman, Mark; Gaetz, Terry; Gorenstein, Paul; Gubarev, Mikhail V.

2010-01-01

Fundamental needs for future x-ray telescopes: a) Sharp images => excellent angular resolution. b) High throughput => large aperture areas. Generation-X optics technical challenges: a) High resolution => precision mirrors & alignment. b) Large apertures => lots of lightweight mirrors. Innovation needed for technical readiness: a) 4 top-level error terms contribute to image size. b) There are approaches to controlling those errors. Innovation needed for manufacturing readiness. Programmatic issues are comparably challenging.

The Portable Remote Imaging Spectrometer (PRISM) Coastal Ocean Sensor

NASA Technical Reports Server (NTRS)

Mouroulis, Pantazis; VanGorp, Byron E.; Green, Robert O.; Eastwppd, Michael; Wilson, Daniel W.; Richardson, Brandon; Dierssen, Heidi

2012-01-01

PRISM is an airborne pushbroom imaging spectrometer intended to address the needs of airborne coastal ocean science research. Its critical characteristics are high throughput and signal-to-noise ratio, high uniformity of response to reduce spectral artifacts, and low polarization sensitivity. We give a brief overview of the instrument and results from laboratory calibration measurements regarding the spatial, spectral, radiometric and polarization characteristics.

Fast Infrared Chemical Imaging with a Quantum Cascade Laser

PubMed Central

2015-01-01

Infrared (IR) spectroscopic imaging systems are a powerful tool for visualizing molecular microstructure of a sample without the need for dyes or stains. Table-top Fourier transform infrared (FT-IR) imaging spectrometers, the current established technology, can record broadband spectral data efficiently but requires scanning the entire spectrum with a low throughput source. The advent of high-intensity, broadly tunable quantum cascade lasers (QCL) has now accelerated IR imaging but results in a fundamentally different type of instrument and approach, namely, discrete frequency IR (DF-IR) spectral imaging. While the higher intensity of the source provides a higher signal per channel, the absence of spectral multiplexing also provides new opportunities and challenges. Here, we couple a rapidly tunable QCL with a high performance microscope equipped with a cooled focal plane array (FPA) detector. Our optical system is conceptualized to provide optimal performance based on recent theory and design rules for high-definition (HD) IR imaging. Multiple QCL units are multiplexed together to provide spectral coverage across the fingerprint region (776.9 to 1904.4 cm–1) in our DF-IR microscope capable of broad spectral coverage, wide-field detection, and diffraction-limited spectral imaging. We demonstrate that the spectral and spatial fidelity of this system is at least as good as the best FT-IR imaging systems. Our configuration provides a speedup for equivalent spectral signal-to-noise ratio (SNR) compared to the best spectral quality from a high-performance linear array system that has 10-fold larger pixels. Compared to the fastest available HD FT-IR imaging system, we demonstrate scanning of large tissue microarrays (TMA) in 3-orders of magnitude smaller time per essential spectral frequency. These advances offer new opportunities for high throughput IR chemical imaging, especially for the measurement of cells and tissues. PMID:25474546

Fast infrared chemical imaging with a quantum cascade laser.

PubMed

Yeh, Kevin; Kenkel, Seth; Liu, Jui-Nung; Bhargava, Rohit

2015-01-06

Infrared (IR) spectroscopic imaging systems are a powerful tool for visualizing molecular microstructure of a sample without the need for dyes or stains. Table-top Fourier transform infrared (FT-IR) imaging spectrometers, the current established technology, can record broadband spectral data efficiently but requires scanning the entire spectrum with a low throughput source. The advent of high-intensity, broadly tunable quantum cascade lasers (QCL) has now accelerated IR imaging but results in a fundamentally different type of instrument and approach, namely, discrete frequency IR (DF-IR) spectral imaging. While the higher intensity of the source provides a higher signal per channel, the absence of spectral multiplexing also provides new opportunities and challenges. Here, we couple a rapidly tunable QCL with a high performance microscope equipped with a cooled focal plane array (FPA) detector. Our optical system is conceptualized to provide optimal performance based on recent theory and design rules for high-definition (HD) IR imaging. Multiple QCL units are multiplexed together to provide spectral coverage across the fingerprint region (776.9 to 1904.4 cm(-1)) in our DF-IR microscope capable of broad spectral coverage, wide-field detection, and diffraction-limited spectral imaging. We demonstrate that the spectral and spatial fidelity of this system is at least as good as the best FT-IR imaging systems. Our configuration provides a speedup for equivalent spectral signal-to-noise ratio (SNR) compared to the best spectral quality from a high-performance linear array system that has 10-fold larger pixels. Compared to the fastest available HD FT-IR imaging system, we demonstrate scanning of large tissue microarrays (TMA) in 3-orders of magnitude smaller time per essential spectral frequency. These advances offer new opportunities for high throughput IR chemical imaging, especially for the measurement of cells and tissues.

Digital micromirror devices in Raman trace detection of explosives

NASA Astrophysics Data System (ADS)

Glimtoft, Martin; Svanqvist, Mattias; Ågren, Matilda; Nordberg, Markus; Östmark, Henric

2016-05-01

Imaging Raman spectroscopy based on tunable filters is an established technique for detecting single explosives particles at stand-off distances. However, large light losses are inherent in the design due to sequential imaging at different wavelengths, leading to effective transmission often well below 1 %. The use of digital micromirror devices (DMD) and compressive sensing (CS) in imaging Raman explosives trace detection can improve light throughput and add significant flexibility compared to existing systems. DMDs are based on mature microelectronics technology, and are compact, scalable, and can be customized for specific tasks, including new functions not available with current technologies. This paper has been focusing on investigating how a DMD can be used when applying CS-based imaging Raman spectroscopy on stand-off explosives trace detection, and evaluating the performance in terms of light throughput, image reconstruction ability and potential detection limits. This type of setup also gives the possibility to combine imaging Raman with non-spatially resolved fluorescence suppression techniques, such as Kerr gating. The system used consists of a 2nd harmonics Nd:YAG laser for sample excitation, collection optics, DMD, CMOScamera and a spectrometer with ICCD camera for signal gating and detection. Initial results for compressive sensing imaging Raman shows a stable reconstruction procedure even at low signals and in presence of interfering background signal. It is also shown to give increased effective light transmission without sacrificing molecular specificity or area coverage compared to filter based imaging Raman. At the same time it adds flexibility so the setup can be customized for new functionality.

Mapping whole-brain activity with cellular resolution by light-sheet microscopy and high-throughput image analysis (Conference Presentation)

NASA Astrophysics Data System (ADS)

Silvestri, Ludovico; Rudinskiy, Nikita; Paciscopi, Marco; Müllenbroich, Marie Caroline; Costantini, Irene; Sacconi, Leonardo; Frasconi, Paolo; Hyman, Bradley T.; Pavone, Francesco S.

2016-03-01

Mapping neuronal activity patterns across the whole brain with cellular resolution is a challenging task for state-of-the-art imaging methods. Indeed, despite a number of technological efforts, quantitative cellular-resolution activation maps of the whole brain have not yet been obtained. Many techniques are limited by coarse resolution or by a narrow field of view. High-throughput imaging methods, such as light sheet microscopy, can be used to image large specimens with high resolution and in reasonable times. However, the bottleneck is then moved from image acquisition to image analysis, since many TeraBytes of data have to be processed to extract meaningful information. Here, we present a full experimental pipeline to quantify neuronal activity in the entire mouse brain with cellular resolution, based on a combination of genetics, optics and computer science. We used a transgenic mouse strain (Arc-dVenus mouse) in which neurons which have been active in the last hours before brain fixation are fluorescently labelled. Samples were cleared with CLARITY and imaged with a custom-made confocal light sheet microscope. To perform an automatic localization of fluorescent cells on the large images produced, we used a novel computational approach called semantic deconvolution. The combined approach presented here allows quantifying the amount of Arc-expressing neurons throughout the whole mouse brain. When applied to cohorts of mice subject to different stimuli and/or environmental conditions, this method helps finding correlations in activity between different neuronal populations, opening the possibility to infer a sort of brain-wide 'functional connectivity' with cellular resolution.

High-throughput method for ear phenotyping and kernel weight estimation in maize using ear digital imaging.

PubMed

Makanza, R; Zaman-Allah, M; Cairns, J E; Eyre, J; Burgueño, J; Pacheco, Ángela; Diepenbrock, C; Magorokosho, C; Tarekegne, A; Olsen, M; Prasanna, B M

2018-01-01

Grain yield, ear and kernel attributes can assist to understand the performance of maize plant under different environmental conditions and can be used in the variety development process to address farmer's preferences. These parameters are however still laborious and expensive to measure. A low-cost ear digital imaging method was developed that provides estimates of ear and kernel attributes i.e., ear number and size, kernel number and size as well as kernel weight from photos of ears harvested from field trial plots. The image processing method uses a script that runs in a batch mode on ImageJ; an open source software. Kernel weight was estimated using the total kernel number derived from the number of kernels visible on the image and the average kernel size. Data showed a good agreement in terms of accuracy and precision between ground truth measurements and data generated through image processing. Broad-sense heritability of the estimated parameters was in the range or higher than that for measured grain weight. Limitation of the method for kernel weight estimation is discussed. The method developed in this work provides an opportunity to significantly reduce the cost of selection in the breeding process, especially for resource constrained crop improvement programs and can be used to learn more about the genetic bases of grain yield determinants.

Three-Dimensional Root Phenotyping with a Novel Imaging and Software Platform1[C][W][OA

PubMed Central

Clark, Randy T.; MacCurdy, Robert B.; Jung, Janelle K.; Shaff, Jon E.; McCouch, Susan R.; Aneshansley, Daniel J.; Kochian, Leon V.

2011-01-01

A novel imaging and software platform was developed for the high-throughput phenotyping of three-dimensional root traits during seedling development. To demonstrate the platform’s capacity, plants of two rice (Oryza sativa) genotypes, Azucena and IR64, were grown in a transparent gellan gum system and imaged daily for 10 d. Rotational image sequences consisting of 40 two-dimensional images were captured using an optically corrected digital imaging system. Three-dimensional root reconstructions were generated and analyzed using a custom-designed software, RootReader3D. Using the automated and interactive capabilities of RootReader3D, five rice root types were classified and 27 phenotypic root traits were measured to characterize these two genotypes. Where possible, measurements from the three-dimensional platform were validated and were highly correlated with conventional two-dimensional measurements. When comparing gellan gum-grown plants with those grown under hydroponic and sand culture, significant differences were detected in morphological root traits (P < 0.05). This highly flexible platform provides the capacity to measure root traits with a high degree of spatial and temporal resolution and will facilitate novel investigations into the development of entire root systems or selected components of root systems. In combination with the extensive genetic resources that are now available, this platform will be a powerful resource to further explore the molecular and genetic determinants of root system architecture. PMID:21454799

A noninvasive, direct real-time PCR method for sex determination in multiple avian species

USGS Publications Warehouse

Brubaker, Jessica L.; Karouna-Renier, Natalie K.; Chen, Yu; Jenko, Kathryn; Sprague, Daniel T.; Henry, Paula F.P.

2011-01-01

Polymerase chain reaction (PCR)-based methods to determine the sex of birds are well established and have seen few modifications since they were first introduced in the 1990s. Although these methods allowed for sex determination in species that were previously difficult to analyse, they were not conducive to high-throughput analysis because of the laboriousness of DNA extraction and gel electrophoresis. We developed a high-throughput real-time PCR-based method for analysis of sex in birds, which uses noninvasive sample collection and avoids DNA extraction and gel electrophoresis.

High-throughput sample processing and sample management; the functional evolution of classical cytogenetic assay towards automation.

PubMed

Ramakumar, Adarsh; Subramanian, Uma; Prasanna, Pataje G S

2015-11-01

High-throughput individual diagnostic dose assessment is essential for medical management of radiation-exposed subjects after a mass casualty. Cytogenetic assays such as the Dicentric Chromosome Assay (DCA) are recognized as the gold standard by international regulatory authorities. DCA is a multi-step and multi-day bioassay. DCA, as described in the IAEA manual, can be used to assess dose up to 4-6 weeks post-exposure quite accurately but throughput is still a major issue and automation is very essential. The throughput is limited, both in terms of sample preparation as well as analysis of chromosome aberrations. Thus, there is a need to design and develop novel solutions that could utilize extensive laboratory automation for sample preparation, and bioinformatics approaches for chromosome-aberration analysis to overcome throughput issues. We have transitioned the bench-based cytogenetic DCA to a coherent process performing high-throughput automated biodosimetry for individual dose assessment ensuring quality control (QC) and quality assurance (QA) aspects in accordance with international harmonized protocols. A Laboratory Information Management System (LIMS) is designed, implemented and adapted to manage increased sample processing capacity, develop and maintain standard operating procedures (SOP) for robotic instruments, avoid data transcription errors during processing, and automate analysis of chromosome-aberrations using an image analysis platform. Our efforts described in this paper intend to bridge the current technological gaps and enhance the potential application of DCA for a dose-based stratification of subjects following a mass casualty. This paper describes one such potential integrated automated laboratory system and functional evolution of the classical DCA towards increasing critically needed throughput. Published by Elsevier B.V.

Biochemical and High Throughput Microscopic Assessment of Fat Mass in Caenorhabditis Elegans

PubMed Central

Pino, Elizabeth C.; Webster, Christopher M.; Carr, Christopher E.; Soukas, Alexander A.

2013-01-01

The nematode C. elegans has emerged as an important model for the study of conserved genetic pathways regulating fat metabolism as it relates to human obesity and its associated pathologies. Several previous methodologies developed for the visualization of C. elegans triglyceride-rich fat stores have proven to be erroneous, highlighting cellular compartments other than lipid droplets. Other methods require specialized equipment, are time-consuming, or yield inconsistent results. We introduce a rapid, reproducible, fixative-based Nile red staining method for the accurate and rapid detection of neutral lipid droplets in C. elegans. A short fixation step in 40% isopropanol makes animals completely permeable to Nile red, which is then used to stain animals. Spectral properties of this lipophilic dye allow it to strongly and selectively fluoresce in the yellow-green spectrum only when in a lipid-rich environment, but not in more polar environments. Thus, lipid droplets can be visualized on a fluorescent microscope equipped with simple GFP imaging capability after only a brief Nile red staining step in isopropanol. The speed, affordability, and reproducibility of this protocol make it ideally suited for high throughput screens. We also demonstrate a paired method for the biochemical determination of triglycerides and phospholipids using gas chromatography mass-spectrometry. This more rigorous protocol should be used as confirmation of results obtained from the Nile red microscopic lipid determination. We anticipate that these techniques will become new standards in the field of C. elegans metabolic research. PMID:23568026

Robust reflective pupil slicing technology

NASA Astrophysics Data System (ADS)

Meade, Jeffrey T.; Behr, Bradford B.; Cenko, Andrew T.; Hajian, Arsen R.

2014-07-01

Tornado Spectral Systems (TSS) has developed the High Throughput Virtual Slit (HTVSTM), robust all-reflective pupil slicing technology capable of replacing the slit in research-, commercial- and MIL-SPEC-grade spectrometer systems. In the simplest configuration, the HTVS allows optical designers to remove the lossy slit from pointsource spectrometers and widen the input slit of long-slit spectrometers, greatly increasing throughput without loss of spectral resolution or cross-dispersion information. The HTVS works by transferring etendue between image plane axes but operating in the pupil domain rather than at a focal plane. While useful for other technologies, this is especially relevant for spectroscopic applications by performing the same spectral narrowing as a slit without throwing away light on the slit aperture. HTVS can be implemented in all-reflective designs and only requires a small number of reflections for significant spectral resolution enhancement-HTVS systems can be efficiently implemented in most wavelength regions. The etendueshifting operation also provides smooth scaling with input spot/image size without requiring reconfiguration for different targets (such as different seeing disk diameters or different fiber core sizes). Like most slicing technologies, HTVS provides throughput increases of several times without resolution loss over equivalent slitbased designs. HTVS technology enables robust slit replacement in point-source spectrometer systems. By virtue of pupilspace operation this technology has several advantages over comparable image-space slicer technology, including the ability to adapt gracefully and linearly to changing source size and better vertical packing of the flux distribution. Additionally, this technology can be implemented with large slicing factors in both fast and slow beams and can easily scale from large, room-sized spectrometers through to small, telescope-mounted devices. Finally, this same technology is directly applicable to multi-fiber spectrometers to achieve similar enhancement. HTVS also provides the ability to anamorphically "stretch" the slit image in long-slit spectrometers, allowing the instrument designer to optimize the plate scale in the dispersion axis and cross-dispersion axes independently without sacrificing spatial information. This allows users to widen the input slit, with the associated gain of throughput and loss of spatial selectivity, while maintaining the spectral resolution of the spectrometer system. This "stretching" places increased requirements on detector focal plane height, as with image slicing techniques, but provides additional degrees of freedom to instrument designers to build the best possible spectrometer systems. We discuss the details of this technology for an astronomical context, covering the applicability from small telescope mounted spectrometers through long-slit imagers and radial-velocity engines. This powerful tool provides additional degrees of freedom when designing a spectrometer, enabling instrument designers to further optimize systems for the required scientific goals.

Analyzing microtomography data with Python and the scikit-image library.

PubMed

Gouillart, Emmanuelle; Nunez-Iglesias, Juan; van der Walt, Stéfan

2017-01-01

The exploration and processing of images is a vital aspect of the scientific workflows of many X-ray imaging modalities. Users require tools that combine interactivity, versatility, and performance. scikit-image is an open-source image processing toolkit for the Python language that supports a large variety of file formats and is compatible with 2D and 3D images. The toolkit exposes a simple programming interface, with thematic modules grouping functions according to their purpose, such as image restoration, segmentation, and measurements. scikit-image users benefit from a rich scientific Python ecosystem that contains many powerful libraries for tasks such as visualization or machine learning. scikit-image combines a gentle learning curve, versatile image processing capabilities, and the scalable performance required for the high-throughput analysis of X-ray imaging data.

Protocols and programs for high-throughput growth and aging phenotyping in yeast.

PubMed

Jung, Paul P; Christian, Nils; Kay, Daniel P; Skupin, Alexander; Linster, Carole L

2015-01-01

In microorganisms, and more particularly in yeasts, a standard phenotyping approach consists in the analysis of fitness by growth rate determination in different conditions. One growth assay that combines high throughput with high resolution involves the generation of growth curves from 96-well plate microcultivations in thermostated and shaking plate readers. To push the throughput of this method to the next level, we have adapted it in this study to the use of 384-well plates. The values of the extracted growth parameters (lag time, doubling time and yield of biomass) correlated well between experiments carried out in 384-well plates as compared to 96-well plates or batch cultures, validating the higher-throughput approach for phenotypic screens. The method is not restricted to the use of the budding yeast Saccharomyces cerevisiae, as shown by consistent results for other species selected from the Hemiascomycete class. Furthermore, we used the 384-well plate microcultivations to develop and validate a higher-throughput assay for yeast Chronological Life Span (CLS), a parameter that is still commonly determined by a cumbersome method based on counting "Colony Forming Units". To accelerate analysis of the large datasets generated by the described growth and aging assays, we developed the freely available software tools GATHODE and CATHODE. These tools allow for semi-automatic determination of growth parameters and CLS behavior from typical plate reader output files. The described protocols and programs will increase the time- and cost-efficiency of a number of yeast-based systems genetics experiments as well as various types of screens.

High Throughput Screening Identifies Novel Lead Compounds with Activity against Larval, Juvenile and Adult Schistosoma mansoni

PubMed Central

Gardner, J. Mark F.; Bell, Andrew S.; Parkinson, Tanya; Bickle, Quentin

2016-01-01

An estimated 600 million people are affected by the helminth disease schistosomiasis caused by parasites of the genus Schistosoma. There is currently only one drug recommended for treating schistosomiasis, praziquantel (PZQ), which is effective against adult worms but not against the juvenile stage. In an attempt to identify improved drugs for treating the disease, we have carried out high throughput screening of a number of small molecule libraries with the aim of identifying lead compounds with balanced activity against all life stages of Schistosoma. A total of almost 300,000 compounds were screened using a high throughput assay based on motility of worm larvae and image analysis of assay plates. Hits were screened against juvenile and adult worms to identify broadly active compounds and against a mammalian cell line to assess cytotoxicity. A number of compounds were identified as promising leads for further chemical optimization. PMID:27128493

Shot-noise limited throughput of soft x-ray ptychography for nanometrology applications

NASA Astrophysics Data System (ADS)

Koek, Wouter; Florijn, Bastiaan; Bäumer, Stefan; Kruidhof, Rik; Sadeghian, Hamed

2018-03-01

Due to its potential for high resolution and three-dimensional imaging, soft x-ray ptychography has received interest for nanometrology applications. We have analyzed the measurement time per unit area when using soft x-ray ptychography for various nanometrology applications including mask inspection and wafer inspection, and are thus able to predict (order of magnitude) throughput figures. Here we show that for a typical measurement system, using a typical sampling strategy, and when aiming for 10-15 nm resolution, it is expected that a wafer-based topology (2.5D) measurement takes approximately 4 minutes per μm2 , and a full three-dimensional measurement takes roughly 6 hours per μm2 . Due to their much higher reflectivity EUV masks can be measured considerably faster; a measurement speed of 0.1 seconds per μm2 is expected. However, such speeds do not allow for full wafer or mask inspection at industrially relevant throughput.

Cox-nnet: An artificial neural network method for prognosis prediction of high-throughput omics data.

PubMed

Ching, Travers; Zhu, Xun; Garmire, Lana X

2018-04-01

Artificial neural networks (ANN) are computing architectures with many interconnections of simple neural-inspired computing elements, and have been applied to biomedical fields such as imaging analysis and diagnosis. We have developed a new ANN framework called Cox-nnet to predict patient prognosis from high throughput transcriptomics data. In 10 TCGA RNA-Seq data sets, Cox-nnet achieves the same or better predictive accuracy compared to other methods, including Cox-proportional hazards regression (with LASSO, ridge, and mimimax concave penalty), Random Forests Survival and CoxBoost. Cox-nnet also reveals richer biological information, at both the pathway and gene levels. The outputs from the hidden layer node provide an alternative approach for survival-sensitive dimension reduction. In summary, we have developed a new method for accurate and efficient prognosis prediction on high throughput data, with functional biological insights. The source code is freely available at https://github.com/lanagarmire/cox-nnet.

Continuing Development of Alternative High-Throughput Screens to Determine Endocrine Disruption, Focusing on Androgen Receptor, Steroidogenesis, and Thyroid Pathways

EPA Science Inventory

The focus of this meeting is the SAP's review and comment on the Agency's proposed high-throughput computational model of androgen receptor pathway activity as an alternative to the current Tier 1 androgen receptor assay (OCSPP 890.1150: Androgen Receptor Binding Rat Prostate Cyt...

Development of an automated large-scale protein-crystallization and monitoring system for high-throughput protein-structure analyses.

PubMed

Hiraki, Masahiko; Kato, Ryuichi; Nagai, Minoru; Satoh, Tadashi; Hirano, Satoshi; Ihara, Kentaro; Kudo, Norio; Nagae, Masamichi; Kobayashi, Masanori; Inoue, Michio; Uejima, Tamami; Oda, Shunichiro; Chavas, Leonard M G; Akutsu, Masato; Yamada, Yusuke; Kawasaki, Masato; Matsugaki, Naohiro; Igarashi, Noriyuki; Suzuki, Mamoru; Wakatsuki, Soichi

2006-09-01

Protein crystallization remains one of the bottlenecks in crystallographic analysis of macromolecules. An automated large-scale protein-crystallization system named PXS has been developed consisting of the following subsystems, which proceed in parallel under unified control software: dispensing precipitants and protein solutions, sealing crystallization plates, carrying robot, incubators, observation system and image-storage server. A sitting-drop crystallization plate specialized for PXS has also been designed and developed. PXS can set up 7680 drops for vapour diffusion per hour, which includes time for replenishing supplies such as disposable tips and crystallization plates. Images of the crystallization drops are automatically recorded according to a preprogrammed schedule and can be viewed by users remotely using web-based browser software. A number of protein crystals were successfully produced and several protein structures could be determined directly from crystals grown by PXS. In other cases, X-ray quality crystals were obtained by further optimization by manual screening based on the conditions found by PXS.

The Preflight Photometric Calibration of the Extreme-Ultraviolet Imaging Telescope EIT

NASA Technical Reports Server (NTRS)

Dere, K. P.; Moses, J. D.; Delaboudiniere, J. -P.; Brunaud, J.; Carabetian, C.; Hochedez, J. -F.; Song, X. Y.; Catura, R. C.; Clette, F.; Defise, J. -M.

2000-01-01

This paper presents the preflight photometric calibration of the Extreme-ultraviolet Imaging Telescope (EIT) aboard the Solar and Heliospheric Observatory (SOHO). The EIT consists of a Ritchey-Chretien telescope with multilayer coatings applied to four quadrants of the primary and secondary mirrors, several filters and a backside-thinned CCD detector. The quadrants of the EIT optics were used to observe the Sun in 4 wavelength bands that peak near 171, 195, 284, and 304 . Before the launch of SOHO, the EIT mirror reflectivities, the filter transmissivities and the CCD quantum efficiency were measured and these values are described here. The instrumental throughput in terms of an effective area is presented for each of the various mirror quadrant and filter wheel combinations. The response to a coronal plasma as a function of temperature is also determined and the expected count rates are compared to the count rates observed in a coronal hole, the quiet Sun and an active region.

The high throughput virtual slit enables compact, inexpensive Raman spectral imagers

NASA Astrophysics Data System (ADS)

Gooding, Edward; Deutsch, Erik R.; Huehnerhoff, Joseph; Hajian, Arsen R.

2018-02-01

Raman spectral imaging is increasingly becoming the tool of choice for field-based applications such as threat, narcotics and hazmat detection; air, soil and water quality monitoring; and material ID. Conventional fiber-coupled point source Raman spectrometers effectively interrogate a small sample area and identify bulk samples via spectral library matching. However, these devices are very slow at mapping over macroscopic areas. In addition, the spatial averaging performed by instruments that collect binned spectra, particularly when used in combination with orbital raster scanning, tends to dilute the spectra of trace particles in a mixture. Our design, employing free space line illumination combined with area imaging, reveals both the spectral and spatial content of heterogeneous mixtures. This approach is well suited to applications such as detecting explosives and narcotics trace particle detection in fingerprints. The patented High Throughput Virtual Slit1 is an innovative optical design that enables compact, inexpensive handheld Raman spectral imagers. HTVS-based instruments achieve significantly higher spectral resolution than can be obtained with conventional designs of the same size. Alternatively, they can be used to build instruments with comparable resolution to large spectrometers, but substantially smaller size, weight and unit cost, all while maintaining high sensitivity. When used in combination with laser line imaging, this design eliminates sample photobleaching and unwanted photochemistry while greatly enhancing mapping speed, all with high selectivity and sensitivity. We will present spectral image data and discuss applications that are made possible by low cost HTVS-enabled instruments.

Quantitative determination of free/bound atazanavir via high-throughput equilibrium dialysis and LC-MS/MS, and the application in ex vivo samples.

PubMed

Xu, Xiaohui Sophia; Rose, Anne; Demers, Roger; Eley, Timothy; Ryan, John; Stouffer, Bruce; Cojocaru, Laura; Arnold, Mark

2014-01-01

The determination of drug-protein binding is important in the pharmaceutical development process because of the impact of protein binding on both the pharmacokinetics and pharmacodynamics of drugs. Equilibrium dialysis is the preferred method to measure the free drug fraction because it is considered to be more accurate. The throughput of equilibrium dialysis has recently been improved by implementing a 96-well format plate. Results/methodology: This manuscript illustrates the successful application of a 96-well rapid equilibrium dialysis (RED) device in the determination of atazanavir plasma-protein binding. This RED method of measuring free fraction was successfully validated and then applied to the analysis of clinical plasma samples taken from HIV-infected pregnant women administered atazanavir. Combined with LC-MS/MS detection, the 96-well format equilibrium dialysis device was suitable for measuring the free and bound concentration of pharmaceutical molecules in a high-throughput mode.

Cytometric analysis of retinopathies in retinal trypsin digests

NASA Astrophysics Data System (ADS)

Ghanian, Zahra; Staniszewski, Kevin; Sorenson, Christine M.; Sheibani, Nader; Ranji, Mahsa

2014-03-01

The objective of this work was to design an automated image cytometry tool for determination of various retinal vascular parameters including extraction of features that are relevant to postnatal retinal vascular development, and the progression of diabetic retinopathy. To confirm the utility and accuracy of the software, retinal trypsin digest from TSP1-/- and diabetic Akita/+; TSP1-/- mice were analyzed. TSP1 is a critical inhibitor of development of retinopathies and lack of TSP1 exacerbates progression of early diabetic retinopathies. Loss of vascular cells of and gain more acellular capillaries as two major signs of diabetic retinopathies were used to classify a retina as normal or injured. This software allows quantification and high throughput assessment of retinopathy changes associated with diabetes.

HTML5 PivotViewer: high-throughput visualization and querying of image data on the web.

PubMed

Taylor, Stephen; Noble, Roger

2014-09-15

Visualization and analysis of large numbers of biological images has generated a bottle neck in research. We present HTML5 PivotViewer, a novel, open source, platform-independent viewer making use of the latest web technologies that allows seamless access to images and associated metadata for each image. This provides a powerful method to allow end users to mine their data. Documentation, examples and links to the software are available from http://www.cbrg.ox.ac.uk/data/pivotviewer/. The software is licensed under GPLv2. © The Author 2014. Published by Oxford University Press.

Resolution modeling of dispersive imaging spectrometers

NASA Astrophysics Data System (ADS)

Silny, John F.

2017-08-01

This paper presents best practices for modeling the resolution of dispersive imaging spectrometers. The differences between sampling, width, and resolution are discussed. It is proposed that the spectral imaging community adopt a standard definition for resolution as the full-width at half maximum of the total line spread function. Resolution should be computed for each of the spectral, cross-scan spatial, and along-scan spatial/temporal dimensions separately. A physical optics resolution model is presented that incorporates the effects of slit diffraction and partial coherence, the result of which is a narrower slit image width and reduced radiometric throughput.

Tools for automating the imaging of zebrafish larvae.

PubMed

Pulak, Rock

2016-03-01

The VAST BioImager system is a set of tools developed for zebrafish researchers who require the collection of images from a large number of 2-7 dpf zebrafish larvae. The VAST BioImager automates larval handling, positioning and orientation tasks. Color images at about 10 μm resolution are collected from the on-board camera of the system. If images of greater resolution and detail are required, this system is mounted on an upright microscope, such as a confocal or fluorescence microscope, to utilize their capabilities. The system loads a larvae, positions it in view of the camera, determines orientation using pattern recognition analysis, and then more precisely positions to user-defined orientation for optimal imaging of any desired tissue or organ system. Multiple images of the same larva can be collected. The specific part of each larva and the desired orientation and position is identified by the researcher and an experiment defining the settings and a series of steps can be saved and repeated for imaging of subsequent larvae. The system captures images, then ejects and loads another larva from either a bulk reservoir, a well of a 96 well plate using the LP Sampler, or individually targeted larvae from a Petri dish or other container using the VAST Pipettor. Alternative manual protocols for handling larvae for image collection are tedious and time consuming. The VAST BioImager automates these steps to allow for greater throughput of assays and screens requiring high-content image collection of zebrafish larvae such as might be used in drug discovery and toxicology studies. Copyright © 2015 The Author. Published by Elsevier Inc. All rights reserved.

Nanoparticle light scattering on interferometric surfaces

NASA Astrophysics Data System (ADS)

Hayrapetyan, K.; Arif, K. M.; Savran, C. A.; Nolte, D. D.

2011-03-01

We present a model based on Mie Surface Double Interaction (MSDI) to explore bead-based detection mechanisms using imaging and scanning. The application goal of this work is to explore the trade-offs between the sensitivity and throughput among various detection methods. Experimentally we use thermal oxide on silicon to establish and control surface interferometric conditions. Surface-captured gold beads are detected using Molecular Interferometric Imaging (MI2) and Spinning-Disc Interferometry (SDI).

Automatic high throughput empty ISO container verification

NASA Astrophysics Data System (ADS)

Chalmers, Alex

2007-04-01

Encouraging results are presented for the automatic analysis of radiographic images of a continuous stream of ISO containers to confirm they are truly empty. A series of image processing algorithms are described that process real-time data acquired during the actual inspection of each container and assigns each to one of the classes "empty", "not empty" or "suspect threat". This research is one step towards achieving fully automated analysis of cargo containers.

High-throughput methods for electron crystallography.

PubMed

Stokes, David L; Ubarretxena-Belandia, Iban; Gonen, Tamir; Engel, Andreas

2013-01-01

Membrane proteins play a tremendously important role in cell physiology and serve as a target for an increasing number of drugs. Structural information is key to understanding their function and for developing new strategies for combating disease. However, the complex physical chemistry associated with membrane proteins has made them more difficult to study than their soluble cousins. Electron crystallography has historically been a successful method for solving membrane protein structures and has the advantage of providing a native lipid environment for these proteins. Specifically, when membrane proteins form two-dimensional arrays within a lipid bilayer, electron microscopy can be used to collect images and diffraction and the corresponding data can be combined to produce a three-dimensional reconstruction, which under favorable conditions can extend to atomic resolution. Like X-ray crystallography, the quality of the structures are very much dependent on the order and size of the crystals. However, unlike X-ray crystallography, high-throughput methods for screening crystallization trials for electron crystallography are not in general use. In this chapter, we describe two alternative methods for high-throughput screening of membrane protein crystallization within the lipid bilayer. The first method relies on the conventional use of dialysis for removing detergent and thus reconstituting the bilayer; an array of dialysis wells in the standard 96-well format allows the use of a liquid-handling robot and greatly increases throughput. The second method relies on titration of cyclodextrin as a chelating agent for detergent; a specialized pipetting robot has been designed not only to add cyclodextrin in a systematic way, but to use light scattering to monitor the reconstitution process. In addition, the use of liquid-handling robots for making negatively stained grids and methods for automatically imaging samples in the electron microscope are described.

Fast and Adaptive Lossless Onboard Hyperspectral Data Compression System

NASA Technical Reports Server (NTRS)

Aranki, Nazeeh I.; Keymeulen, Didier; Kimesh, Matthew A.

2012-01-01

Modern hyperspectral imaging systems are able to acquire far more data than can be downlinked from a spacecraft. Onboard data compression helps to alleviate this problem, but requires a system capable of power efficiency and high throughput. Software solutions have limited throughput performance and are power-hungry. Dedicated hardware solutions can provide both high throughput and power efficiency, while taking the load off of the main processor. Thus a hardware compression system was developed. The implementation uses a field-programmable gate array (FPGA). The implementation is based on the fast lossless (FL) compression algorithm reported in Fast Lossless Compression of Multispectral-Image Data (NPO-42517), NASA Tech Briefs, Vol. 30, No. 8 (August 2006), page 26, which achieves excellent compression performance and has low complexity. This algorithm performs predictive compression using an adaptive filtering method, and uses adaptive Golomb coding. The implementation also packetizes the coded data. The FL algorithm is well suited for implementation in hardware. In the FPGA implementation, one sample is compressed every clock cycle, which makes for a fast and practical realtime solution for space applications. Benefits of this implementation are: 1) The underlying algorithm achieves a combination of low complexity and compression effectiveness that exceeds that of techniques currently in use. 2) The algorithm requires no training data or other specific information about the nature of the spectral bands for a fixed instrument dynamic range. 3) Hardware acceleration provides a throughput improvement of 10 to 100 times vs. the software implementation. A prototype of the compressor is available in software, but it runs at a speed that does not meet spacecraft requirements. The hardware implementation targets the Xilinx Virtex IV FPGAs, and makes the use of this compressor practical for Earth satellites as well as beyond-Earth missions with hyperspectral instruments.

Time-Domain Microfluidic Fluorescence Lifetime Flow Cytometry for High-Throughput Förster Resonance Energy Transfer Screening

PubMed Central

Nedbal, Jakub; Visitkul, Viput; Ortiz-Zapater, Elena; Weitsman, Gregory; Chana, Prabhjoat; Matthews, Daniel R; Ng, Tony; Ameer-Beg, Simon M

2015-01-01

Sensing ion or ligand concentrations, physico-chemical conditions, and molecular dimerization or conformation change is possible by assays involving fluorescent lifetime imaging. The inherent low throughput of imaging impedes rigorous statistical data analysis on large cell numbers. We address this limitation by developing a fluorescence lifetime-measuring flow cytometer for fast fluorescence lifetime quantification in living or fixed cell populations. The instrument combines a time-correlated single photon counting epifluorescent microscope with microfluidics cell-handling system. The associated computer software performs burst integrated fluorescence lifetime analysis to assign fluorescence lifetime, intensity, and burst duration to each passing cell. The maximum safe throughput of the instrument reaches 3,000 particles per minute. Living cells expressing spectroscopic rulers of varying peptide lengths were distinguishable by Förster resonant energy transfer measured by donor fluorescence lifetime. An epidermal growth factor (EGF)-stimulation assay demonstrated the technique's capacity to selectively quantify EGF receptor phosphorylation in cells, which was impossible by measuring sensitized emission on a standard flow cytometer. Dual-color fluorescence lifetime detection and cell-specific chemical environment sensing were exemplified using di-4-ANEPPDHQ, a lipophilic environmentally sensitive dye that exhibits changes in its fluorescence lifetime as a function of membrane lipid order. To our knowledge, this instrument opens new applications in flow cytometry which were unavailable due to technological limitations of previously reported fluorescent lifetime flow cytometers. The presented technique is sensitive to lifetimes of most popular fluorophores in the 0.5–5 ns range including fluorescent proteins and is capable of detecting multi-exponential fluorescence lifetime decays. This instrument vastly enhances the throughput of experiments involving fluorescence lifetime measurements, thereby providing statistically significant quantitative data for analysis of large cell populations. © 2014 International Society for Advancement of Cytometry PMID:25523156

FPGA Implementation of Stereo Disparity with High Throughput for Mobility Applications

NASA Technical Reports Server (NTRS)

Villalpando, Carlos Y.; Morfopolous, Arin; Matthies, Larry; Goldberg, Steven

2011-01-01

High speed stereo vision can allow unmanned robotic systems to navigate safely in unstructured terrain, but the computational cost can exceed the capacity of typical embedded CPUs. In this paper, we describe an end-to-end stereo computation co-processing system optimized for fast throughput that has been implemented on a single Virtex 4 LX160 FPGA. This system is capable of operating on images from a 1024 x 768 3CCD (true RGB) camera pair at 15 Hz. Data enters the FPGA directly from the cameras via Camera Link and is rectified, pre-filtered and converted into a disparity image all within the FPGA, incurring no CPU load. Once complete, a rectified image and the final disparity image are read out over the PCI bus, for a bandwidth cost of 68 MB/sec. Within the FPGA there are 4 distinct algorithms: Camera Link capture, Bilinear rectification, Bilateral subtraction pre-filtering and the Sum of Absolute Difference (SAD) disparity. Each module will be described in brief along with the data flow and control logic for the system. The system has been successfully fielded upon the Carnegie Mellon University's National Robotics Engineering Center (NREC) Crusher system during extensive field trials in 2007 and 2008 and is being implemented for other surface mobility systems at JPL.

Multifocal multiphoton microscopy with adaptive optical correction

NASA Astrophysics Data System (ADS)

Coelho, Simao; Poland, Simon; Krstajic, Nikola; Li, David; Monypenny, James; Walker, Richard; Tyndall, David; Ng, Tony; Henderson, Robert; Ameer-Beg, Simon

2013-02-01

Fluorescence lifetime imaging microscopy (FLIM) is a well established approach for measuring dynamic signalling events inside living cells, including detection of protein-protein interactions. The improvement in optical penetration of infrared light compared with linear excitation due to Rayleigh scattering and low absorption have provided imaging depths of up to 1mm in brain tissue but significant image degradation occurs as samples distort (aberrate) the infrared excitation beam. Multiphoton time-correlated single photon counting (TCSPC) FLIM is a method for obtaining functional, high resolution images of biological structures. In order to achieve good statistical accuracy TCSPC typically requires long acquisition times. We report the development of a multifocal multiphoton microscope (MMM), titled MegaFLI. Beam parallelization performed via a 3D Gerchberg-Saxton (GS) algorithm using a Spatial Light Modulator (SLM), increases TCSPC count rate proportional to the number of beamlets produced. A weighted 3D GS algorithm is employed to improve homogeneity. An added benefit is the implementation of flexible and adaptive optical correction. Adaptive optics performed by means of Zernike polynomials are used to correct for system induced aberrations. Here we present results with significant improvement in throughput obtained using a novel complementary metal-oxide-semiconductor (CMOS) 1024 pixel single-photon avalanche diode (SPAD) array, opening the way to truly high-throughput FLIM.

Characterisation of a CZT detector for dosimetry of molecular radiotherapy

NASA Astrophysics Data System (ADS)

McAreavey, L. H.; Harkness-Brennan, L. J.; Colosimo, S. J.; Judson, D. S.; Boston, A. J.; Boston, H. C.; Nolan, P. J.; Flux, G. D.; Denis-Bacelar, A. M.; Harris, B.; Radley, I.; Carroll, M.

2017-03-01

A pixelated cadmium zinc telluride (CZT) detector has been characterised for the purpose of developing a quantitative single photon emission computed tomography (SPECT) system for dosimetry of molecular radiotherapy (MRT). This is the aim of the Dosimetric Imaging with CZT (DEPICT) project, which is a collaboration between the University of Liverpool, The Royal Marsden Hospital, The Royal Liverpool and Broadgreen University Hospital, and the commercial partner Kromek. CZT is a direct band gap semiconductor with superior energy resolution and stopping power compared to scintillator detectors used in current SPECT systems. The inherent detector properties have been investigated and operational parameters such as bias voltage and peaking time have been selected to optimise the performance of the system. Good energy resolution is required to discriminate γ-rays that are scattered as they are emitted from the body and within the collimator, and high photon throughput is essential due to the high activities of isotopes administered in MRT. The system has an average measured electronic noise of 3.31 keV full width at half maximum (FWHM), determined through the use of an internal pulser. The energy response of the system was measured across the energy region of interest 59.5 keV to 364.5 keV and found to be linear. The reverse bias voltage and peaking time producing the optimum FWHM and maximum photon throughput were 600 V and 0.5 μs respectively. The average dead time of the system was measured as 4.84 μs and charge sharing was quantified to be 0.71 % at 59.5 keV . A pixel sensitivity calibration map was created and planar images of the medical imaging isotopes 99mTc and 123I were acquired by coupling the device to a prototype collimator, thereby demonstrating the suitability of the detector for the DEPICT project.

Ground Field-Based Hyperspectral Imaging: A Preliminary Study to Assess the Potential of Established Vegetation Indices to Infer Variation in Water-Use Efficiency.

NASA Astrophysics Data System (ADS)

Pelech, E. A.; McGrath, J.; Pederson, T.; Bernacchi, C.

2017-12-01

Increases in the global average temperature will consequently induce a higher occurrence of severe environmental conditions such as drought on arable land. To mitigate these threats, crops for fuel and food must be bred for higher water-use efficiencies (WUE). Defining genomic variation through high-throughput phenotypic analysis in field conditions has the potential to relieve the major bottleneck in linking desirable genetic traits to the associated phenotypic response. This can subsequently enable breeders to create new agricultural germplasm that supports the need for higher water-use efficient crops. From satellites to field-based aerial and ground sensors, the reflectance properties of vegetation measured by hyperspectral imaging is becoming a rapid high-throughput phenotyping technique. A variety of physiological traits can be inferred by regression analysis with leaf reflectance which is controlled by the properties and abundance of water, carbon, nitrogen and pigments. Although, given that the current established vegetation indices are designed to accentuate these properties from spectral reflectance, it becomes a challenge to infer relative measurements of WUE at a crop canopy scale without ground-truth data collection. This study aims to correlate established biomass and canopy-water-content indices with ground-truth data. Five bioenergy sorghum genotypes (Sorghum bicolor L. Moench) that have differences in WUE and wild-type Tobacco (Nicotiana tabacum var. Samsun) under irrigated and rainfed field conditions were examined. A linear regression analysis was conducted to determine if variation in canopy water content and biomass, driven by natural genotypic and artificial treatment influences, can be inferred using established vegetation indices. The results from this study will elucidate the ability of ground field-based hyperspectral imaging to assess variation in water content, biomass and water-use efficiency. This can lead to improved opportunities to select ideal genotypes for an increasing water-limited environment and to help parameterize and validate terrestrial vegetation models that require a better representation of genetic variation within crop species.

External optical imaging of freely moving mice with green fluorescent protein-expressing metastatic tumors

NASA Astrophysics Data System (ADS)

Yang, Meng; Baranov, Eugene; Shimada, Hiroshi; Moossa, A. R.; Hoffman, Robert M.

2000-04-01

We report here a new approach to genetically engineering tumors to become fluorescence such that they can be imaged externally in freely-moving animals. We describe here external high-resolution real-time fluorescent optical imaging of metastatic tumors in live mice. Stable high-level green flourescent protein (GFP)-expressing human and rodent cell lines enable tumors and metastasis is formed from them to be externally imaged from freely-moving mice. Real-time tumor and metastatic growth were quantitated from whole-body real-time imaging in GFP-expressing melanoma and colon carcinoma models. This GFP optical imaging system is highly appropriate for high throughput in vivo drug screening.

Plant phenomics: an overview of image acquisition technologies and image data analysis algorithms.

PubMed

Perez-Sanz, Fernando; Navarro, Pedro J; Egea-Cortines, Marcos

2017-11-01

The study of phenomes or phenomics has been a central part of biology. The field of automatic phenotype acquisition technologies based on images has seen an important advance in the last years. As with other high-throughput technologies, it addresses a common set of problems, including data acquisition and analysis. In this review, we give an overview of the main systems developed to acquire images. We give an in-depth analysis of image processing with its major issues and the algorithms that are being used or emerging as useful to obtain data out of images in an automatic fashion. © The Author 2017. Published by Oxford University Press.

A high-throughput microtiter plate based method for the determination of peracetic acid and hydrogen peroxide.

PubMed

Putt, Karson S; Pugh, Randall B

2013-01-01

Peracetic acid is gaining usage in numerous industries who have found a myriad of uses for its antimicrobial activity. However, rapid high throughput quantitation methods for peracetic acid and hydrogen peroxide are lacking. Herein, we describe the development of a high-throughput microtiter plate based assay based upon the well known and trusted titration chemical reactions. The adaptation of these titration chemistries to rapid plate based absorbance methods for the sequential determination of hydrogen peroxide specifically and the total amount of peroxides present in solution are described. The results of these methods were compared to those of a standard titration and found to be in good agreement. Additionally, the utility of the developed method is demonstrated through the generation of degradation curves of both peracetic acid and hydrogen peroxide in a mixed solution.

A High-Throughput Microtiter Plate Based Method for the Determination of Peracetic Acid and Hydrogen Peroxide

PubMed Central

Putt, Karson S.; Pugh, Randall B.

2013-01-01

Peracetic acid is gaining usage in numerous industries who have found a myriad of uses for its antimicrobial activity. However, rapid high throughput quantitation methods for peracetic acid and hydrogen peroxide are lacking. Herein, we describe the development of a high-throughput microtiter plate based assay based upon the well known and trusted titration chemical reactions. The adaptation of these titration chemistries to rapid plate based absorbance methods for the sequential determination of hydrogen peroxide specifically and the total amount of peroxides present in solution are described. The results of these methods were compared to those of a standard titration and found to be in good agreement. Additionally, the utility of the developed method is demonstrated through the generation of degradation curves of both peracetic acid and hydrogen peroxide in a mixed solution. PMID:24260173

Applications of pathology-assisted image analysis of immunohistochemistry-based biomarkers in oncology.

PubMed

Shinde, V; Burke, K E; Chakravarty, A; Fleming, M; McDonald, A A; Berger, A; Ecsedy, J; Blakemore, S J; Tirrell, S M; Bowman, D

2014-01-01

Immunohistochemistry-based biomarkers are commonly used to understand target inhibition in key cancer pathways in preclinical models and clinical studies. Automated slide-scanning and advanced high-throughput image analysis software technologies have evolved into a routine methodology for quantitative analysis of immunohistochemistry-based biomarkers. Alongside the traditional pathology H-score based on physical slides, the pathology world is welcoming digital pathology and advanced quantitative image analysis, which have enabled tissue- and cellular-level analysis. An automated workflow was implemented that includes automated staining, slide-scanning, and image analysis methodologies to explore biomarkers involved in 2 cancer targets: Aurora A and NEDD8-activating enzyme (NAE). The 2 workflows highlight the evolution of our immunohistochemistry laboratory and the different needs and requirements of each biological assay. Skin biopsies obtained from MLN8237 (Aurora A inhibitor) phase 1 clinical trials were evaluated for mitotic and apoptotic index, while mitotic index and defects in chromosome alignment and spindles were assessed in tumor biopsies to demonstrate Aurora A inhibition. Additionally, in both preclinical xenograft models and an acute myeloid leukemia phase 1 trial of the NAE inhibitor MLN4924, development of a novel image algorithm enabled measurement of downstream pathway modulation upon NAE inhibition. In the highlighted studies, developing a biomarker strategy based on automated image analysis solutions enabled project teams to confirm target and pathway inhibition and understand downstream outcomes of target inhibition with increased throughput and quantitative accuracy. These case studies demonstrate a strategy that combines a pathologist's expertise with automated image analysis to support oncology drug discovery and development programs.

Reconstructing the regulatory circuit of cell fate determination in yeast mating response.

PubMed

Shao, Bin; Yuan, Haiyu; Zhang, Rongfei; Wang, Xuan; Zhang, Shuwen; Ouyang, Qi; Hao, Nan; Luo, Chunxiong

2017-07-01

Massive technological advances enabled high-throughput measurements of proteomic changes in biological processes. However, retrieving biological insights from large-scale protein dynamics data remains a challenging task. Here we used the mating differentiation in yeast Saccharomyces cerevisiae as a model and developed integrated experimental and computational approaches to analyze the proteomic dynamics during the process of cell fate determination. When exposed to a high dose of mating pheromone, the yeast cell undergoes growth arrest and forms a shmoo-like morphology; however, at intermediate doses, chemotropic elongated growth is initialized. To understand the gene regulatory networks that control this differentiation switch, we employed a high-throughput microfluidic imaging system that allows real-time and simultaneous measurements of cell growth and protein expression. Using kinetic modeling of protein dynamics, we classified the stimulus-dependent changes in protein abundance into two sources: global changes due to physiological alterations and gene-specific changes. A quantitative framework was proposed to decouple gene-specific regulatory modes from the growth-dependent global modulation of protein abundance. Based on the temporal patterns of gene-specific regulation, we established the network architectures underlying distinct cell fates using a reverse engineering method and uncovered the dose-dependent rewiring of gene regulatory network during mating differentiation. Furthermore, our results suggested a potential crosstalk between the pheromone response pathway and the target of rapamycin (TOR)-regulated ribosomal biogenesis pathway, which might underlie a cell differentiation switch in yeast mating response. In summary, our modeling approach addresses the distinct impacts of the global and gene-specific regulation on the control of protein dynamics and provides new insights into the mechanisms of cell fate determination. We anticipate that our integrated experimental and modeling strategies could be widely applicable to other biological systems.

Automated imaging system for single molecules

DOEpatents

Schwartz, David Charles; Runnheim, Rodney; Forrest, Daniel

2012-09-18

There is provided a high throughput automated single molecule image collection and processing system that requires minimal initial user input. The unique features embodied in the present disclosure allow automated collection and initial processing of optical images of single molecules and their assemblies. Correct focus may be automatically maintained while images are collected. Uneven illumination in fluorescence microscopy is accounted for, and an overall robust imaging operation is provided yielding individual images prepared for further processing in external systems. Embodiments described herein are useful in studies of any macromolecules such as DNA, RNA, peptides and proteins. The automated image collection and processing system and method of same may be implemented and deployed over a computer network, and may be ergonomically optimized to facilitate user interaction.

Trypsin and MALDI matrix pre-coated targets simplify sample preparation for mapping proteomic distributions within biological tissues by imaging mass spectrometry

PubMed Central

Zubair, Faizan; Laibinis, Paul E.; Swisher, William G.; Yang, Junhai; Spraggins, Jeffrey M.; Norris, Jeremy L.; Caprioli, Richard M.

2017-01-01

Prefabricated surfaces containing α-cyano-4-hydroxycinnamic acid and trypsin have been developed to facilitate enzymatic digestion of endogenous tissue proteins prior to matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS). Tissue sections are placed onto slides that were previously coated with α-cyano-4-hydroxycinnamic acid and trypsin. After incubation to promote enzymatic digestion, the tissue is analyzed by MALDI IMS to determine the spatial distribution of the tryptic fragments. The peptides detected in the MALDI IMS dataset were identified by Liquid chromatography-tandem mass spectrometry/mass spectrometry. Protein identification was further confirmed by correlating the localization of unique tryptic fragments originating from common parent proteins. Using this procedure, proteins with molecular weights as large as 300 kDa were identified and their distributions were imaged in sections of rat brain. In particular, large proteins such as myristoylated alanine-rich C-kinase substrate (29.8 kDa) and spectrin alpha chain, non-erythrocytic 1 (284 kDa) were detected that are not observed without trypsin. The pre-coated targets simplify workflow and increase sample throughput by decreasing the sample preparation time. Further, the approach allows imaging at higher spatial resolution compared with robotic spotters that apply one drop at a time. PMID:27676701

Discriminant Analysis of Defective and Non-Defective Field Pea (Pisum sativum L.) into Broad Market Grades Based on Digital Image Features.

PubMed

McDonald, Linda S; Panozzo, Joseph F; Salisbury, Phillip A; Ford, Rebecca

2016-01-01

Field peas (Pisum sativum L.) are generally traded based on seed appearance, which subjectively defines broad market-grades. In this study, we developed an objective Linear Discriminant Analysis (LDA) model to classify market grades of field peas based on seed colour, shape and size traits extracted from digital images. Seeds were imaged in a high-throughput system consisting of a camera and laser positioned over a conveyor belt. Six colour intensity digital images were captured (under 405, 470, 530, 590, 660 and 850nm light) for each seed, and surface height was measured at each pixel by laser. Colour, shape and size traits were compiled across all seed in each sample to determine the median trait values. Defective and non-defective seed samples were used to calibrate and validate the model. Colour components were sufficient to correctly classify all non-defective seed samples into correct market grades. Defective samples required a combination of colour, shape and size traits to achieve 87% and 77% accuracy in market grade classification of calibration and validation sample-sets respectively. Following these results, we used the same colour, shape and size traits to develop an LDA model which correctly classified over 97% of all validation samples as defective or non-defective.

Indexing amyloid peptide diffraction from serial femtosecond crystallography: new algorithms for sparse patterns

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brewster, Aaron S.; Sawaya, Michael R.; University of California, Los Angeles, CA 90095-1570

2015-02-01

Special methods are required to interpret sparse diffraction patterns collected from peptide crystals at X-ray free-electron lasers. Bragg spots can be indexed from composite-image powder rings, with crystal orientations then deduced from a very limited number of spot positions. Still diffraction patterns from peptide nanocrystals with small unit cells are challenging to index using conventional methods owing to the limited number of spots and the lack of crystal orientation information for individual images. New indexing algorithms have been developed as part of the Computational Crystallography Toolbox (cctbx) to overcome these challenges. Accurate unit-cell information derived from an aggregate data setmore » from thousands of diffraction patterns can be used to determine a crystal orientation matrix for individual images with as few as five reflections. These algorithms are potentially applicable not only to amyloid peptides but also to any set of diffraction patterns with sparse properties, such as low-resolution virus structures or high-throughput screening of still images captured by raster-scanning at synchrotron sources. As a proof of concept for this technique, successful integration of X-ray free-electron laser (XFEL) data to 2.5 Å resolution for the amyloid segment GNNQQNY from the Sup35 yeast prion is presented.« less

Discriminant Analysis of Defective and Non-Defective Field Pea (Pisum sativum L.) into Broad Market Grades Based on Digital Image Features

PubMed Central

McDonald, Linda S.; Panozzo, Joseph F.; Salisbury, Phillip A.; Ford, Rebecca

2016-01-01

Field peas (Pisum sativum L.) are generally traded based on seed appearance, which subjectively defines broad market-grades. In this study, we developed an objective Linear Discriminant Analysis (LDA) model to classify market grades of field peas based on seed colour, shape and size traits extracted from digital images. Seeds were imaged in a high-throughput system consisting of a camera and laser positioned over a conveyor belt. Six colour intensity digital images were captured (under 405, 470, 530, 590, 660 and 850nm light) for each seed, and surface height was measured at each pixel by laser. Colour, shape and size traits were compiled across all seed in each sample to determine the median trait values. Defective and non-defective seed samples were used to calibrate and validate the model. Colour components were sufficient to correctly classify all non-defective seed samples into correct market grades. Defective samples required a combination of colour, shape and size traits to achieve 87% and 77% accuracy in market grade classification of calibration and validation sample-sets respectively. Following these results, we used the same colour, shape and size traits to develop an LDA model which correctly classified over 97% of all validation samples as defective or non-defective. PMID:27176469

Advancing ovarian folliculometry with selective plane illumination microscopy

NASA Astrophysics Data System (ADS)

Lin, Hsiao-Chun Amy; Dutta, Rahul; Mandal, Subhamoy; Kind, Alexander; Schnieke, Angelika; Razansky, Daniel

2016-12-01

Determination of ovarian status and follicle monitoring are common methods of diagnosing female infertility. We evaluated the suitability of selective plane illumination microscopy (SPIM) for the study of ovarian follicles. The large field of view and fast acquisition speed of our SPIM system enables rendering of volumetric image stacks from intact whole porcine ovarian follicles, clearly visualizing follicular features including follicle volume and average diameter (70 μm-2.5 mm), their spherical asymmetry parameters, size of developing cumulus oophorus complexes (40 μm-110 μm), and follicular wall thickness (90 μm-120 μm). Follicles at all developmental stages were identified. A distribution of the theca thickness was measured for each follicle, and a relationship between these distributions and the stages of follicular development was discerned. The ability of the system to non-destructively generate sub-cellular resolution 3D images of developing follicles, with excellent image contrast and high throughput capacity compared to conventional histology, suggests that it can be used to monitor follicular development and identify structural abnormalities indicative of ovarian ailments. Accurate folliculometric measurements provided by SPIM images can immensely help the understanding of ovarian physiology and provide important information for the proper management of ovarian diseases.

International Ultraviolet Explorer Final Archive

NASA Technical Reports Server (NTRS)

1997-01-01

CSC processed IUE images through the Final Archive Data Processing System. Raw images were obtained from both NDADS and the IUEGTC optical disk platters for processing on the Alpha cluster, and from the IUEGTC optical disk platters for DECstation processing. Input parameters were obtained from the IUE database. Backup tapes of data to send to VILSPA were routinely made on the Alpha cluster. IPC handled more than 263 requests for priority NEWSIPS processing during the contract. Staff members also answered various questions and requests for information and sent copies of IUE documents to requesters. CSC implemented new processing capabilities into the NEWSIPS processing systems as they became available. In addition, steps were taken to improve efficiency and throughput whenever possible. The node TORTE was reconfigured as the I/O server for Alpha processing in May. The number of Alpha nodes used for the NEWSIPS processing queue was increased to a maximum of six in measured fashion in order to understand the dependence of throughput on the number of nodes and to be able to recognize when a point of diminishing returns was reached. With Project approval, generation of the VD FITS files was dropped in July. This action not only saved processing time but, even more significantly, also reduced the archive storage media requirements, and the time required to perform the archiving, drastically. The throughput of images verified through CDIVS and processed through NEWSIPS for the contract period is summarized below. The number of images of a given dispersion type and camera that were processed in any given month reflects several factors, including the availability of the required NEWSIPS software system, the availability of the corresponding required calibrations (e.g., the LWR high-dispersion ripple correction and absolute calibration), and the occurrence of reprocessing efforts such as that conducted to incorporate the updated SWP sensitivity-degradation correction in May.

Directed molecular evolution to design advanced red fluorescent proteins.

PubMed

Subach, Fedor V; Piatkevich, Kiryl D; Verkhusha, Vladislav V

2011-11-29

Fluorescent proteins have become indispensable imaging tools for biomedical research. Continuing progress in fluorescence imaging, however, requires probes with additional colors and properties optimized for emerging techniques. Here we summarize strategies for development of red-shifted fluorescent proteins. We discuss possibilities for knowledge-based rational design based on the photochemistry of fluorescent proteins and the position of the chromophore in protein structure. We consider advances in library design by mutagenesis, protein expression systems and instrumentation for high-throughput screening that should yield improved fluorescent proteins for advanced imaging applications.

Reflective optical imaging systems with balanced distortion

DOEpatents

Hudyma, Russell M.

2001-01-01

Optical systems compatible with extreme ultraviolet radiation comprising four reflective elements for projecting a mask image onto a substrate are described. The four optical elements comprise, in order from object to image, convex, concave, convex and concave mirrors. The optical systems are particularly suited for step and scan lithography methods. The invention enables the use of larger slit dimensions associated with ring field scanning optics, improves wafer throughput, and allows higher semiconductor device density. The inventive optical systems are characterized by reduced dynamic distortion because the static distortion is balanced across the slit width.

High-throughput Analysis of Large Microscopy Image Datasets on CPU-GPU Cluster Platforms

PubMed Central

Teodoro, George; Pan, Tony; Kurc, Tahsin M.; Kong, Jun; Cooper, Lee A. D.; Podhorszki, Norbert; Klasky, Scott; Saltz, Joel H.

2014-01-01

Analysis of large pathology image datasets offers significant opportunities for the investigation of disease morphology, but the resource requirements of analysis pipelines limit the scale of such studies. Motivated by a brain cancer study, we propose and evaluate a parallel image analysis application pipeline for high throughput computation of large datasets of high resolution pathology tissue images on distributed CPU-GPU platforms. To achieve efficient execution on these hybrid systems, we have built runtime support that allows us to express the cancer image analysis application as a hierarchical data processing pipeline. The application is implemented as a coarse-grain pipeline of stages, where each stage may be further partitioned into another pipeline of fine-grain operations. The fine-grain operations are efficiently managed and scheduled for computation on CPUs and GPUs using performance aware scheduling techniques along with several optimizations, including architecture aware process placement, data locality conscious task assignment, data prefetching, and asynchronous data copy. These optimizations are employed to maximize the utilization of the aggregate computing power of CPUs and GPUs and minimize data copy overheads. Our experimental evaluation shows that the cooperative use of CPUs and GPUs achieves significant improvements on top of GPU-only versions (up to 1.6×) and that the execution of the application as a set of fine-grain operations provides more opportunities for runtime optimizations and attains better performance than coarser-grain, monolithic implementations used in other works. An implementation of the cancer image analysis pipeline using the runtime support was able to process an image dataset consisting of 36,848 4Kx4K-pixel image tiles (about 1.8TB uncompressed) in less than 4 minutes (150 tiles/second) on 100 nodes of a state-of-the-art hybrid cluster system. PMID:25419546

A high throughput array microscope for the mechanical characterization of biomaterials

NASA Astrophysics Data System (ADS)

Cribb, Jeremy; Osborne, Lukas D.; Hsiao, Joe Ping-Lin; Vicci, Leandra; Meshram, Alok; O'Brien, E. Tim; Spero, Richard Chasen; Taylor, Russell; Superfine, Richard

2015-02-01

In the last decade, the emergence of high throughput screening has enabled the development of novel drug therapies and elucidated many complex cellular processes. Concurrently, the mechanobiology community has developed tools and methods to show that the dysregulation of biophysical properties and the biochemical mechanisms controlling those properties contribute significantly to many human diseases. Despite these advances, a complete understanding of the connection between biomechanics and disease will require advances in instrumentation that enable parallelized, high throughput assays capable of probing complex signaling pathways, studying biology in physiologically relevant conditions, and capturing specimen and mechanical heterogeneity. Traditional biophysical instruments are unable to meet this need. To address the challenge of large-scale, parallelized biophysical measurements, we have developed an automated array high-throughput microscope system that utilizes passive microbead diffusion to characterize mechanical properties of biomaterials. The instrument is capable of acquiring data on twelve-channels simultaneously, where each channel in the system can independently drive two-channel fluorescence imaging at up to 50 frames per second. We employ this system to measure the concentration-dependent apparent viscosity of hyaluronan, an essential polymer found in connective tissue and whose expression has been implicated in cancer progression.

Projection Reduction Exposure with Variable Axis Immersion Lenses (PREVAIL)-A High Throughput E-Beam Projection Approach for Next Generation Lithography

NASA Astrophysics Data System (ADS)

Pfeiffer, Hans

1999-12-01

Projection reduction exposure with variable axis immersion lenses (PREVAIL) represents the high throughput e-beam projection approach to next generation lithography (NGL), which IBM is pursuing in cooperation with Nikon Corporation as an alliance partner. This paper discusses the challenges and accomplishments of the PREVAIL project. The supreme challenge facing all e-beam lithography approaches has been and still is throughput. Since the throughput of e-beam projection systems is severely limited by the available optical field size, the key to success is the ability to overcome this limitation. The PREVAIL technique overcomes field-limiting off-axis aberrations through the use of variable axis lenses, which electronically shift the optical axis simultaneously with the deflected beam, so that the beam effectively remains on axis. The resist images obtained with the proof-of-concept (POC) system demonstrate that PREVAIL effectively eliminates off-axis aberrations affecting both the resolution and placement accuracy of pixels. As part of the POC system a high emittance gun has been developed to provide uniform illumination of the patterned subfield, and to fill the large numerical aperture projection optics designed to significantly reduce beam blur caused by Coulombinteraction.

THE RABIT: A RAPID AUTOMATED BIODOSIMETRY TOOL FOR RADIOLOGICAL TRIAGE

PubMed Central

Garty, Guy; Chen, Youhua; Salerno, Alessio; Turner, Helen; Zhang, Jian; Lyulko, Oleksandra; Bertucci, Antonella; Xu, Yanping; Wang, Hongliang; Simaan, Nabil; Randers-Pehrson, Gerhard; Yao, Y. Lawrence; Amundson, Sally A.; Brenner, David J.

2010-01-01

In response to the recognized need for high throughput biodosimetry methods for use after large scale radiological events, a logical approach is complete automation of standard biodosimetric assays that are currently performed manually. We describe progress to date on the RABIT (Rapid Automated BIodosimetry Tool), designed to score micronuclei or γ-H2AX fluorescence in lymphocytes derived from a single drop of blood from a fingerstick. The RABIT system is designed to be completely automated, from the input of the capillary blood sample into the machine, to the output of a dose estimate. Improvements in throughput are achieved through use of a single drop of blood, optimization of the biological protocols for in-situ analysis in multi-well plates, implementation of robotic plate and liquid handling, and new developments in high-speed imaging. Automating well-established bioassays represents a promising approach to high-throughput radiation biodosimetry, both because high throughputs can be achieved, but also because the time to deployment is potentially much shorter than for a new biological assay. Here we describe the development of each of the individual modules of the RABIT system, and show preliminary data from key modules. Ongoing is system integration, followed by calibration and validation. PMID:20065685

A Fully Automated High-Throughput Flow Cytometry Screening System Enabling Phenotypic Drug Discovery.

PubMed

Joslin, John; Gilligan, James; Anderson, Paul; Garcia, Catherine; Sharif, Orzala; Hampton, Janice; Cohen, Steven; King, Miranda; Zhou, Bin; Jiang, Shumei; Trussell, Christopher; Dunn, Robert; Fathman, John W; Snead, Jennifer L; Boitano, Anthony E; Nguyen, Tommy; Conner, Michael; Cooke, Mike; Harris, Jennifer; Ainscow, Ed; Zhou, Yingyao; Shaw, Chris; Sipes, Dan; Mainquist, James; Lesley, Scott

2018-05-01

The goal of high-throughput screening is to enable screening of compound libraries in an automated manner to identify quality starting points for optimization. This often involves screening a large diversity of compounds in an assay that preserves a connection to the disease pathology. Phenotypic screening is a powerful tool for drug identification, in that assays can be run without prior understanding of the target and with primary cells that closely mimic the therapeutic setting. Advanced automation and high-content imaging have enabled many complex assays, but these are still relatively slow and low throughput. To address this limitation, we have developed an automated workflow that is dedicated to processing complex phenotypic assays for flow cytometry. The system can achieve a throughput of 50,000 wells per day, resulting in a fully automated platform that enables robust phenotypic drug discovery. Over the past 5 years, this screening system has been used for a variety of drug discovery programs, across many disease areas, with many molecules advancing quickly into preclinical development and into the clinic. This report will highlight a diversity of approaches that automated flow cytometry has enabled for phenotypic drug discovery.

Modified Pressure System for Imaging Egg Cracks

USDA-ARS?s Scientific Manuscript database

One aspect of grading table eggs is shell checks or cracks. Currently, USDA voluntary regulations require that humans grade a representative sample of all eggs processed. However, as processing plants and packing facilities continue to increase their volume and throughput, human graders are having ...

Modified Pressure System for Imaging Egg Cracks

USDA-ARS?s Scientific Manuscript database

Abstract One aspect of grading table eggs is shell checks or cracks. Currently, USDA voluntary regulations require that humans grade a representative sample of all eggs processed. However, as processing plants and packing facilities continue to increase their volume and throughput, human graders a...

High-throughput crystal-optimization strategies in the South Paris Yeast Structural Genomics Project: one size fits all?

PubMed

Leulliot, Nicolas; Trésaugues, Lionel; Bremang, Michael; Sorel, Isabelle; Ulryck, Nathalie; Graille, Marc; Aboulfath, Ilham; Poupon, Anne; Liger, Dominique; Quevillon-Cheruel, Sophie; Janin, Joël; van Tilbeurgh, Herman

2005-06-01

Crystallization has long been regarded as one of the major bottlenecks in high-throughput structural determination by X-ray crystallography. Structural genomics projects have addressed this issue by using robots to set up automated crystal screens using nanodrop technology. This has moved the bottleneck from obtaining the first crystal hit to obtaining diffraction-quality crystals, as crystal optimization is a notoriously slow process that is difficult to automatize. This article describes the high-throughput optimization strategies used in the Yeast Structural Genomics project, with selected successful examples.

On the importance of image formation optics in the design of infrared spectroscopic imaging systems

PubMed Central

Mayerich, David; van Dijk, Thomas; Walsh, Michael; Schulmerich, Matthew; Carney, P. Scott

2014-01-01

Infrared spectroscopic imaging provides micron-scale spatial resolution with molecular contrast. While recent work demonstrates that sample morphology affects the recorded spectrum, considerably less attention has been focused on the effects of the optics, including the condenser and objective. This analysis is extremely important, since it will be possible to understand effects on recorded data and provides insight for reducing optical effects through rigorous microscope design. Here, we present a theoretical description and experimental results that demonstrate the effects of commonly-employed cassegranian optics on recorded spectra. We first combine an explicit model of image formation and a method for quantifying and visualizing the deviations in recorded spectra as a function of microscope optics. We then verify these simulations with measurements obtained from spatially heterogeneous samples. The deviation of the computed spectrum from the ideal case is quantified via a map which we call a deviation map. The deviation map is obtained as a function of optical elements by systematic simulations. Examination of deviation maps demonstrates that the optimal optical configuration for minimal deviation is contrary to prevailing practice in which throughput is maximized for an instrument without a sample. This report should be helpful for understanding recorded spectra as a function of the optics, the analytical limits of recorded data determined by the optical design, and potential routes for optimization of imaging systems. PMID:24936526

On the importance of image formation optics in the design of infrared spectroscopic imaging systems.

PubMed

Mayerich, David; van Dijk, Thomas; Walsh, Michael J; Schulmerich, Matthew V; Carney, P Scott; Bhargava, Rohit

2014-08-21

Infrared spectroscopic imaging provides micron-scale spatial resolution with molecular contrast. While recent work demonstrates that sample morphology affects the recorded spectrum, considerably less attention has been focused on the effects of the optics, including the condenser and objective. This analysis is extremely important, since it will be possible to understand effects on recorded data and provides insight for reducing optical effects through rigorous microscope design. Here, we present a theoretical description and experimental results that demonstrate the effects of commonly-employed cassegranian optics on recorded spectra. We first combine an explicit model of image formation and a method for quantifying and visualizing the deviations in recorded spectra as a function of microscope optics. We then verify these simulations with measurements obtained from spatially heterogeneous samples. The deviation of the computed spectrum from the ideal case is quantified via a map which we call a deviation map. The deviation map is obtained as a function of optical elements by systematic simulations. Examination of deviation maps demonstrates that the optimal optical configuration for minimal deviation is contrary to prevailing practice in which throughput is maximized for an instrument without a sample. This report should be helpful for understanding recorded spectra as a function of the optics, the analytical limits of recorded data determined by the optical design, and potential routes for optimization of imaging systems.

Temporal and spatial resolution required for imaging myocardial function

NASA Astrophysics Data System (ADS)

Eusemann, Christian D.; Robb, Richard A.

2004-05-01

4-D functional analysis of myocardial mechanics is an area of significant interest and research in cardiology and vascular/interventional radiology. Current multidimensional analysis is limited by insufficient temporal resolution of x-ray and magnetic resonance based techniques, but recent improvements in system design holds hope for faster and higher resolution scans to improve images of moving structures allowing more accurate functional studies, such as in the heart. This paper provides a basis for the requisite temporal and spatial resolution for useful imaging during individual segments of the cardiac cycle. Multiple sample rates during systole and diastole are compared to determine an adequate sample frequency to reduce regional myocardial tracking errors. Concurrently, out-of-plane resolution has to be sufficiently high to minimize partial volume effect. Temporal resolution and out-of-plane spatial resolution are related factors that must be considered together. The data used for this study is a DSR dynamic volume image dataset with high temporal and spatial resolution using implanted fiducial markers to track myocardial motion. The results of this study suggest a reduced exposure and scan time for x-ray and magnetic resonance imaging methods, since a lower sample rate during systole is sufficient, whereas the period of rapid filling during diastole requires higher sampling. This could potentially reduce the cost of these procedures and allow higher patient throughput.

Ultra-high throughput detection of single cell β-galactosidase activity in droplets using micro-optical lens array

NASA Astrophysics Data System (ADS)

Lim, Jiseok; Vrignon, Jérémy; Gruner, Philipp; Karamitros, Christos S.; Konrad, Manfred; Baret, Jean-Christophe

2013-11-01

We demonstrate the use of a hybrid microfluidic-micro-optical system for the screening of enzymatic activity at the single cell level. Escherichia coli β-galactosidase activity is revealed by a fluorogenic assay in 100 pl droplets. Individual droplets containing cells are screened by measuring their fluorescence signal using a high-speed camera. The measurement is parallelized over 100 channels equipped with microlenses and analyzed by image processing. A reinjection rate of 1 ml of emulsion per minute was reached corresponding to more than 105 droplets per second, an analytical throughput larger than those obtained using flow cytometry.

Quantitative characterization of conformational-specific protein-DNA binding using a dual-spectral interferometric imaging biosensor.

PubMed

Zhang, Xirui; Daaboul, George G; Spuhler, Philipp S; Dröge, Peter; Ünlü, M Selim

2016-03-14

DNA-binding proteins play crucial roles in the maintenance and functions of the genome and yet, their specific binding mechanisms are not fully understood. Recently, it was discovered that DNA-binding proteins recognize specific binding sites to carry out their functions through an indirect readout mechanism by recognizing and capturing DNA conformational flexibility and deformation. High-throughput DNA microarray-based methods that provide large-scale protein-DNA binding information have shown effective and comprehensive analysis of protein-DNA binding affinities, but do not provide information of DNA conformational changes in specific protein-DNA complexes. Building on the high-throughput capability of DNA microarrays, we demonstrate a quantitative approach that simultaneously measures the amount of protein binding to DNA and nanometer-scale DNA conformational change induced by protein binding in a microarray format. Both measurements rely on spectral interferometry on a layered substrate using a single optical instrument in two distinct modalities. In the first modality, we quantitate the amount of binding of protein to surface-immobilized DNA in each DNA spot using a label-free spectral reflectivity technique that accurately measures the surface densities of protein and DNA accumulated on the substrate. In the second modality, for each DNA spot, we simultaneously measure DNA conformational change using a fluorescence vertical sectioning technique that determines average axial height of fluorophores tagged to specific nucleotides of the surface-immobilized DNA. The approach presented in this paper, when combined with current high-throughput DNA microarray-based technologies, has the potential to serve as a rapid and simple method for quantitative and large-scale characterization of conformational specific protein-DNA interactions.

In vivo Monitoring of Transcriptional Dynamics After Lower-Limb Muscle Injury Enables Quantitative Classification of Healing

PubMed Central

Aguilar, Carlos A.; Shcherbina, Anna; Ricke, Darrell O.; Pop, Ramona; Carrigan, Christopher T.; Gifford, Casey A.; Urso, Maria L.; Kottke, Melissa A.; Meissner, Alexander

2015-01-01

Traumatic lower-limb musculoskeletal injuries are pervasive amongst athletes and the military and typically an individual returns to activity prior to fully healing, increasing a predisposition for additional injuries and chronic pain. Monitoring healing progression after a musculoskeletal injury typically involves different types of imaging but these approaches suffer from several disadvantages. Isolating and profiling transcripts from the injured site would abrogate these shortcomings and provide enumerative insights into the regenerative potential of an individual’s muscle after injury. In this study, a traumatic injury was administered to a mouse model and healing progression was examined from 3 hours to 1 month using high-throughput RNA-Sequencing (RNA-Seq). Comprehensive dissection of the genome-wide datasets revealed the injured site to be a dynamic, heterogeneous environment composed of multiple cell types and thousands of genes undergoing significant expression changes in highly regulated networks. Four independent approaches were used to determine the set of genes, isoforms, and genetic pathways most characteristic of different time points post-injury and two novel approaches were developed to classify injured tissues at different time points. These results highlight the possibility to quantitatively track healing progression in situ via transcript profiling using high- throughput sequencing. PMID:26381351

Maximization Network Throughput Based on Improved Genetic Algorithm and Network Coding for Optical Multicast Networks

NASA Astrophysics Data System (ADS)

Wei, Chengying; Xiong, Cuilian; Liu, Huanlin

2017-12-01

Maximal multicast stream algorithm based on network coding (NC) can improve the network's throughput for wavelength-division multiplexing (WDM) networks, which however is far less than the network's maximal throughput in terms of theory. And the existing multicast stream algorithms do not give the information distribution pattern and routing in the meantime. In the paper, an improved genetic algorithm is brought forward to maximize the optical multicast throughput by NC and to determine the multicast stream distribution by hybrid chromosomes construction for multicast with single source and multiple destinations. The proposed hybrid chromosomes are constructed by the binary chromosomes and integer chromosomes, while the binary chromosomes represent optical multicast routing and the integer chromosomes indicate the multicast stream distribution. A fitness function is designed to guarantee that each destination can receive the maximum number of decoding multicast streams. The simulation results showed that the proposed method is far superior over the typical maximal multicast stream algorithms based on NC in terms of network throughput in WDM networks.

High-throughput analysis using non-depletive SPME: challenges and applications to the determination of free and total concentrations in small sample volumes.

PubMed

Boyacı, Ezel; Bojko, Barbara; Reyes-Garcés, Nathaly; Poole, Justen J; Gómez-Ríos, Germán Augusto; Teixeira, Alexandre; Nicol, Beate; Pawliszyn, Janusz

2018-01-18

In vitro high-throughput non-depletive quantitation of chemicals in biofluids is of growing interest in many areas. Some of the challenges facing researchers include the limited volume of biofluids, rapid and high-throughput sampling requirements, and the lack of reliable methods. Coupled to the above, growing interest in the monitoring of kinetics and dynamics of miniaturized biosystems has spurred the demand for development of novel and revolutionary methodologies for analysis of biofluids. The applicability of solid-phase microextraction (SPME) is investigated as a potential technology to fulfill the aforementioned requirements. As analytes with sufficient diversity in their physicochemical features, nicotine, N,N-Diethyl-meta-toluamide, and diclofenac were selected as test compounds for the study. The objective was to develop methodologies that would allow repeated non-depletive sampling from 96-well plates, using 100 µL of sample. Initially, thin film-SPME was investigated. Results revealed substantial depletion and consequent disruption in the system. Therefore, new ultra-thin coated fibers were developed. The applicability of this device to the described sampling scenario was tested by determining the protein binding of the analytes. Results showed good agreement with rapid equilibrium dialysis. The presented method allows high-throughput analysis using small volumes, enabling fast reliable free and total concentration determinations without disruption of system equilibrium.

Fast assembling of neuron fragments in serial 3D sections.

PubMed

Chen, Hanbo; Iascone, Daniel Maxim; da Costa, Nuno Maçarico; Lein, Ed S; Liu, Tianming; Peng, Hanchuan

2017-09-01

Reconstructing neurons from 3D image-stacks of serial sections of thick brain tissue is very time-consuming and often becomes a bottleneck in high-throughput brain mapping projects. We developed NeuronStitcher, a software suite for stitching non-overlapping neuron fragments reconstructed in serial 3D image sections. With its efficient algorithm and user-friendly interface, NeuronStitcher has been used successfully to reconstruct very large and complex human and mouse neurons.

Recent advances in MRI technology: Implications for image quality and patient safety

PubMed Central

Sobol, Wlad T.

2012-01-01

Recent advances in MRI technology are presented, with emphasis on how this new technology impacts clinical operations (better image quality, faster exam times, and improved throughput). In addition, implications for patient safety are discussed with emphasis on the risk of patient injury due to either high local specific absorption rate (SAR) or large cumulative energy doses delivered during long exam times. Patient comfort issues are examined as well. PMID:23961024

Applications of Molecular Imaging

PubMed Central

Galbán, Craig; Galbán, Stefanie; Van Dort, Marcian; Luker, Gary D.; Bhojani, Mahaveer S.; Rehemtualla, Alnawaz; Ross, Brian D.

2015-01-01

Today molecular imaging technologies play a central role in clinical oncology. The use of imaging techniques in early cancer detection, treatment response and new therapy development is steadily growing and has already significantly impacted clinical management of cancer. In this chapter we will overview three different molecular imaging technologies used for the understanding of disease biomarkers, drug development, or monitoring therapeutic outcome. They are (1) optical imaging (bioluminescence and fluorescence imaging) (2) magnetic resonance imaging (MRI), and (3) nuclear imaging (e.g, single photon emission computed tomography (SPECT) and positron emission tomography (PET)). We will review the use of molecular reporters of biological processes (e.g. apoptosis and protein kinase activity) for high throughput drug screening and new cancer therapies, diffusion MRI as a biomarker for early treatment response and PET and SPECT radioligands in oncology. PMID:21075334

Radiomics: Extracting more information from medical images using advanced feature analysis

PubMed Central

Lambin, Philippe; Rios-Velazquez, Emmanuel; Leijenaar, Ralph; Carvalho, Sara; van Stiphout, Ruud G.P.M.; Granton, Patrick; Zegers, Catharina M.L.; Gillies, Robert; Boellard, Ronald; Dekker, André; Aerts, Hugo J.W.L.

2015-01-01

Solid cancers are spatially and temporally heterogeneous. This limits the use of invasive biopsy based molecular assays but gives huge potential for medical imaging, which has the ability to capture intra-tumoural heterogeneity in a non-invasive way. During the past decades, medical imaging innovations with new hardware, new imaging agents and standardised protocols, allows the field to move towards quantitative imaging. Therefore, also the development of automated and reproducible analysis methodologies to extract more information from image-based features is a requirement. Radiomics – the high-throughput extraction of large amounts of image features from radiographic images – addresses this problem and is one of the approaches that hold great promises but need further validation in multi-centric settings and in the laboratory. PMID:22257792

Flexible Measurement of Bioluminescent Reporters Using an Automated Longitudinal Luciferase Imaging Gas- and Temperature-optimized Recorder (ALLIGATOR).

PubMed

Crosby, Priya; Hoyle, Nathaniel P; O'Neill, John S

2017-12-13

Luciferase-based reporters of cellular gene expression are in widespread use for both longitudinal and end-point assays of biological activity. In circadian rhythms research, for example, clock gene fusions with firefly luciferase give rise to robust rhythms in cellular bioluminescence that persist over many days. Technical limitations associated with photomultiplier tubes (PMT) or conventional microscopy-based methods for bioluminescence quantification have typically demanded that cells and tissues be maintained under quite non-physiological conditions during recording, with a trade-off between sensitivity and throughput. Here, we report a refinement of prior methods that allows long-term bioluminescence imaging with high sensitivity and throughput which supports a broad range of culture conditions, including variable gas and humidity control, and that accepts many different tissue culture plates and dishes. This automated longitudinal luciferase imaging gas- and temperature-optimized recorder (ALLIGATOR) also allows the observation of spatial variations in luciferase expression across a cell monolayer or tissue, which cannot readily be observed by traditional methods. We highlight how the ALLIGATOR provides vastly increased flexibility for the detection of luciferase activity when compared with existing methods.

High-throughput sensing and noninvasive imaging of protein nuclear transport by using reconstitution of split Renilla luciferase.

PubMed

Kim, Sung Bae; Ozawa, Takeaki; Watanabe, Shigeaki; Umezawa, Yoshio

2004-08-10

Nucleocytoplasmic trafficking of functional proteins plays a key role in regulating gene expressions in response to extracellular signals. We developed a genetically encoded bioluminescent indicator for monitoring the nuclear trafficking of target proteins in vitro and in vivo. The principle is based on reconstitution of split fragments of Renilla reniformis (Rluc) by protein splicing with a DnaE intein (a catalytic subunit of DNA polymerase III). A target cytosolic protein fused to the N-terminal half of Rluc is expressed in mammalian cells. If the protein translocates into the nucleus, the Rluc moiety meets the C-terminal half of Rluc, and full-length Rluc is reconstituted by protein splicing. We demonstrated quantitative cell-based in vitro sensing of ligand-induced translocation of androgen receptor, which allowed high-throughput screening of exo- and endogenous agonists and antagonists. Furthermore, the indicator enabled noninvasive in vivo imaging of the androgen receptor translocation in the brains of living mice with a charge-coupled device imaging system. These rapid and quantitative analyses in vitro and in vivo provide a wide variety of applications for screening pharmacological or toxicological compounds and testing them in living animals.

Scientific Performance Analysis of the SYZ Telescope Design versus the RC Telescope Design

NASA Astrophysics Data System (ADS)

Ma, Donglin; Cai, Zheng

2018-02-01

Recently, Su et al. propose an innovative design, referred as the “SYZ” design, for China’s new project of a 12 m optical-infrared telescope. The SYZ telescope design consists of three aspheric mirrors with non-zero power, including a relay mirror below the primary mirror. SYZ design yields a good imaging quality and has a relatively flat field curvature at Nasmyth focus. To evaluate the science-compatibility of this three-mirror telescope, in this paper, we thoroughly compare the performance of SYZ design with that of Ritchey–Chrétien (RC) design, a conventional two-mirror telescope design. Further, we propose the Observing Information Throughput (OIT) as a metric for quantitatively evaluating the telescopes’ science performance. We find that although a SYZ telescope yields a superb imaging quality over a large field of view, a two-mirror (RC) telescope design holds a higher overall throughput, a better diffraction-limited imaging quality in the central field of view (FOV < 5‧) which is better for the performance of extreme Adaptive Optics (AO), and a generally better scientific performance with a higher OIT value. D. Ma & Z. Cai contributed equally to this paper.

Ultra-high frequency ultrasound biomicroscopy and high throughput cardiovascular phenotyping in a large scale mouse mutagenesis screen

NASA Astrophysics Data System (ADS)

Liu, Xiaoqin; Francis, Richard; Tobita, Kimimasa; Kim, Andy; Leatherbury, Linda; Lo, Cecilia W.

2013-02-01

Ultrasound biomicroscopy (UBM) is ideally suited for phenotyping fetal mice for congenital heart disease (CHD), as imaging can be carried out noninvasively to provide both hemodynamic and structural information essential for CHD diagnosis. Using the UBM (Vevo 2100; 40Hz) in conjunction with the clinical ultrasound system (Acuson Sequioa C512; 15Hz), we developed a two-step screening protocol to scan thousands fetuses derived from ENU mutagenized pedigrees. A wide spectrum of CHD was detected by the UBM, which were subsequently confirmed with follow-up necropsy and histopathology examination with episcopic fluorescence image capture. CHD observed included outflow anomalies, left/right heart obstructive lesions, septal/valvular defects and cardiac situs anomalies. Meanwhile, various extracardiac defects were found, such as polydactyly, craniofacial defects, exencephaly, omphalocele-cleft palate, most of which were associated with cardiac defects. Our analyses showed the UBM was better at assessing cardiac structure and blood flow profiles, while conventional ultrasound allowed higher throughput low-resolution screening. Our study showed the integration of conventional clinical ultrasound imaging with the UBM for fetal mouse cardiovascular phenotyping can maximize the detection and recovery of CHD mutants.

High-throughput Titration of Luciferase-expressing Recombinant Viruses

PubMed Central

Garcia, Vanessa; Krishnan, Ramya; Davis, Colin; Batenchuk, Cory; Le Boeuf, Fabrice; Abdelbary, Hesham; Diallo, Jean-Simon

2014-01-01

Standard plaque assays to determine infectious viral titers can be time consuming, are not amenable to a high volume of samples, and cannot be done with viruses that do not form plaques. As an alternative to plaque assays, we have developed a high-throughput titration method that allows for the simultaneous titration of a high volume of samples in a single day. This approach involves infection of the samples with a Firefly luciferase tagged virus, transfer of the infected samples onto an appropriate permissive cell line, subsequent addition of luciferin, reading of plates in order to obtain luminescence readings, and finally the conversion from luminescence to viral titers. The assessment of cytotoxicity using a metabolic viability dye can be easily incorporated in the workflow in parallel and provide valuable information in the context of a drug screen. This technique provides a reliable, high-throughput method to determine viral titers as an alternative to a standard plaque assay. PMID:25285536

Temporal Data Set Reduction Based on D-Optimality for Quantitative FLIM-FRET Imaging.

PubMed

Omer, Travis; Intes, Xavier; Hahn, Juergen

2015-01-01

Fluorescence lifetime imaging (FLIM) when paired with Förster resonance energy transfer (FLIM-FRET) enables the monitoring of nanoscale interactions in living biological samples. FLIM-FRET model-based estimation methods allow the quantitative retrieval of parameters such as the quenched (interacting) and unquenched (non-interacting) fractional populations of the donor fluorophore and/or the distance of the interactions. The quantitative accuracy of such model-based approaches is dependent on multiple factors such as signal-to-noise ratio and number of temporal points acquired when sampling the fluorescence decays. For high-throughput or in vivo applications of FLIM-FRET, it is desirable to acquire a limited number of temporal points for fast acquisition times. Yet, it is critical to acquire temporal data sets with sufficient information content to allow for accurate FLIM-FRET parameter estimation. Herein, an optimal experimental design approach based upon sensitivity analysis is presented in order to identify the time points that provide the best quantitative estimates of the parameters for a determined number of temporal sampling points. More specifically, the D-optimality criterion is employed to identify, within a sparse temporal data set, the set of time points leading to optimal estimations of the quenched fractional population of the donor fluorophore. Overall, a reduced set of 10 time points (compared to a typical complete set of 90 time points) was identified to have minimal impact on parameter estimation accuracy (≈5%), with in silico and in vivo experiment validations. This reduction of the number of needed time points by almost an order of magnitude allows the use of FLIM-FRET for certain high-throughput applications which would be infeasible if the entire number of time sampling points were used.

20170308 - Higher Throughput Toxicokinetics to Allow ...

EPA Pesticide Factsheets

As part of "Ongoing EDSP Directions & Activities" I will present CSS research on high throughput toxicokinetics, including in vitro data and models to allow rapid determination of the real world doses that may cause endocrine disruption. This is a presentation as part of the U.S. Environmental Protection Agency – Japan Ministry of the Environment 12th Bilateral Meeting on Endocrine Disruption Test Methods Development.

High content screening of ToxCast compounds using Vala Sciences’ complex cell culturing systems (SOT)

EPA Science Inventory

US EPA’s ToxCast research program evaluates bioactivity for thousands of chemicals utilizing high-throughput screening assays to inform chemical testing decisions. Vala Sciences provides high content, multiplexed assays that utilize quantitative cell-based digital image analysis....

Evaluating chemical safety: ToxCast, Tipping Points and Virtual Tissues (Tamburro Symposium)

EPA Science Inventory

This presentation provides an overview of high-throughput toxicology at the NCCT using high-content imaging and computational models for analyzing chemical safety. In In particular, this work outlines the derivation of toxicological "tipping points" from in vitro concentration- a...

Non-Gaussian Distribution of DNA Barcode Extension In Nanochannels Using High-throughput Imaging

NASA Astrophysics Data System (ADS)

Sheats, Julian; Reinhart, Wesley; Reifenberger, Jeff; Gupta, Damini; Muralidhar, Abhiram; Cao, Han; Dorfman, Kevin

2015-03-01

We present experimental data for the extension of internal segments of highly confined DNA using a high-­throughput experimental setup. Barcode­-labeled E. coli genomic DNA molecules were imaged at a high areal density in square nanochannels with sizes ranging from 40 nm to 51 nm in width. Over 25,000 molecules were used to obtain more than 1,000,000 measurements for genomic distances between 2,500 bp and 100,000 bp. The distribution of extensions has positive excess kurtosis and is skew­ left due to weak backfolding in the channel. As a result, the two Odijk theories for the chain extension and variance bracket the experimental data. We compared to predictions of a harmonic approximation for the confinement free energy and show that it produces a substantial error in the variance. These results suggest an inherent error associated with any statistical analysis of barcoded DNA that relies on harmonic models for chain extension. Present address: Department of Chemical and Biological Engineering, Princeton University.

A data set from flash X-ray imaging of carboxysomes

NASA Astrophysics Data System (ADS)

Hantke, Max F.; Hasse, Dirk; Ekeberg, Tomas; John, Katja; Svenda, Martin; Loh, Duane; Martin, Andrew V.; Timneanu, Nicusor; Larsson, Daniel S. D.; van der Schot, Gijs; Carlsson, Gunilla H.; Ingelman, Margareta; Andreasson, Jakob; Westphal, Daniel; Iwan, Bianca; Uetrecht, Charlotte; Bielecki, Johan; Liang, Mengning; Stellato, Francesco; Deponte, Daniel P.; Bari, Sadia; Hartmann, Robert; Kimmel, Nils; Kirian, Richard A.; Seibert, M. Marvin; Mühlig, Kerstin; Schorb, Sebastian; Ferguson, Ken; Bostedt, Christoph; Carron, Sebastian; Bozek, John D.; Rolles, Daniel; Rudenko, Artem; Foucar, Lutz; Epp, Sascha W.; Chapman, Henry N.; Barty, Anton; Andersson, Inger; Hajdu, Janos; Maia, Filipe R. N. C.

2016-08-01

Ultra-intense femtosecond X-ray pulses from X-ray lasers permit structural studies on single particles and biomolecules without crystals. We present a large data set on inherently heterogeneous, polyhedral carboxysome particles. Carboxysomes are cell organelles that vary in size and facilitate up to 40% of Earth’s carbon fixation by cyanobacteria and certain proteobacteria. Variation in size hinders crystallization. Carboxysomes appear icosahedral in the electron microscope. A protein shell encapsulates a large number of Rubisco molecules in paracrystalline arrays inside the organelle. We used carboxysomes with a mean diameter of 115±26 nm from Halothiobacillus neapolitanus. A new aerosol sample-injector allowed us to record 70,000 low-noise diffraction patterns in 12 min. Every diffraction pattern is a unique structure measurement and high-throughput imaging allows sampling the space of structural variability. The different structures can be separated and phased directly from the diffraction data and open a way for accurate, high-throughput studies on structures and structural heterogeneity in biology and elsewhere.

Repurposing High-Throughput Image Assays Enables Biological Activity Prediction for Drug Discovery.

PubMed

Simm, Jaak; Klambauer, Günter; Arany, Adam; Steijaert, Marvin; Wegner, Jörg Kurt; Gustin, Emmanuel; Chupakhin, Vladimir; Chong, Yolanda T; Vialard, Jorge; Buijnsters, Peter; Velter, Ingrid; Vapirev, Alexander; Singh, Shantanu; Carpenter, Anne E; Wuyts, Roel; Hochreiter, Sepp; Moreau, Yves; Ceulemans, Hugo

2018-05-17

In both academia and the pharmaceutical industry, large-scale assays for drug discovery are expensive and often impractical, particularly for the increasingly important physiologically relevant model systems that require primary cells, organoids, whole organisms, or expensive or rare reagents. We hypothesized that data from a single high-throughput imaging assay can be repurposed to predict the biological activity of compounds in other assays, even those targeting alternate pathways or biological processes. Indeed, quantitative information extracted from a three-channel microscopy-based screen for glucocorticoid receptor translocation was able to predict assay-specific biological activity in two ongoing drug discovery projects. In these projects, repurposing increased hit rates by 50- to 250-fold over that of the initial project assays while increasing the chemical structure diversity of the hits. Our results suggest that data from high-content screens are a rich source of information that can be used to predict and replace customized biological assays. Copyright © 2018 Elsevier Ltd. All rights reserved.

Comprehensive Analysis of Immunological Synapse Phenotypes Using Supported Lipid Bilayers.

PubMed

Valvo, Salvatore; Mayya, Viveka; Seraia, Elena; Afrose, Jehan; Novak-Kotzer, Hila; Ebner, Daniel; Dustin, Michael L

2017-01-01

Supported lipid bilayers (SLB) formed on glass substrates have been a useful tool for study of immune cell signaling since the early 1980s. The mobility of lipid-anchored proteins in the system, first described for antibodies binding to synthetic phospholipid head groups, allows for the measurement of two-dimensional binding reactions and signaling processes in a single imaging plane over time or for fixed samples. The fragility of SLB and the challenges of building and validating individual substrates limit most experimenters to ~10 samples per day, perhaps increasing this few-fold when examining fixed samples. Successful experiments might then require further days to fully analyze. We present methods for automation of many steps in SLB formation, imaging in 96-well glass bottom plates, and analysis that enables >100-fold increase in throughput for fixed samples and wide-field fluorescence. This increased throughput will allow better coverage of relevant parameters and more comprehensive analysis of aspects of the immunological synapse that are well reconstituted by SLB.

Neural network Hilbert transform based filtered backprojection for fast inline x-ray inspection

NASA Astrophysics Data System (ADS)

Janssens, Eline; De Beenhouwer, Jan; Van Dael, Mattias; De Schryver, Thomas; Van Hoorebeke, Luc; Verboven, Pieter; Nicolai, Bart; Sijbers, Jan

2018-03-01

X-ray imaging is an important tool for quality control since it allows to inspect the interior of products in a non-destructive way. Conventional x-ray imaging, however, is slow and expensive. Inline x-ray inspection, on the other hand, can pave the way towards fast and individual quality control, provided that a sufficiently high throughput can be achieved at a minimal cost. To meet these criteria, an inline inspection acquisition geometry is proposed where the object moves and rotates on a conveyor belt while it passes a fixed source and detector. Moreover, for this acquisition geometry, a new neural-network-based reconstruction algorithm is introduced: the neural network Hilbert transform based filtered backprojection. The proposed algorithm is evaluated both on simulated and real inline x-ray data and has shown to generate high quality reconstructions of 400  ×  400 reconstruction pixels within 200 ms, thereby meeting the high throughput criteria.

Synthetic Biomaterials to Rival Nature's Complexity-a Path Forward with Combinatorics, High-Throughput Discovery, and High-Content Analysis.

PubMed

Zhang, Douglas; Lee, Junmin; Kilian, Kristopher A

2017-10-01

Cells in tissue receive a host of soluble and insoluble signals in a context-dependent fashion, where integration of these cues through a complex network of signal transduction cascades will define a particular outcome. Biomaterials scientists and engineers are tasked with designing materials that can at least partially recreate this complex signaling milieu towards new materials for biomedical applications. In this progress report, recent advances in high throughput techniques and high content imaging approaches that are facilitating the discovery of efficacious biomaterials are described. From microarrays of synthetic polymers, peptides and full-length proteins, to designer cell culture systems that present multiple biophysical and biochemical cues in tandem, it is discussed how the integration of combinatorics with high content imaging and analysis is essential to extracting biologically meaningful information from large scale cellular screens to inform the design of next generation biomaterials. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

High Content Imaging (HCI) on Miniaturized Three-Dimensional (3D) Cell Cultures

PubMed Central

Joshi, Pranav; Lee, Moo-Yeal

2015-01-01

High content imaging (HCI) is a multiplexed cell staining assay developed for better understanding of complex biological functions and mechanisms of drug action, and it has become an important tool for toxicity and efficacy screening of drug candidates. Conventional HCI assays have been carried out on two-dimensional (2D) cell monolayer cultures, which in turn limit predictability of drug toxicity/efficacy in vivo; thus, there has been an urgent need to perform HCI assays on three-dimensional (3D) cell cultures. Although 3D cell cultures better mimic in vivo microenvironments of human tissues and provide an in-depth understanding of the morphological and functional features of tissues, they are also limited by having relatively low throughput and thus are not amenable to high-throughput screening (HTS). One attempt of making 3D cell culture amenable for HTS is to utilize miniaturized cell culture platforms. This review aims to highlight miniaturized 3D cell culture platforms compatible with current HCI technology. PMID:26694477

Urological diagnosis using clinical PACS

NASA Astrophysics Data System (ADS)

Mills, Stephen F.; Spetz, Kevin S.; Dwyer, Samuel J., III

1995-05-01

Urological diagnosis using fluoroscopy images has traditionally been performed using radiographic films. Images are generally acquired in conjunction with the application of a contrast agent, processed to create analog films, and inspected to ensure satisfactory image quality prior to being provided to a radiologist for reading. In the case of errors the entire process must be repeated. In addition, the radiologist must then often go to a particular reading room, possibly in a remote part of the healthcare facility, to read the images. The integration of digital fluoroscopy modalities with clinical PACS has the potential to significantly improve the urological diagnosis process by providing high-speed access to images at a variety of locations within a healthcare facility without costly film processing. The PACS additionally provides a cost-effective and reliable means of long-term storage and allows several medical users to simultaneously view the same images at different locations. The installation of a digital data interface between the existing clinically operational PACS at the University of Virginia Health Sciences Center and a digital urology fluoroscope is described. Preliminary user interviews that have been conducted to determine the clinical effectiveness of PACS workstations for urological diagnosis are discussed. The specific suitability of the workstation medium is discussed, as are overall advantages and disadvantages of the hardcopy and softcopy media in terms of efficiency, timeliness and cost. Throughput metrics and some specific parameters of gray-scale viewing stations and the expected system impacts resulting from the integration of a urology fluoroscope with PACS are also discussed.

OPTICAL PROCESSING OF INFORMATION: Multistage optoelectronic two-dimensional image switches

NASA Astrophysics Data System (ADS)

Fedorov, V. B.

1994-06-01

The implementation principles and the feasibility of construction of high-throughput multistage optoelectronic switches, capable of transmitting data in the form of two-dimensional images along interconnected pairs of optical channels, are considered. Different ways of realising compact switches are proposed. They are based on the use of polarisation-sensitive elements, arrays of modulators of the plane of polarisation of light, arrays of objectives, and free-space optics. Optical systems of such switches can theoretically ensure that the resolution and optical losses in two-dimensional image transmission are limited only by diffraction. Estimates are obtained of the main maximum-performance parameters of the proposed optoelectronic image switches.

Hyperspectral imaging using the single-pixel Fourier transform technique

NASA Astrophysics Data System (ADS)

Jin, Senlin; Hui, Wangwei; Wang, Yunlong; Huang, Kaicheng; Shi, Qiushuai; Ying, Cuifeng; Liu, Dongqi; Ye, Qing; Zhou, Wenyuan; Tian, Jianguo

2017-03-01

Hyperspectral imaging technology is playing an increasingly important role in the fields of food analysis, medicine and biotechnology. To improve the speed of operation and increase the light throughput in a compact equipment structure, a Fourier transform hyperspectral imaging system based on a single-pixel technique is proposed in this study. Compared with current imaging spectrometry approaches, the proposed system has a wider spectral range (400-1100 nm), a better spectral resolution (1 nm) and requires fewer measurement data (a sample rate of 6.25%). The performance of this system was verified by its application to the non-destructive testing of potatoes.

Novel microscale approaches for easy, rapid determination of protein stability in academic and commercial settings

PubMed Central

Alexander, Crispin G.; Wanner, Randy; Johnson, Christopher M.; Breitsprecher, Dennis; Winter, Gerhard; Duhr, Stefan; Baaske, Philipp; Ferguson, Neil

2014-01-01

Chemical denaturant titrations can be used to accurately determine protein stability. However, data acquisition is typically labour intensive, has low throughput and is difficult to automate. These factors, combined with high protein consumption, have limited the adoption of chemical denaturant titrations in commercial settings. Thermal denaturation assays can be automated, sometimes with very high throughput. However, thermal denaturation assays are incompatible with proteins that aggregate at high temperatures and large extrapolation of stability parameters to physiological temperatures can introduce significant uncertainties. We used capillary-based instruments to measure chemical denaturant titrations by intrinsic fluorescence and microscale thermophoresis. This allowed higher throughput, consumed several hundred-fold less protein than conventional, cuvette-based methods yet maintained the high quality of the conventional approaches. We also established efficient strategies for automated, direct determination of protein stability at a range of temperatures via chemical denaturation, which has utility for characterising stability for proteins that are difficult to purify in high yield. This approach may also have merit for proteins that irreversibly denature or aggregate in classical thermal denaturation assays. We also developed procedures for affinity ranking of protein–ligand interactions from ligand-induced changes in chemical denaturation data, and proved the principle for this by correctly ranking the affinity of previously unreported peptide–PDZ domain interactions. The increased throughput, automation and low protein consumption of protein stability determinations afforded by using capillary-based methods to measure denaturant titrations, can help to revolutionise protein research. We believe that the strategies reported are likely to find wide applications in academia, biotherapeutic formulation and drug discovery programmes. PMID:25262836

1-Million droplet array with wide-field fluorescence imaging for digital PCR.

PubMed

Hatch, Andrew C; Fisher, Jeffrey S; Tovar, Armando R; Hsieh, Albert T; Lin, Robert; Pentoney, Stephen L; Yang, David L; Lee, Abraham P

2011-11-21

Digital droplet reactors are useful as chemical and biological containers to discretize reagents into picolitre or nanolitre volumes for analysis of single cells, organisms, or molecules. However, most DNA based assays require processing of samples on the order of tens of microlitres and contain as few as one to as many as millions of fragments to be detected. Presented in this work is a droplet microfluidic platform and fluorescence imaging setup designed to better meet the needs of the high-throughput and high-dynamic-range by integrating multiple high-throughput droplet processing schemes on the chip. The design is capable of generating over 1-million, monodisperse, 50 picolitre droplets in 2-7 minutes that then self-assemble into high density 3-dimensional sphere packing configurations in a large viewing chamber for visualization and analysis. This device then undergoes on-chip polymerase chain reaction (PCR) amplification and fluorescence detection to digitally quantify the sample's nucleic acid contents. Wide-field fluorescence images are captured using a low cost 21-megapixel digital camera and macro-lens with an 8-12 cm(2) field-of-view at 1× to 0.85× magnification, respectively. We demonstrate both end-point and real-time imaging ability to perform on-chip quantitative digital PCR analysis of the entire droplet array. Compared to previous work, this highly integrated design yields a 100-fold increase in the number of on-chip digitized reactors with simultaneous fluorescence imaging for digital PCR based assays.

Technology insertion of a COTS RAID server as an image buffer in the image chain of the Defense Mapping Agency's Digital Production System

NASA Astrophysics Data System (ADS)

Mehring, James W.; Thomas, Scott D.

1995-11-01

The Data Services Segment of the Defense Mapping Agency's Digital Production System provides a digital archive of imagery source data for use by DMA's cartographic user's. This system was developed in the mid-1980's and is currently undergoing modernization. This paper addresses the modernization of the imagery buffer function that was performed by custom hardware in the baseline system and is being replaced by a RAID Server based on commercial off the shelf (COTS) hardware. The paper briefly describes the baseline DMA image system and the modernization program, that is currently under way. Throughput benchmark measurements were made to make design configuration decisions for a commercial off the shelf (COTS) RAID Server to perform as system image buffer. The test program began with performance measurements of the RAID read and write operations between the RAID arrays and the server CPU for RAID levels 0, 5 and 0+1. Interface throughput measurements were made for the HiPPI interface between the RAID Server and the image archive and processing system as well as the client side interface between a custom interface board that provides the interface between the internal bus of the RAID Server and the Input- Output Processor (IOP) external wideband network currently in place in the DMA system to service client workstations. End to end measurements were taken from the HiPPI interface through the RAID write and read operations to the IOP output interface.

GiA Roots: software for the high throughput analysis of plant root system architecture.

PubMed

Galkovskyi, Taras; Mileyko, Yuriy; Bucksch, Alexander; Moore, Brad; Symonova, Olga; Price, Charles A; Topp, Christopher N; Iyer-Pascuzzi, Anjali S; Zurek, Paul R; Fang, Suqin; Harer, John; Benfey, Philip N; Weitz, Joshua S

2012-07-26

Characterizing root system architecture (RSA) is essential to understanding the development and function of vascular plants. Identifying RSA-associated genes also represents an underexplored opportunity for crop improvement. Software tools are needed to accelerate the pace at which quantitative traits of RSA are estimated from images of root networks. We have developed GiA Roots (General Image Analysis of Roots), a semi-automated software tool designed specifically for the high-throughput analysis of root system images. GiA Roots includes user-assisted algorithms to distinguish root from background and a fully automated pipeline that extracts dozens of root system phenotypes. Quantitative information on each phenotype, along with intermediate steps for full reproducibility, is returned to the end-user for downstream analysis. GiA Roots has a GUI front end and a command-line interface for interweaving the software into large-scale workflows. GiA Roots can also be extended to estimate novel phenotypes specified by the end-user. We demonstrate the use of GiA Roots on a set of 2393 images of rice roots representing 12 genotypes from the species Oryza sativa. We validate trait measurements against prior analyses of this image set that demonstrated that RSA traits are likely heritable and associated with genotypic differences. Moreover, we demonstrate that GiA Roots is extensible and an end-user can add functionality so that GiA Roots can estimate novel RSA traits. In summary, we show that the software can function as an efficient tool as part of a workflow to move from large numbers of root images to downstream analysis.

Quantification of diffusion tensor imaging in normal white matter maturation of early childhood using an automated processing pipeline.

PubMed

Loh, K B; Ramli, N; Tan, L K; Roziah, M; Rahmat, K; Ariffin, H

2012-07-01

The degree and status of white matter myelination can be sensitively monitored using diffusion tensor imaging (DTI). This study looks at the measurement of fractional anistropy (FA) and mean diffusivity (MD) using an automated ROI with an existing DTI atlas. Anatomical MRI and structural DTI were performed cross-sectionally on 26 normal children (newborn to 48 months old), using 1.5-T MRI. The automated processing pipeline was implemented to convert diffusion-weighted images into the NIfTI format. DTI-TK software was used to register the processed images to the ICBM DTI-81 atlas, while AFNI software was used for automated atlas-based volumes of interest (VOIs) and statistical value extraction. DTI exhibited consistent grey-white matter contrast. Triphasic temporal variation of the FA and MD values was noted, with FA increasing and MD decreasing rapidly early in the first 12 months. The second phase lasted 12-24 months during which the rate of FA and MD changes was reduced. After 24 months, the FA and MD values plateaued. DTI is a superior technique to conventional MR imaging in depicting WM maturation. The use of the automated processing pipeline provides a reliable environment for quantitative analysis of high-throughput DTI data. Diffusion tensor imaging outperforms conventional MRI in depicting white matter maturation. • DTI will become an important clinical tool for diagnosing paediatric neurological diseases. • DTI appears especially helpful for developmental abnormalities, tumours and white matter disease. • An automated processing pipeline assists quantitative analysis of high throughput DTI data.

Beamforming transmission in IEEE 802.11ac under time-varying channels.

PubMed

Yu, Heejung; Kim, Taejoon

2014-01-01

The IEEE 802.11ac wireless local area network (WLAN) standard has adopted beamforming (BF) schemes to improve spectral efficiency and throughput with multiple antennas. To design the transmit beam, a channel sounding process to feedback channel state information (CSI) is required. Due to sounding overhead, throughput increases with the amount of transmit data under static channels. Under practical channel conditions with mobility, however, the mismatch between the transmit beam and the channel at transmission time causes performance loss when transmission duration after channel sounding is too long. When the fading rate, payload size, and operating signal-to-noise ratio are given, the optimal transmission duration (i.e., packet length) can be determined to maximize throughput. The relationship between packet length and throughput is also investigated for single-user and multiuser BF modes.

Beamforming Transmission in IEEE 802.11ac under Time-Varying Channels

PubMed Central

2014-01-01

The IEEE 802.11ac wireless local area network (WLAN) standard has adopted beamforming (BF) schemes to improve spectral efficiency and throughput with multiple antennas. To design the transmit beam, a channel sounding process to feedback channel state information (CSI) is required. Due to sounding overhead, throughput increases with the amount of transmit data under static channels. Under practical channel conditions with mobility, however, the mismatch between the transmit beam and the channel at transmission time causes performance loss when transmission duration after channel sounding is too long. When the fading rate, payload size, and operating signal-to-noise ratio are given, the optimal transmission duration (i.e., packet length) can be determined to maximize throughput. The relationship between packet length and throughput is also investigated for single-user and multiuser BF modes. PMID:25152927

UAV-Based Thermal Imaging for High-Throughput Field Phenotyping of Black Poplar Response to Drought

PubMed Central

Ludovisi, Riccardo; Tauro, Flavia; Salvati, Riccardo; Khoury, Sacha; Mugnozza Scarascia, Giuseppe; Harfouche, Antoine

2017-01-01

Poplars are fast-growing, high-yielding forest tree species, whose cultivation as second-generation biofuel crops is of increasing interest and can efficiently meet emission reduction goals. Yet, breeding elite poplar trees for drought resistance remains a major challenge. Worldwide breeding programs are largely focused on intra/interspecific hybridization, whereby Populus nigra L. is a fundamental parental pool. While high-throughput genotyping has resulted in unprecedented capabilities to rapidly decode complex genetic architecture of plant stress resistance, linking genomics to phenomics is hindered by technically challenging phenotyping. Relying on unmanned aerial vehicle (UAV)-based remote sensing and imaging techniques, high-throughput field phenotyping (HTFP) aims at enabling highly precise and efficient, non-destructive screening of genotype performance in large populations. To efficiently support forest-tree breeding programs, ground-truthing observations should be complemented with standardized HTFP. In this study, we develop a high-resolution (leaf level) HTFP approach to investigate the response to drought of a full-sib F2 partially inbred population (termed here ‘POP6’), whose F1 was obtained from an intraspecific P. nigra controlled cross between genotypes with highly divergent phenotypes. We assessed the effects of two water treatments (well-watered and moderate drought) on a population of 4603 trees (503 genotypes) hosted in two adjacent experimental plots (1.67 ha) by conducting low-elevation (25 m) flights with an aerial drone and capturing 7836 thermal infrared (TIR) images. TIR images were undistorted, georeferenced, and orthorectified to obtain radiometric mosaics. Canopy temperature (Tc) was extracted using two independent semi-automated segmentation techniques, eCognition- and Matlab-based, to avoid the mixed-pixel problem. Overall, results showed that the UAV platform-based thermal imaging enables to effectively assess genotype variability under drought stress conditions. Tc derived from aerial thermal imagery presented a good correlation with ground-truth stomatal conductance (gs) in both segmentation techniques. Interestingly, the HTFP approach was instrumental to detect drought-tolerant response in 25% of the population. This study shows the potential of UAV-based thermal imaging for field phenomics of poplar and other tree species. This is anticipated to have tremendous implications for accelerating forest tree genetic improvement against abiotic stress. PMID:29021803

UAV-Based Thermal Imaging for High-Throughput Field Phenotyping of Black Poplar Response to Drought.

PubMed

Ludovisi, Riccardo; Tauro, Flavia; Salvati, Riccardo; Khoury, Sacha; Mugnozza Scarascia, Giuseppe; Harfouche, Antoine

2017-01-01

Poplars are fast-growing, high-yielding forest tree species, whose cultivation as second-generation biofuel crops is of increasing interest and can efficiently meet emission reduction goals. Yet, breeding elite poplar trees for drought resistance remains a major challenge. Worldwide breeding programs are largely focused on intra/interspecific hybridization, whereby Populus nigra L. is a fundamental parental pool. While high-throughput genotyping has resulted in unprecedented capabilities to rapidly decode complex genetic architecture of plant stress resistance, linking genomics to phenomics is hindered by technically challenging phenotyping. Relying on unmanned aerial vehicle (UAV)-based remote sensing and imaging techniques, high-throughput field phenotyping (HTFP) aims at enabling highly precise and efficient, non-destructive screening of genotype performance in large populations. To efficiently support forest-tree breeding programs, ground-truthing observations should be complemented with standardized HTFP. In this study, we develop a high-resolution (leaf level) HTFP approach to investigate the response to drought of a full-sib F 2 partially inbred population (termed here 'POP6'), whose F 1 was obtained from an intraspecific P. nigra controlled cross between genotypes with highly divergent phenotypes. We assessed the effects of two water treatments (well-watered and moderate drought) on a population of 4603 trees (503 genotypes) hosted in two adjacent experimental plots (1.67 ha) by conducting low-elevation (25 m) flights with an aerial drone and capturing 7836 thermal infrared (TIR) images. TIR images were undistorted, georeferenced, and orthorectified to obtain radiometric mosaics. Canopy temperature ( T c ) was extracted using two independent semi-automated segmentation techniques, eCognition- and Matlab-based, to avoid the mixed-pixel problem. Overall, results showed that the UAV platform-based thermal imaging enables to effectively assess genotype variability under drought stress conditions. T c derived from aerial thermal imagery presented a good correlation with ground-truth stomatal conductance ( g s ) in both segmentation techniques. Interestingly, the HTFP approach was instrumental to detect drought-tolerant response in 25% of the population. This study shows the potential of UAV-based thermal imaging for field phenomics of poplar and other tree species. This is anticipated to have tremendous implications for accelerating forest tree genetic improvement against abiotic stress.

A novel quantitative assay of mitophagy: Combining high content fluorescence microscopy and mitochondrial DNA load to quantify mitophagy and identify novel pharmacological tools against pathogenic heteroplasmic mtDNA.

PubMed

Diot, Alan; Hinks-Roberts, Alex; Lodge, Tiffany; Liao, Chunyan; Dombi, Eszter; Morten, Karl; Brady, Stefen; Fratter, Carl; Carver, Janet; Muir, Rebecca; Davis, Ryan; Green, Charlotte J; Johnston, Iain; Hilton-Jones, David; Sue, Carolyn; Mortiboys, Heather; Poulton, Joanna

2015-10-01

Mitophagy is a cellular mechanism for the recycling of mitochondrial fragments. This process is able to improve mitochondrial DNA (mtDNA) quality in heteroplasmic mtDNA disease, in which mutant mtDNA co-exists with normal mtDNA. In disorders where the load of mutant mtDNA determines disease severity it is likely to be an important determinant of disease progression. Measuring mitophagy is technically demanding. We used pharmacological modulators of autophagy to validate two techniques for quantifying mitophagy. First we used the IN Cell 1000 analyzer to quantify mitochondrial co-localisation with LC3-II positive autophagosomes. Unlike conventional fluorescence and electron microscopy, this high-throughput system is sufficiently sensitive to detect transient low frequency autophagosomes. Secondly, because mitophagy preferentially removes pathogenic heteroplasmic mtDNA mutants, we developed a heteroplasmy assay based on loss of m.3243A>G mtDNA, during culture conditions requiring oxidative metabolism ("energetic stress"). The effects of the pharmacological modulators on these two measures were consistent, confirming that the high throughput imaging output (autophagosomes co-localising with mitochondria) reflects mitochondrial quality control. To further validate these methods, we performed a more detailed study using metformin, the most commonly prescribed antidiabetic drug that is still sometimes used in Maternally Inherited Diabetes and Deafness (MIDD). This confirmed our initial findings and revealed that metformin inhibits mitophagy at clinically relevant concentrations, suggesting that it may have novel therapeutic uses. Copyright © 2015. Published by Elsevier Ltd.

Efficient Sample Delay Calculation for 2-D and 3-D Ultrasound Imaging.

PubMed

Ibrahim, Aya; Hager, Pascal A; Bartolini, Andrea; Angiolini, Federico; Arditi, Marcel; Thiran, Jean-Philippe; Benini, Luca; De Micheli, Giovanni

2017-08-01

Ultrasound imaging is a reference medical diagnostic technique, thanks to its blend of versatility, effectiveness, and moderate cost. The core computation of all ultrasound imaging methods is based on simple formulae, except for those required to calculate acoustic propagation delays with high precision and throughput. Unfortunately, advanced three-dimensional (3-D) systems require the calculation or storage of billions of such delay values per frame, which is a challenge. In 2-D systems, this requirement can be four orders of magnitude lower, but efficient computation is still crucial in view of low-power implementations that can be battery-operated, enabling usage in numerous additional scenarios. In this paper, we explore two smart designs of the delay generation function. To quantify their hardware cost, we implement them on FPGA and study their footprint and performance. We evaluate how these architectures scale to different ultrasound applications, from a low-power 2-D system to a next-generation 3-D machine. When using numerical approximations, we demonstrate the ability to generate delay values with sufficient throughput to support 10 000-channel 3-D imaging at up to 30 fps while using 63% of a Virtex 7 FPGA, requiring 24 MB of external memory accessed at about 32 GB/s bandwidth. Alternatively, with similar FPGA occupation, we show an exact calculation method that reaches 24 fps on 1225-channel 3-D imaging and does not require external memory at all. Both designs can be scaled to use a negligible amount of resources for 2-D imaging in low-power applications and for ultrafast 2-D imaging at hundreds of frames per second.

Multimodal ophthalmic imaging using spectrally encoded scanning laser ophthalmoscopy and optical coherence tomography

NASA Astrophysics Data System (ADS)

El-Haddad, Mohamed T.; Malone, Joseph D.; Li, Jianwei D.; Bozic, Ivan; Arquitola, Amber M.; Joos, Karen M.; Patel, Shriji N.; Tao, Yuankai K.

2017-08-01

Ophthalmic surgery involves manipulation of delicate, layered tissue structures on milli- to micrometer scales. Traditional surgical microscopes provide an inherently two-dimensional view of the surgical field with limited depth perception which precludes accurate depth-resolved visualization of these tissue layers, and limits the development of novel surgical techniques. We demonstrate multimodal swept-source spectrally encoded scanning laser ophthalmoscopy and optical coherence tomography (SS-SESLO-OCT) to address current limitations of image-guided ophthalmic microsurgery. SS-SESLO-OCT provides inherently co-registered en face and cross-sectional field-of-views (FOVs) at a line rate of 400 kHz and >2 GPix/s throughput. We show in vivo imaging of the anterior segment and retinal fundus of a healthy volunteer, and preliminary results of multi-volumetric mosaicking for ultrawide-field retinal imaging with 90° FOV. Additionally, a scan-head was rapid-prototyped with a modular architecture which enabled integration of SS-SESLO-OCT with traditional surgical microscope and slit-lamp imaging optics. Ex vivo surgical maneuvers were simulated in cadaveric porcine eyes. The system throughput enabled volumetric acquisition at 10 volumes-per-second (vps) and allowed visualization of surgical dynamics in corneal sweeps, compressions, and dissections, and retinal sweeps, compressions, and elevations. SESLO en face images enabled simple real-time co-registration with the surgical microscope FOV, and OCT cross-sections provided depth-resolved visualization of instrument-tissue interactions. Finally, we demonstrate novel augmented-reality integration with the surgical view using segmentation overlays to aid surgical guidance. SS-SESLO-OCT may benefit clinical diagnostics by enabling aiming, registration, and mosaicking; and intraoperative imaging by allowing for real-time surgical feedback, instrument tracking, and overlays of computationally extracted biomarkers of disease.

High-Throughput Epitope Binning Assays on Label-Free Array-Based Biosensors Can Yield Exquisite Epitope Discrimination That Facilitates the Selection of Monoclonal Antibodies with Functional Activity

PubMed Central

Abdiche, Yasmina Noubia; Miles, Adam; Eckman, Josh; Foletti, Davide; Van Blarcom, Thomas J.; Yeung, Yik Andy; Pons, Jaume; Rajpal, Arvind

2014-01-01

Here, we demonstrate how array-based label-free biosensors can be applied to the multiplexed interaction analysis of large panels of analyte/ligand pairs, such as the epitope binning of monoclonal antibodies (mAbs). In this application, the larger the number of mAbs that are analyzed for cross-blocking in a pairwise and combinatorial manner against their specific antigen, the higher the probability of discriminating their epitopes. Since cross-blocking of two mAbs is necessary but not sufficient for them to bind an identical epitope, high-resolution epitope binning analysis determined by high-throughput experiments can enable the identification of mAbs with similar but unique epitopes. We demonstrate that a mAb's epitope and functional activity are correlated, thereby strengthening the relevance of epitope binning data to the discovery of therapeutic mAbs. We evaluated two state-of-the-art label-free biosensors that enable the parallel analysis of 96 unique analyte/ligand interactions and nearly ten thousand total interactions per unattended run. The IBIS-MX96 is a microarray-based surface plasmon resonance imager (SPRi) integrated with continuous flow microspotting technology whereas the Octet-HTX is equipped with disposable fiber optic sensors that use biolayer interferometry (BLI) detection. We compared their throughput, versatility, ease of sample preparation, and sample consumption in the context of epitope binning assays. We conclude that the main advantages of the SPRi technology are its exceptionally low sample consumption, facile sample preparation, and unparalleled unattended throughput. In contrast, the BLI technology is highly flexible because it allows for the simultaneous interaction analysis of 96 independent analyte/ligand pairs, ad hoc sensor replacement and on-line reloading of an analyte- or ligand-array. Thus, the complementary use of these two platforms can expedite applications that are relevant to the discovery of therapeutic mAbs, depending upon the sample availability, and the number and diversity of the interactions being studied. PMID:24651868

Dual Telecentric Lens System For Projection Onto Tilted Toroidal Screen

NASA Technical Reports Server (NTRS)

Gold, Ronald S.; Hudyma, Russell M.

1995-01-01

System of two optical assemblies for projecting image onto tilted toroidal screen. One projection lens optimized for red and green spectral region; other for blue. Dual-channel approach offers several advantages which include: simplified color filtering, simplified chromatic aberration corrections, less complex polarizing prism arrangement, and increased throughput of blue light energy. Used in conjunction with any source of imagery, designed especially to project images formed by reflection of light from liquid-crystal light valve (LCLV).

Coded aperture imaging with self-supporting uniformly redundant arrays. [Patent application

DOEpatents

Fenimore, E.E.

1980-09-26

A self-supporting uniformly redundant array pattern for coded aperture imaging. The invention utilizes holes which are an integer times smaller in each direction than holes in conventional URA patterns. A balance correlation function is generated where holes are represented by 1's, nonholes are represented by -1's, and supporting area is represented by 0's. The self-supporting array can be used for low energy applications where substrates would greatly reduce throughput.

Applying Zeeman Doppler imaging to solar spectra

NASA Astrophysics Data System (ADS)

Hussain, G. A. J.; Saar, S. H.; Collier Cameron, A.

2004-03-01

A new generation of spectro-polarimeters with high throughput (e.g. CFHT/ESPADONS and LBT/PEPSI) is becoming available. This opportunity can be exploited using Zeeman Doppler imaging (ZDI), a technique that inverts time-series of Stokes V spectra to map stellar surface magnetic fields (Semel 1989). ZDI is assisted by ``Least squares deconvolution'' (LSD), which sums up the signal from 1000's of photospheric lines to produce a mean deconvolved profile with higher S:N (Donati & Collier Cameron 1997).

Wavelength-Scanning SPR Imaging Sensors Based on an Acousto-Optic Tunable Filter and a White Light Laser

PubMed Central

Zeng, Youjun; Wang, Lei; Wu, Shu-Yuen; He, Jianan; Qu, Junle; Li, Xuejin; Ho, Ho-Pui; Gu, Dayong; Gao, Bruce Zhi; Shao, Yonghong

2017-01-01

A fast surface plasmon resonance (SPR) imaging biosensor system based on wavelength interrogation using an acousto-optic tunable filter (AOTF) and a white light laser is presented. The system combines the merits of a wide-dynamic detection range and high sensitivity offered by the spectral approach with multiplexed high-throughput data collection and a two-dimensional (2D) biosensor array. The key feature is the use of AOTF to realize wavelength scan from a white laser source and thus to achieve fast tracking of the SPR dip movement caused by target molecules binding to the sensor surface. Experimental results show that the system is capable of completing a SPR dip measurement within 0.35 s. To the best of our knowledge, this is the fastest time ever reported in the literature for imaging spectral interrogation. Based on a spectral window with a width of approximately 100 nm, a dynamic detection range and resolution of 4.63 × 10−2 refractive index unit (RIU) and 1.27 × 10−6 RIU achieved in a 2D-array sensor is reported here. The spectral SPR imaging sensor scheme has the capability of performing fast high-throughput detection of biomolecular interactions from 2D sensor arrays. The design has no mechanical moving parts, thus making the scheme completely solid-state. PMID:28067766

Imaging of Cell-Cell Communication in a Vertical Orientation Reveals High-Resolution Structure of Immunological Synapse and Novel PD-1 Dynamics

PubMed Central

Jang, Joon Hee; Huang, Yu; Zheng, Peilin; Jo, Myeong Chan; Bertolet, Grant; Qin, Lidong; Liu, Dongfang

2015-01-01

The immunological synapse (IS) is one of the most pivotal communication strategies in immune cells. Understanding the molecular basis of the IS provides critical information regarding how immune cells mount an effective immune response. Fluorescence microscopy provides a fundamental tool to study the IS. However, current imaging techniques for studying the IS cannot sufficiently achieve high resolution in real cell-cell conjugates. Here we present a new device that allows for high-resolution imaging of the IS with conventional confocal microscopy in a high-throughput manner. Combining micropits and single cell trap arrays, we have developed a new microfluidic platform that allows visualization of the IS in vertically “stacked” cells. Using this vertical cell pairing (VCP) system, we investigated the dynamics of the inhibitory synapse mediated by an inhibitory receptor, programed death protein-1 (PD-1) and the cytotoxic synapse at the single cell level. In addition to the technique innovation, we demonstrated novel biological findings by this VCP device, including novel distribution of F-actin and cytolytic granules at the IS, PD-1 microclusters in the NK IS, and kinetics of cytotoxicity. We propose that this high-throughput, cost-effective, easy-to-use VCP system, along with conventional imaging techniques, can be used to address a number of significant biological questions in a variety of disciplines. PMID:26123352

Neural network control of focal position during time-lapse microscopy of cells.

PubMed

Wei, Ling; Roberts, Elijah

2018-05-09

Live-cell microscopy is quickly becoming an indispensable technique for studying the dynamics of cellular processes. Maintaining the specimen in focus during image acquisition is crucial for high-throughput applications, especially for long experiments or when a large sample is being continuously scanned. Automated focus control methods are often expensive, imperfect, or ill-adapted to a specific application and are a bottleneck for widespread adoption of high-throughput, live-cell imaging. Here, we demonstrate a neural network approach for automatically maintaining focus during bright-field microscopy. Z-stacks of yeast cells growing in a microfluidic device were collected and used to train a convolutional neural network to classify images according to their z-position. We studied the effect on prediction accuracy of the various hyperparameters of the neural network, including downsampling, batch size, and z-bin resolution. The network was able to predict the z-position of an image with ±1 μm accuracy, outperforming human annotators. Finally, we used our neural network to control microscope focus in real-time during a 24 hour growth experiment. The method robustly maintained the correct focal position compensating for 40 μm of focal drift and was insensitive to changes in the field of view. About ~100 annotated z-stacks were required to train the network making our method quite practical for custom autofocus applications.

Hierarchical classification strategy for Phenotype extraction from epidermal growth factor receptor endocytosis screening.

PubMed

Cao, Lu; Graauw, Marjo de; Yan, Kuan; Winkel, Leah; Verbeek, Fons J

2016-05-03

Endocytosis is regarded as a mechanism of attenuating the epidermal growth factor receptor (EGFR) signaling and of receptor degradation. There is increasing evidence becoming available showing that breast cancer progression is associated with a defect in EGFR endocytosis. In order to find related Ribonucleic acid (RNA) regulators in this process, high-throughput imaging with fluorescent markers is used to visualize the complex EGFR endocytosis process. Subsequently a dedicated automatic image and data analysis system is developed and applied to extract the phenotype measurement and distinguish different developmental episodes from a huge amount of images acquired through high-throughput imaging. For the image analysis, a phenotype measurement quantifies the important image information into distinct features or measurements. Therefore, the manner in which prominent measurements are chosen to represent the dynamics of the EGFR process becomes a crucial step for the identification of the phenotype. In the subsequent data analysis, classification is used to categorize each observation by making use of all prominent measurements obtained from image analysis. Therefore, a better construction for a classification strategy will support to raise the performance level in our image and data analysis system. In this paper, we illustrate an integrated analysis method for EGFR signalling through image analysis of microscopy images. Sophisticated wavelet-based texture measurements are used to obtain a good description of the characteristic stages in the EGFR signalling. A hierarchical classification strategy is designed to improve the recognition of phenotypic episodes of EGFR during endocytosis. Different strategies for normalization, feature selection and classification are evaluated. The results of performance assessment clearly demonstrate that our hierarchical classification scheme combined with a selected set of features provides a notable improvement in the temporal analysis of EGFR endocytosis. Moreover, it is shown that the addition of the wavelet-based texture features contributes to this improvement. Our workflow can be applied to drug discovery to analyze defected EGFR endocytosis processes.

Large-scale Topographical Screen for Investigation of Physical Neural-Guidance Cues

NASA Astrophysics Data System (ADS)

Li, Wei; Tang, Qing Yuan; Jadhav, Amol D.; Narang, Ankit; Qian, Wei Xian; Shi, Peng; Pang, Stella W.

2015-03-01

A combinatorial approach was used to present primary neurons with a large library of topographical features in the form of micropatterned substrate for high-throughput screening of physical neural-guidance cues that can effectively promote different aspects of neuronal development, including axon and dendritic outgrowth. Notably, the neuronal-guidance capability of specific features was automatically identified using a customized image processing software, thus significantly increasing the screening throughput with minimal subjective bias. Our results indicate that the anisotropic topographies promote axonal and in some cases dendritic extension relative to the isotropic topographies, while dendritic branching showed preference to plain substrates over the microscale features. The results from this work can be readily applied towards engineering novel biomaterials with precise surface topography that can serve as guidance conduits for neuro-regenerative applications. This novel topographical screening strategy combined with the automated processing capability can also be used for high-throughput screening of chemical or genetic regulatory factors in primary neurons.

Efficient Strategies for Estimating the Spatial Coherence of Backscatter

PubMed Central

Hyun, Dongwoon; Crowley, Anna Lisa C.; Dahl, Jeremy J.

2017-01-01

The spatial coherence of ultrasound backscatter has been proposed to reduce clutter in medical imaging, to measure the anisotropy of the scattering source, and to improve the detection of blood flow. These techniques rely on correlation estimates that are obtained using computationally expensive strategies. In this study, we assess existing spatial coherence estimation methods and propose three computationally efficient modifications: a reduced kernel, a downsampled receive aperture, and the use of an ensemble correlation coefficient. The proposed methods are implemented in simulation and in vivo studies. Reducing the kernel to a single sample improved computational throughput and improved axial resolution. Downsampling the receive aperture was found to have negligible effect on estimator variance, and improved computational throughput by an order of magnitude for a downsample factor of 4. The ensemble correlation estimator demonstrated lower variance than the currently used average correlation. Combining the three methods, the throughput was improved 105-fold in simulation with a downsample factor of 4 and 20-fold in vivo with a downsample factor of 2. PMID:27913342

Cox-nnet: An artificial neural network method for prognosis prediction of high-throughput omics data

PubMed Central

Ching, Travers; Zhu, Xun

2018-01-01

Artificial neural networks (ANN) are computing architectures with many interconnections of simple neural-inspired computing elements, and have been applied to biomedical fields such as imaging analysis and diagnosis. We have developed a new ANN framework called Cox-nnet to predict patient prognosis from high throughput transcriptomics data. In 10 TCGA RNA-Seq data sets, Cox-nnet achieves the same or better predictive accuracy compared to other methods, including Cox-proportional hazards regression (with LASSO, ridge, and mimimax concave penalty), Random Forests Survival and CoxBoost. Cox-nnet also reveals richer biological information, at both the pathway and gene levels. The outputs from the hidden layer node provide an alternative approach for survival-sensitive dimension reduction. In summary, we have developed a new method for accurate and efficient prognosis prediction on high throughput data, with functional biological insights. The source code is freely available at https://github.com/lanagarmire/cox-nnet. PMID:29634719

LOCATE: a mouse protein subcellular localization database

PubMed Central

Fink, J. Lynn; Aturaliya, Rajith N.; Davis, Melissa J.; Zhang, Fasheng; Hanson, Kelly; Teasdale, Melvena S.; Kai, Chikatoshi; Kawai, Jun; Carninci, Piero; Hayashizaki, Yoshihide; Teasdale, Rohan D.

2006-01-01

We present here LOCATE, a curated, web-accessible database that houses data describing the membrane organization and subcellular localization of proteins from the FANTOM3 Isoform Protein Sequence set. Membrane organization is predicted by the high-throughput, computational pipeline MemO. The subcellular locations of selected proteins from this set were determined by a high-throughput, immunofluorescence-based assay and by manually reviewing >1700 peer-reviewed publications. LOCATE represents the first effort to catalogue the experimentally verified subcellular location and membrane organization of mammalian proteins using a high-throughput approach and provides localization data for ∼40% of the mouse proteome. It is available at . PMID:16381849

Identifying Candidate Chemical-Disease Linkages ...

EPA Pesticide Factsheets

Presentation at meeting on Environmental and Epigenetic Determinants of IBD in New York, NY on identifying candidate chemical-disease linkages by using AOPs to identify molecular initiating events and using relevant high throughput assays to screen for candidate chemicals. This hazard information is combined with exposure models to inform risk assessment. Presentation at meeting on Environmental and Epigenetic Determinants of IBD in New York, NY on identifying candidate chemical-disease linkages by using AOPs to identify molecular initiating events and using relevant high throughput assays to screen for candidate chemicals. This hazard information is combined with exposure models to inform risk assessment.

One Size Fits All: Evaluation of the Transferability of a New "Learning" Histologic Image Analysis Application.

PubMed

Arlt, Janine; Homeyer, André; Sänger, Constanze; Dahmen, Uta; Dirsch, Olaf

2016-01-01

Quantitative analysis of histologic slides is of importance for pathology and also to address surgical questions. Recently, a novel application was developed for the automated quantification of whole-slide images. The aim of this study was to test and validate the underlying image analysis algorithm with respect to user friendliness, accuracy, and transferability to different histologic scenarios. The algorithm splits the images into tiles of a predetermined size and identifies the tissue class of each tile. In the training procedure, the user specifies example tiles of the different tissue classes. In the subsequent analysis procedure, the algorithm classifies each tile into the previously specified classes. User friendliness was evaluated by recording training time and testing reproducibility of the training procedure of users with different background. Accuracy was determined with respect to single and batch analysis. Transferability was demonstrated by analyzing tissue of different organs (rat liver, kidney, small bowel, and spleen) and with different stainings (glutamine synthetase and hematoxylin-eosin). Users of different educational background could apply the program efficiently after a short introduction. When analyzing images with similar properties, accuracy of >90% was reached in single images as well as in batch mode. We demonstrated that the novel application is user friendly and very accurate. With the "training" procedure the application can be adapted to novel image characteristics simply by giving examples of relevant tissue structures. Therefore, it is suitable for the fast and efficient analysis of high numbers of fully digitalized histologic sections, potentially allowing "high-throughput" quantitative "histomic" analysis.

High-speed quantitative phase imaging using time-stretch spectral shearing contrast (Conference Presentation)

NASA Astrophysics Data System (ADS)

Bosworth, Bryan; Foster, Mark A.

2017-02-01

Photonic time-stretch microscopy (TSM) provides an ideal platform for high-throughput imaging flow cytometry, affording extremely high shutter speeds and frame rates with high sensitivity. In order to resolve weakly scattering cells in biofluid and solve the issue of signal-to-noise in cell labeling specificity of biomarkers in imaging flow cytometry, several quantitative phase (QP) techniques have recently been adapted to TSM. However, these techniques have relied primarily on sensitive free-space optical configurations to generate full electric field measurements. The present work draws from the field of ultrashort pulse characterization to leverage the coherence of the ultrashort optical pulses integral to all TSM systems in order to do self-referenced single-shot quantitative phase imaging in a TSM system. Self-referencing is achieved via spectral shearing interferometry in an exceptionally stable and straightforward Sagnac loop incorporating an electro-optic phase modulator and polarization-maintaining fiber that produce sheared and unsheared copies of the pulse train with an inter-pulse delay determined by polarization mode dispersion. The spectral interferogram then yields a squared amplitude and a phase derivative image that can be integrated for conventional phase. We apply this spectral shearing contrast microscope to acquire QP images on a high-speed flow microscope at 90-MHz line rates with <400 pixels per line. We also consider the extension of this technique to compressed sensing (CS) acquisition by intensity modulating the interference spectra with pseudorandom binary waveforms to reconstruct the images from a highly sub-Nyquist number of random inner products, providing a path to even higher operating rates and reduced data storage requirements.

Reflective optical imaging system with balanced distortion

DOEpatents

Chapman, Henry N.; Hudyma, Russell M.; Shafer, David R.; Sweeney, Donald W.

1999-01-01

An optical system compatible with short wavelength (extreme ultraviolet) An optical system compatible with short wavelength (extreme ultraviolet) radiation comprising four reflective elements for projecting a mask image onto a substrate. The four optical elements comprise, in order from object to image, convex, concave, convex and concave mirrors. The optical system is particularly suited for step and scan lithography methods. The invention enables the use of larger slit dimensions associated with ring field scanning optics, improves wafer throughput and allows higher semiconductor device density. The inventive optical system is characterized by reduced dynamic distortion because the static distortion is balanced across the slit width.

"First generation" automated DNA sequencing technology.

PubMed

Slatko, Barton E; Kieleczawa, Jan; Ju, Jingyue; Gardner, Andrew F; Hendrickson, Cynthia L; Ausubel, Frederick M

2011-10-01

Beginning in the 1980s, automation of DNA sequencing has greatly increased throughput, reduced costs, and enabled large projects to be completed more easily. The development of automation technology paralleled the development of other aspects of DNA sequencing: better enzymes and chemistry, separation and imaging technology, sequencing protocols, robotics, and computational advancements (including base-calling algorithms with quality scores, database developments, and sequence analysis programs). Despite the emergence of high-throughput sequencing platforms, automated Sanger sequencing technology remains useful for many applications. This unit provides background and a description of the "First-Generation" automated DNA sequencing technology. It also includes protocols for using the current Applied Biosystems (ABI) automated DNA sequencing machines. © 2011 by John Wiley & Sons, Inc.

UAV-based high-throughput phenotyping in legume crops

NASA Astrophysics Data System (ADS)

Sankaran, Sindhuja; Khot, Lav R.; Quirós, Juan; Vandemark, George J.; McGee, Rebecca J.

2016-05-01

In plant breeding, one of the biggest obstacles in genetic improvement is the lack of proven rapid methods for measuring plant responses in field conditions. Therefore, the major objective of this research was to evaluate the feasibility of utilizing high-throughput remote sensing technology for rapid measurement of phenotyping traits in legume crops. The plant responses of several chickpea and peas varieties to the environment were assessed with an unmanned aerial vehicle (UAV) integrated with multispectral imaging sensors. Our preliminary assessment showed that the vegetation indices are strongly correlated (p<0.05) with seed yield of legume crops. Results endorse the potential of UAS-based sensing technology to rapidly measure those phenotyping traits.

High Throughput, High Yield Fabrication of High Quantum Efficiency Back-Illuminated Photon Counting, Far UV, UV, and Visible Detector Arrays

NASA Technical Reports Server (NTRS)

Nikzad, Shouleh; Hoenk, M. E.; Carver, A. G.; Jones, T. J.; Greer, F.; Hamden, E.; Goodsall, T.

2013-01-01

In this paper we discuss the high throughput end-to-end post fabrication processing of high performance delta-doped and superlattice-doped silicon imagers for UV, visible, and NIR applications. As an example, we present our results on far ultraviolet and ultraviolet quantum efficiency (QE) in a photon counting, detector array. We have improved the QE by nearly an order of magnitude over microchannel plates (MCPs) that are the state-of-the-art UV detectors for many NASA space missions as well as defense applications. These achievements are made possible by precision interface band engineering of Molecular Beam Epitaxy (MBE) and Atomic Layer Deposition (ALD).

Goodman High Throughput Spectrograph | SOAR

Science.gov Websites

SPARTAN Near-IR Camera Ohio State Infrared Imager/Spectrograph (OSIRIS) - NO LONGER AVAILABLE SOAR 320-850 nm wavelength range. The paper describing the instrument is Clemens et al. (2004) Applying for IRAF. Publishing results based on Goodman data?: ADS link to 2004 SPIE Goodman Spectrograph paper

Using ToxCast data to reconstruct dynamic cell state trajectories and estimate toxicological points of departure

EPA Science Inventory

AbstractBackground. High-throughput in vitro screening is an important tool for evaluating the potential biological activity of the thousands of existing chemicals in commerce and the hundreds more introduced each year. Among the assay technologies available, high-content imaging...

RootScan: Software for high-throughput analysis of root anatomical traits

USDA-ARS?s Scientific Manuscript database

RootScan is a program for semi-automated image analysis of anatomical phenes in root cross-sections. RootScan uses pixel value thresholds to separate the cross-section from its background and to visually dissect it into tissue regions. Area measurements and object counts are performed within various...

The X-ray astronomy satellite ASCA

NASA Technical Reports Server (NTRS)

Tanaka, Yasuo; Inoue, Hajime; Holt, Stephen S.

1994-01-01

Advanced Satellite for Cosmology and Astrophysics (ASCA) is a high-throughput X-ray astronomy observatory which is capable of simultaneous imaging and spectroscopic observations over a wide energy range 0.5-10 keV. The scientific capabilities of ASCA and some aspects related to its operation and observations are briefly described.

A Dedicated Micro-Tomography Beamline For The Australian Synchrotron

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mayo, Sheridan C.; Stevenson, Andrew W.; Wilkins, Stephen W.

2010-07-23

A dedicated micro-tomography beamline is proposed for the Australian Synchrotron. It will enable high-resolution micro-tomography with resolution below a micron and supporting phase-contrast imaging modes. A key feature of the beamline will be high-throughput/high-speed operation enabling near real-time micro-tomography.

Development of rapid and sensitive high throughput pharmacologic assays for marine phycotoxins.

PubMed

Van Dolah, F M; Finley, E L; Haynes, B L; Doucette, G J; Moeller, P D; Ramsdell, J S

1994-01-01

The lack of rapid, high throughput assays is a major obstacle to many aspects of research on marine phycotoxins. Here we describe the application of microplate scintillation technology to develop high throughput assays for several classes of marine phycotoxin based on their differential pharmacologic actions. High throughput "drug discovery" format microplate receptor binding assays developed for brevetoxins/ciguatoxins and for domoic acid are described. Analysis for brevetoxins/ciguatoxins is carried out by binding competition with [3H] PbTx-3 for site 5 on the voltage dependent sodium channel in rat brain synaptosomes. Analysis of domoic acid is based on binding competition with [3H] kainic acid for the kainate/quisqualate glutamate receptor using frog brain synaptosomes. In addition, a high throughput microplate 45Ca flux assay for determination of maitotoxins is described. These microplate assays can be completed within 3 hours, have sensitivities of less than 1 ng, and can analyze dozens of samples simultaneously. The assays have been demonstrated to be useful for assessing algal toxicity and for assay-guided purification of toxins, and are applicable to the detection of biotoxins in seafood.

High-throughput continuous hydrothermal synthesis of nanomaterials (part II): unveiling the as-prepared CexZryYzO2-δ phase diagram.

PubMed

Quesada-Cabrera, Raul; Weng, Xiaole; Hyett, Geoff; Clark, Robin J H; Wang, Xue Z; Darr, Jawwad A

2013-09-09

High-throughput continuous hydrothermal flow synthesis was used to manufacture 66 unique nanostructured oxide samples in the Ce-Zr-Y-O system. This synthesis approach resulted in a significant increase in throughput compared to that of conventional batch or continuous hydrothermal synthesis methods. The as-prepared library samples were placed into a wellplate for both automated high-throughput powder X-ray diffraction and Raman spectroscopy data collection, which allowed comprehensive structural characterization and phase mapping. The data suggested that a continuous cubic-like phase field connects all three Ce-Zr-O, Ce-Y-O, and Y-Zr-O binary systems together with a smooth and steady transition between the structures of neighboring compositions. The continuous hydrothermal process led to as-prepared crystallite sizes in the range of 2-7 nm (as determined by using the Scherrer equation).

High-Throughput Density Measurement Using Magnetic Levitation.

PubMed

Ge, Shencheng; Wang, Yunzhe; Deshler, Nicolas J; Preston, Daniel J; Whitesides, George M

2018-06-20

This work describes the development of an integrated analytical system that enables high-throughput density measurements of diamagnetic particles (including cells) using magnetic levitation (MagLev), 96-well plates, and a flatbed scanner. MagLev is a simple and useful technique with which to carry out density-based analysis and separation of a broad range of diamagnetic materials with different physical forms (e.g., liquids, solids, gels, pastes, gums, etc.); one major limitation, however, is the capacity to perform high-throughput density measurements. This work addresses this limitation by (i) re-engineering the shape of the magnetic fields so that the MagLev system is compatible with 96-well plates, and (ii) integrating a flatbed scanner (and simple optical components) to carry out imaging of the samples that levitate in the system. The resulting system is compatible with both biological samples (human erythrocytes) and nonbiological samples (simple liquids and solids, such as 3-chlorotoluene, cholesterol crystals, glass beads, copper powder, and polymer beads). The high-throughput capacity of this integrated MagLev system will enable new applications in chemistry (e.g., analysis and separation of materials) and biochemistry (e.g., cellular responses under environmental stresses) in a simple and label-free format on the basis of a universal property of all matter, i.e., density.

Micro-patterned agarose gel devices for single-cell high-throughput microscopy of E. coli cells.

PubMed

Priest, David G; Tanaka, Nobuyuki; Tanaka, Yo; Taniguchi, Yuichi

2017-12-21

High-throughput microscopy of bacterial cells elucidated fundamental cellular processes including cellular heterogeneity and cell division homeostasis. Polydimethylsiloxane (PDMS)-based microfluidic devices provide advantages including precise positioning of cells and throughput, however device fabrication is time-consuming and requires specialised skills. Agarose pads are a popular alternative, however cells often clump together, which hinders single cell quantitation. Here, we imprint agarose pads with micro-patterned 'capsules', to trap individual cells and 'lines', to direct cellular growth outwards in a straight line. We implement this micro-patterning into multi-pad devices called CapsuleHotel and LineHotel for high-throughput imaging. CapsuleHotel provides ~65,000 capsule structures per mm 2 that isolate individual Escherichia coli cells. In contrast, LineHotel provides ~300 line structures per mm that direct growth of micro-colonies. With CapsuleHotel, a quantitative single cell dataset of ~10,000 cells across 24 samples can be acquired and analysed in under 1 hour. LineHotel allows tracking growth of > 10 micro-colonies across 24 samples simultaneously for up to 4 generations. These easy-to-use devices can be provided in kit format, and will accelerate discoveries in diverse fields ranging from microbiology to systems and synthetic biology.

Crop 3D-a LiDAR based platform for 3D high-throughput crop phenotyping.

PubMed

Guo, Qinghua; Wu, Fangfang; Pang, Shuxin; Zhao, Xiaoqian; Chen, Linhai; Liu, Jin; Xue, Baolin; Xu, Guangcai; Li, Le; Jing, Haichun; Chu, Chengcai

2018-03-01

With the growing population and the reducing arable land, breeding has been considered as an effective way to solve the food crisis. As an important part in breeding, high-throughput phenotyping can accelerate the breeding process effectively. Light detection and ranging (LiDAR) is an active remote sensing technology that is capable of acquiring three-dimensional (3D) data accurately, and has a great potential in crop phenotyping. Given that crop phenotyping based on LiDAR technology is not common in China, we developed a high-throughput crop phenotyping platform, named Crop 3D, which integrated LiDAR sensor, high-resolution camera, thermal camera and hyperspectral imager. Compared with traditional crop phenotyping techniques, Crop 3D can acquire multi-source phenotypic data in the whole crop growing period and extract plant height, plant width, leaf length, leaf width, leaf area, leaf inclination angle and other parameters for plant biology and genomics analysis. In this paper, we described the designs, functions and testing results of the Crop 3D platform, and briefly discussed the potential applications and future development of the platform in phenotyping. We concluded that platforms integrating LiDAR and traditional remote sensing techniques might be the future trend of crop high-throughput phenotyping.

A Barcoding Strategy Enabling Higher-Throughput Library Screening by Microscopy.

PubMed

Chen, Robert; Rishi, Harneet S; Potapov, Vladimir; Yamada, Masaki R; Yeh, Vincent J; Chow, Thomas; Cheung, Celia L; Jones, Austin T; Johnson, Terry D; Keating, Amy E; DeLoache, William C; Dueber, John E

2015-11-20

Dramatic progress has been made in the design and build phases of the design-build-test cycle for engineering cells. However, the test phase usually limits throughput, as many outputs of interest are not amenable to rapid analytical measurements. For example, phenotypes such as motility, morphology, and subcellular localization can be readily measured by microscopy, but analysis of these phenotypes is notoriously slow. To increase throughput, we developed microscopy-readable barcodes (MiCodes) composed of fluorescent proteins targeted to discernible organelles. In this system, a unique barcode can be genetically linked to each library member, making possible the parallel analysis of phenotypes of interest via microscopy. As a first demonstration, we MiCoded a set of synthetic coiled-coil leucine zipper proteins to allow an 8 × 8 matrix to be tested for specific interactions in micrographs consisting of mixed populations of cells. A novel microscopy-readable two-hybrid fluorescence localization assay for probing candidate interactions in the cytosol was also developed using a bait protein targeted to the peroxisome and a prey protein tagged with a fluorescent protein. This work introduces a generalizable, scalable platform for making microscopy amenable to higher-throughput library screening experiments, thereby coupling the power of imaging with the utility of combinatorial search paradigms.

Introduction of High Throughput Magnetic Resonance T2-Weighted Image Texture Analysis for WHO Grade 2 and 3 Gliomas.

PubMed

Kinoshita, Manabu; Sakai, Mio; Arita, Hideyuki; Shofuda, Tomoko; Chiba, Yasuyoshi; Kagawa, Naoki; Watanabe, Yoshiyuki; Hashimoto, Naoya; Fujimoto, Yasunori; Yoshimine, Toshiki; Nakanishi, Katsuyuki; Kanemura, Yonehiro

2016-01-01

Reports have suggested that tumor textures presented on T2-weighted images correlate with the genetic status of glioma. Therefore, development of an image analyzing framework that is capable of objective and high throughput image texture analysis for large scale image data collection is needed. The current study aimed to address the development of such a framework by introducing two novel parameters for image textures on T2-weighted images, i.e., Shannon entropy and Prewitt filtering. Twenty-two WHO grade 2 and 28 grade 3 glioma patients were collected whose pre-surgical MRI and IDH1 mutation status were available. Heterogeneous lesions showed statistically higher Shannon entropy than homogenous lesions (p = 0.006) and ROC curve analysis proved that Shannon entropy on T2WI was a reliable indicator for discrimination of homogenous and heterogeneous lesions (p = 0.015, AUC = 0.73). Lesions with well-defined borders exhibited statistically higher Edge mean and Edge median values using Prewitt filtering than those with vague lesion borders (p = 0.0003 and p = 0.0005 respectively). ROC curve analysis also proved that both Edge mean and median values were promising indicators for discrimination of lesions with vague and well defined borders and both Edge mean and median values performed in a comparable manner (p = 0.0002, AUC = 0.81 and p < 0.0001, AUC = 0.83, respectively). Finally, IDH1 wild type gliomas showed statistically lower Shannon entropy on T2WI than IDH1 mutated gliomas (p = 0.007) but no difference was observed between IDH1 wild type and mutated gliomas in Edge median values using Prewitt filtering. The current study introduced two image metrics that reflect lesion texture described on T2WI. These two metrics were validated by readings of a neuro-radiologist who was blinded to the results. This observation will facilitate further use of this technique in future large scale image analysis of glioma.

Toward a Low-Cost System for High-Throughput Image-Based Phenotyping of Root System Architecture

NASA Astrophysics Data System (ADS)

Davis, T. W.; Schneider, D. J.; Cheng, H.; Shaw, N.; Kochian, L. V.; Shaff, J. E.

2015-12-01

Root system architecture is being studied more closely for improved nutrient acquisition, stress tolerance and carbon sequestration by relating the genetic material that corresponds to preferential physical features. This information can help direct plant breeders in addressing the growing concerns regarding the global demand on crops and fossil fuels. To help support this incentive comes a need to make high-throughput image-based phenotyping of plant roots, at the individual plant scale, simpler and more affordable. Our goal is to create an affordable and portable product for simple image collection, processing and management that will extend root phenotyping to institutions with limited funding (e.g., in developing countries). Thus, a new integrated system has been developed using the Raspberry Pi single-board computer. Similar to other 3D-based imaging platforms, the system utilizes a stationary camera to photograph a rotating crop root system (e.g., rice, maize or sorghum) that is suspended either in a gel or on a mesh (for hydroponics). In contrast, the new design takes advantage of powerful open-source hardware and software to reduce the system costs, simplify the imaging process, and manage the large datasets produced by the high-resolution photographs. A newly designed graphical user interface (GUI) unifies the system controls (e.g., adjusting camera and motor settings and orchestrating the motor motion with image capture), making it easier to accommodate a variety of experiments. During each imaging session, integral metadata necessary for reproducing experiment results are collected (e.g., plant type and age, growing conditions and treatments, camera settings) using hierarchical data format files. These metadata are searchable within the GUI and can be selected and extracted for further analysis. The GUI also supports an image previewer that performs limited image processing (e.g., thresholding and cropping). Root skeletonization, 3D reconstruction and trait calculation (e.g., rooting depth, rooting angle, total volume of roots) is being developed in conjunction with this project.

Out-of-focal plane imaging by leakage radiation microscopy

NASA Astrophysics Data System (ADS)

Zhu, Liangfu; Zhang, Douguo; Wang, Ruxue; Wen, Xiaolei; Wang, Pei; Ming, Hai; Badugu, Ramachandram; Lakowicz, Joseph R.

2017-09-01

Leakage radiation microscopy (LRM) is used to investigate the optical properties of surfaces. The front-focal plane (FFP) image with LRM reveals the structural features on the surfaces. A back-focal plane (BFP) image with LRM reveals the angular distribution of the radiation. Herein, we experimentally demonstrate that the out-of-focal plane (OFP) images present a link between the FFP and BFP images and provide optical information that cannot be resolved by either FFP or BFP images. The OFP image provides a link between the spatial location of the emission and the angular distribution from the same location, and thus information about the film’s discontinuity, nonuniformity or variable thickness can be uncovered. The use of OFP imaging will extend the scope and applications of the LRM and coupled emission imaging, which are powerful tools in nanophotonics and high throughput fluorescence screening.

Simulation tools for analyzer-based x-ray phase contrast imaging system with a conventional x-ray source

NASA Astrophysics Data System (ADS)

Caudevilla, Oriol; Zhou, Wei; Stoupin, Stanislav; Verman, Boris; Brankov, J. G.

2016-09-01

Analyzer-based X-ray phase contrast imaging (ABI) belongs to a broader family of phase-contrast (PC) X-ray imaging modalities. Unlike the conventional X-ray radiography, which measures only X-ray absorption, in PC imaging one can also measures the X-rays deflection induced by the object refractive properties. It has been shown that refraction imaging provides better contrast when imaging the soft tissue, which is of great interest in medical imaging applications. In this paper, we introduce a simulation tool specifically designed to simulate the analyzer-based X-ray phase contrast imaging system with a conventional polychromatic X-ray source. By utilizing ray tracing and basic physical principles of diffraction theory our simulation tool can predicting the X-ray beam profile shape, the energy content, the total throughput (photon count) at the detector. In addition we can evaluate imaging system point-spread function for various system configurations.

Quantitative Image Restoration in Bright Field Optical Microscopy.

PubMed

Gutiérrez-Medina, Braulio; Sánchez Miranda, Manuel de Jesús

2017-11-07

Bright field (BF) optical microscopy is regarded as a poor method to observe unstained biological samples due to intrinsic low image contrast. We introduce quantitative image restoration in bright field (QRBF), a digital image processing method that restores out-of-focus BF images of unstained cells. Our procedure is based on deconvolution, using a point spread function modeled from theory. By comparing with reference images of bacteria observed in fluorescence, we show that QRBF faithfully recovers shape and enables quantify size of individual cells, even from a single input image. We applied QRBF in a high-throughput image cytometer to assess shape changes in Escherichia coli during hyperosmotic shock, finding size heterogeneity. We demonstrate that QRBF is also applicable to eukaryotic cells (yeast). Altogether, digital restoration emerges as a straightforward alternative to methods designed to generate contrast in BF imaging for quantitative analysis. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Optimizing ultrafast wide field-of-view illumination for high-throughput multi-photon imaging and screening of mutant fluorescent proteins

NASA Astrophysics Data System (ADS)

Stoltzfus, Caleb; Mikhailov, Alexandr; Rebane, Aleksander

2017-02-01

Fluorescence induced by 1wo-photon absorption (2PA) and three-photon absorption (3PA) is becoming an increasingly important tool for deep-tissue microscopy, especially in conjunction with genetically-encoded functional probes such as fluorescent proteins (FPs). Unfortunately, the efficacy of the multi-photon excitation of FPs is notoriously low, and because relations between a biological fluorophore's nonlinear-optical properties and its molecular structure are inherently complex, there are no practical avenues available that would allow boosting the performance of current FPs. Here we describe a novel method, where we apply directed evolution to optimize the 2PA properties of EGFP. Key to the success of this approach consists in high-throughput screening of mutants that would allow selection of variants with promising 2PA and 3PA properties in a broad near-IR excitation range of wavelength. For this purpose, we construct and test a wide field-of-view (FOV), femtosecond imaging system that we then use to quantify the multi-photon excited fluorescence in the 550- 1600 nm range of tens of thousands of E. coli colonies expressing randomly mutated FPs in a standard 10 cm diameter Petri dish configuration. We present a quantitative analysis of different factors that are currently limiting the maximum throughput of the femtosecond multi-photon screening techniques and also report on quantitative measurement of absolute 2PA and 3PA cross sections spectra.

Pixel Perfect

DOE Office of Scientific and Technical Information (OSTI.GOV)

Perrine, Kenneth A.; Hopkins, Derek F.; Lamarche, Brian L.

2005-09-01

Biologists and computer engineers at Pacific Northwest National Laboratory have specified, designed, and implemented a hardware/software system for performing real-time, multispectral image processing on a confocal microscope. This solution is intended to extend the capabilities of the microscope, enabling scientists to conduct advanced experiments on cell signaling and other kinds of protein interactions. FRET (fluorescence resonance energy transfer) techniques are used to locate and monitor protein activity. In FRET, it is critical that spectral images be precisely aligned with each other despite disturbances in the physical imaging path caused by imperfections in lenses and cameras, and expansion and contraction ofmore » materials due to temperature changes. The central importance of this work is therefore automatic image registration. This runs in a framework that guarantees real-time performance (processing pairs of 1024x1024, 8-bit images at 15 frames per second) and enables the addition of other types of advanced image processing algorithms such as image feature characterization. The supporting system architecture consists of a Visual Basic front-end containing a series of on-screen interfaces for controlling various aspects of the microscope and a script engine for automation. One of the controls is an ActiveX component written in C++ for handling the control and transfer of images. This component interfaces with a pair of LVDS image capture boards and a PCI board containing a 6-million gate Xilinx Virtex-II FPGA. Several types of image processing are performed on the FPGA in a pipelined fashion, including the image registration. The FPGA offloads work that would otherwise need to be performed by the main CPU and has a guaranteed real-time throughput. Image registration is performed in the FPGA by applying a cubic warp on one image to precisely align it with the other image. Before each experiment, an automated calibration procedure is run in order to set up the cubic warp. During image acquisitions, the cubic warp is evaluated by way of forward differencing. Unwanted pixelation artifacts are minimized by bilinear sampling. The resulting system is state-of-the-art for biological imaging. Precisely registered images enable the reliable use of FRET techniques. In addition, real-time image processing performance allows computed images to be fed back and displayed to scientists immediately, and the pipelined nature of the FPGA allows additional image processing algorithms to be incorporated into the system without slowing throughput.« less

Review of high fidelity imaging spectrometer design for remote sensing

NASA Astrophysics Data System (ADS)

Mouroulis, Pantazis; Green, Robert O.

2018-04-01

We review the design and assessment techniques that underlie a number of successfully deployed space and airborne imaging spectrometers that have been demonstrated to achieve demanding specifications in terms of throughput and response uniformity. The principles are illustrated with telescope designs as well as spectrometer examples from the Offner and Dyson families. We also show how the design space can be extended with the use of freeform surfaces and provide additional design examples with grating as well as prism dispersive elements.

Computational efficient segmentation of cell nuclei in 2D and 3D fluorescent micrographs

NASA Astrophysics Data System (ADS)

De Vylder, Jonas; Philips, Wilfried

2011-02-01

This paper proposes a new segmentation technique developed for the segmentation of cell nuclei in both 2D and 3D fluorescent micrographs. The proposed method can deal with both blurred edges as with touching nuclei. Using a dual scan line algorithm its both memory as computational efficient, making it interesting for the analysis of images coming from high throughput systems or the analysis of 3D microscopic images. Experiments show good results, i.e. recall of over 0.98.

ACS Data Handbook v.6.0

NASA Astrophysics Data System (ADS)

Gonzaga, S.; et al.

2011-03-01

ACS was designed to provide a deep, wide-field survey capability from the visible to near-IR using the Wide Field Camera (WFC), high resolution imaging from the near-UV to near-IR with the now-defunct High Resolution Camera (HRC), and solar-blind far-UV imaging using the Solar Blind Camera (SBC). The discovery efficiency of ACS's Wide Field Channel (i.e., the product of WFC's field of view and throughput) is 10 times greater than that of WFPC2. The failure of ACS's CCD electronics in January 2007 brought a temporary halt to CCD imaging until Servicing Mission 4 in May 2009, when WFC functionality was restored. Unfortunately, the high-resolution optical imaging capability of HRC was not recovered.

HTML5 PivotViewer: high-throughput visualization and querying of image data on the web

PubMed Central

Taylor, Stephen; Noble, Roger

2014-01-01

Motivation: Visualization and analysis of large numbers of biological images has generated a bottle neck in research. We present HTML5 PivotViewer, a novel, open source, platform-independent viewer making use of the latest web technologies that allows seamless access to images and associated metadata for each image. This provides a powerful method to allow end users to mine their data. Availability and implementation: Documentation, examples and links to the software are available from http://www.cbrg.ox.ac.uk/data/pivotviewer/. The software is licensed under GPLv2. Contact:  stephen.taylor@imm.ox.ac.uk and roger@coritsu.com PMID:24849578

Imaging and identification of waterborne parasites using a chip-scale microscope.

PubMed

Lee, Seung Ah; Erath, Jessey; Zheng, Guoan; Ou, Xiaoze; Willems, Phil; Eichinger, Daniel; Rodriguez, Ana; Yang, Changhuei

2014-01-01

We demonstrate a compact portable imaging system for the detection of waterborne parasites in resource-limited settings. The previously demonstrated sub-pixel sweeping microscopy (SPSM) technique is a lens-less imaging scheme that can achieve high-resolution (<1 µm) bright-field imaging over a large field-of-view (5.7 mm×4.3 mm). A chip-scale microscope system, based on the SPSM technique, can be used for automated and high-throughput imaging of protozoan parasite cysts for the effective diagnosis of waterborne enteric parasite infection. We successfully imaged and identified three major types of enteric parasite cysts, Giardia, Cryptosporidium, and Entamoeba, which can be found in fecal samples from infected patients. We believe that this compact imaging system can serve well as a diagnostic device in challenging environments, such as rural settings or emergency outbreaks.

Time-Correlated Single-Photon Counting Fluorescence Imaging of Lipid Domains In Raft-Mimicking Giant Unilamellar Vesicles

NASA Astrophysics Data System (ADS)

Clarke, James; Cheng, Kwan; Shindell, Orrin; Wang, Exing

We have designed and constructed a high-throughput electrofusion chamber and an incubator to fabricate Giant Unilamellar Vesicles (GUVs) consisting of high-melting lipids, low-melting lipids, cholesterol and both ordered and disordered phase sensitive fluorescent probes (DiIC12, dehydroergosterol and BODIPY-Cholesterol). GUVs were formed in a 3 stage pulse sequence electrofusion process with voltages ranging from 50mVpp to 2.2Vpp and frequencies from 5Hz to 10Hz. Steady state and time-correlated single-photon counting (TCSPC) fluorescence lifetime (FLIM) based confocal and/or multi-photon microscopic techniques were used to characterize phase separated lipid domains in GUVs. Confocal imaging measures the probe concentration and the chemical environment of the system. TCSPC techniques determine the chemical environment through the perturbation of fluorescent lifetimes of the probes in the system. The above techniques will be applied to investigate the protein-lipid interactions involving domain formation. Specifically, the mechanisms governing lipid domain formations in the above systems that mimic the lipid rafts in cells will be explored. Murchison Fellowship at Trinity University.

Analysis and Testing of Mobile Wireless Networks

NASA Technical Reports Server (NTRS)

Alena, Richard; Evenson, Darin; Rundquist, Victor; Clancy, Daniel (Technical Monitor)

2002-01-01

Wireless networks are being used to connect mobile computing elements in more applications as the technology matures. There are now many products (such as 802.11 and 802.11b) which ran in the ISM frequency band and comply with wireless network standards. They are being used increasingly to link mobile Intranet into Wired networks. Standard methods of analyzing and testing their performance and compatibility are needed to determine the limits of the technology. This paper presents analytical and experimental methods of determining network throughput, range and coverage, and interference sources. Both radio frequency (BE) domain and network domain analysis have been applied to determine wireless network throughput and range in the outdoor environment- Comparison of field test data taken under optimal conditions, with performance predicted from RF analysis, yielded quantitative results applicable to future designs. Layering multiple wireless network- sooners can increase performance. Wireless network components can be set to different radio frequency-hopping sequences or spreading functions, allowing more than one sooner to coexist. Therefore, we ran multiple 802.11-compliant systems concurrently in the same geographical area to determine interference effects and scalability, The results can be used to design of more robust networks which have multiple layers of wireless data communication paths and provide increased throughput overall.

Multi-angle lensless digital holography for depth resolved imaging on a chip.

PubMed

Su, Ting-Wei; Isikman, Serhan O; Bishara, Waheb; Tseng, Derek; Erlinger, Anthony; Ozcan, Aydogan

2010-04-26

A multi-angle lensfree holographic imaging platform that can accurately characterize both the axial and lateral positions of cells located within multi-layered micro-channels is introduced. In this platform, lensfree digital holograms of the micro-objects on the chip are recorded at different illumination angles using partially coherent illumination. These digital holograms start to shift laterally on the sensor plane as the illumination angle of the source is tilted. Since the exact amount of this lateral shift of each object hologram can be calculated with an accuracy that beats the diffraction limit of light, the height of each cell from the substrate can be determined over a large field of view without the use of any lenses. We demonstrate the proof of concept of this multi-angle lensless imaging platform by using light emitting diodes to characterize various sized microparticles located on a chip with sub-micron axial and lateral localization over approximately 60 mm(2) field of view. Furthermore, we successfully apply this lensless imaging approach to simultaneously characterize blood samples located at multi-layered micro-channels in terms of the counts, individual thicknesses and the volumes of the cells at each layer. Because this platform does not require any lenses, lasers or other bulky optical/mechanical components, it provides a compact and high-throughput alternative to conventional approaches for cytometry and diagnostics applications involving lab on a chip systems.

High-throughput determination of biochemical oxygen demand (BOD) by a microplate-based biosensor.

PubMed

Pang, Hei-Leung; Kwok, Nga-Yan; Chan, Pak-Ho; Yeung, Chi-Hung; Lo, Waihung; Wong, Kwok-Yin

2007-06-01

The use of the conventional 5-day biochemical oxygen demand (BOD5) method in BOD determination is greatly hampered by its time-consuming sampling procedure and its technical difficulty in the handling of a large pool of wastewater samples. Thus, it is highly desirable to develop a fast and high-throughput biosensor for BOD measurements. This paper describes the construction of a microplate-based biosensor consisting of an organically modified silica (ORMOSIL) oxygen sensing film for high-throughput determination of BOD in wastewater. The ORMOSIL oxygen sensing film was prepared by reacting tetramethoxysilane with dimethyldimethoxysilane in the presence of the oxygen-sensitive dye tris(4,7-diphenyl-1,10-phenanthroline)ruthenium-(II) chloride. The silica composite formed a homogeneous, crack-free oxygen sensing film on polystyrene microtiter plates with high stability, and the embedded ruthenium dye interacted with the dissolved oxygen in wastewater according to the Stern-Volmer relation. The bacterium Stenotrophomonas maltophilia was loaded into the ORMOSIL/ PVA composite (deposited on the top of the oxygen sensing film) and used to metabolize the organic compounds in wastewater. This BOD biosensor was found to be able to determine the BOD values of wastewater samples within 20 min by monitoring the dissolved oxygen concentrations. Moreover, the BOD values determined by the BOD biosensor were in good agreement with those obtained by the conventional BOD5 method.

An improved method for differentiating cell-bound from internalized particles by imaging flow cytometry.

PubMed

Smirnov, Asya; Solga, Michael D; Lannigan, Joanne; Criss, Alison K

2015-08-01

Recognition, binding, internalization, and elimination of pathogens and cell debris are important functions of professional as well as non-professional phagocytes. However, high-throughput methods for quantifying cell-associated particles and discriminating bound from internalized particles have been lacking. Here we describe a protocol for using imaging flow cytometry to quantify the attached and phagocytosed particles that are associated with a population of cells. Cells were exposed to fluorescent particles, fixed, and exposed to an antibody of a different fluorophore that recognizes the particles. The antibody is added without cell permeabilization, such that the antibody only binds extracellular particles. Cells with and without associated particles were identified by imaging flow cytometry. For each cell with associated particles, a spot count algorithm was employed to quantify the number of extracellular (double fluorescent) and intracellular (single fluorescent) particles per cell, from which the percent particle internalization was determined. The spot count algorithm was empirically validated by examining the fluorescence and phase contrast images acquired by the flow cytometer. We used this protocol to measure binding and internalization of the bacterium Neisseria gonorrhoeae by primary human neutrophils, using different bacterial variants and under different cellular conditions. The results acquired using imaging flow cytometry agreed with findings that were previously obtained using conventional immunofluorescence microscopy. This protocol provides a rapid, powerful method for measuring the association and internalization of any particle by any cell type. Copyright © 2015 Elsevier B.V. All rights reserved.

Micropatterned comet assay enables high throughput and sensitive DNA damage quantification

PubMed Central

Ge, Jing; Chow, Danielle N.; Fessler, Jessica L.; Weingeist, David M.; Wood, David K.; Engelward, Bevin P.

2015-01-01

The single cell gel electrophoresis assay, also known as the comet assay, is a versatile method for measuring many classes of DNA damage, including base damage, abasic sites, single strand breaks and double strand breaks. However, limited throughput and difficulties with reproducibility have limited its utility, particularly for clinical and epidemiological studies. To address these limitations, we created a microarray comet assay. The use of a micrometer scale array of cells increases the number of analysable comets per square centimetre and enables automated imaging and analysis. In addition, the platform is compatible with standard 24- and 96-well plate formats. Here, we have assessed the consistency and sensitivity of the microarray comet assay. We showed that the linear detection range for H2O2-induced DNA damage in human lymphoblastoid cells is between 30 and 100 μM, and that within this range, inter-sample coefficient of variance was between 5 and 10%. Importantly, only 20 comets were required to detect a statistically significant induction of DNA damage for doses within the linear range. We also evaluated sample-to-sample and experiment-to-experiment variation and found that for both conditions, the coefficient of variation was lower than what has been reported for the traditional comet assay. Finally, we also show that the assay can be performed using a 4× objective (rather than the standard 10× objective for the traditional assay). This adjustment combined with the microarray format makes it possible to capture more than 50 analysable comets in a single image, which can then be automatically analysed using in-house software. Overall, throughput is increased more than 100-fold compared to the traditional assay. Together, the results presented here demonstrate key advances in comet assay technology that improve the throughput, sensitivity, and robustness, thus enabling larger scale clinical and epidemiological studies. PMID:25527723

Micropatterned comet assay enables high throughput and sensitive DNA damage quantification.

PubMed

Ge, Jing; Chow, Danielle N; Fessler, Jessica L; Weingeist, David M; Wood, David K; Engelward, Bevin P

2015-01-01

The single cell gel electrophoresis assay, also known as the comet assay, is a versatile method for measuring many classes of DNA damage, including base damage, abasic sites, single strand breaks and double strand breaks. However, limited throughput and difficulties with reproducibility have limited its utility, particularly for clinical and epidemiological studies. To address these limitations, we created a microarray comet assay. The use of a micrometer scale array of cells increases the number of analysable comets per square centimetre and enables automated imaging and analysis. In addition, the platform is compatible with standard 24- and 96-well plate formats. Here, we have assessed the consistency and sensitivity of the microarray comet assay. We showed that the linear detection range for H2O2-induced DNA damage in human lymphoblastoid cells is between 30 and 100 μM, and that within this range, inter-sample coefficient of variance was between 5 and 10%. Importantly, only 20 comets were required to detect a statistically significant induction of DNA damage for doses within the linear range. We also evaluated sample-to-sample and experiment-to-experiment variation and found that for both conditions, the coefficient of variation was lower than what has been reported for the traditional comet assay. Finally, we also show that the assay can be performed using a 4× objective (rather than the standard 10× objective for the traditional assay). This adjustment combined with the microarray format makes it possible to capture more than 50 analysable comets in a single image, which can then be automatically analysed using in-house software. Overall, throughput is increased more than 100-fold compared to the traditional assay. Together, the results presented here demonstrate key advances in comet assay technology that improve the throughput, sensitivity, and robustness, thus enabling larger scale clinical and epidemiological studies. © The Author 2014. Published by Oxford University Press on behalf of the Mutagenesis Society. All rights reserved. For permissions, please email: journals.permissions@oup.com.

NiftyPET: a High-throughput Software Platform for High Quantitative Accuracy and Precision PET Imaging and Analysis.

PubMed

Markiewicz, Pawel J; Ehrhardt, Matthias J; Erlandsson, Kjell; Noonan, Philip J; Barnes, Anna; Schott, Jonathan M; Atkinson, David; Arridge, Simon R; Hutton, Brian F; Ourselin, Sebastien

2018-01-01

We present a standalone, scalable and high-throughput software platform for PET image reconstruction and analysis. We focus on high fidelity modelling of the acquisition processes to provide high accuracy and precision quantitative imaging, especially for large axial field of view scanners. All the core routines are implemented using parallel computing available from within the Python package NiftyPET, enabling easy access, manipulation and visualisation of data at any processing stage. The pipeline of the platform starts from MR and raw PET input data and is divided into the following processing stages: (1) list-mode data processing; (2) accurate attenuation coefficient map generation; (3) detector normalisation; (4) exact forward and back projection between sinogram and image space; (5) estimation of reduced-variance random events; (6) high accuracy fully 3D estimation of scatter events; (7) voxel-based partial volume correction; (8) region- and voxel-level image analysis. We demonstrate the advantages of this platform using an amyloid brain scan where all the processing is executed from a single and uniform computational environment in Python. The high accuracy acquisition modelling is achieved through span-1 (no axial compression) ray tracing for true, random and scatter events. Furthermore, the platform offers uncertainty estimation of any image derived statistic to facilitate robust tracking of subtle physiological changes in longitudinal studies. The platform also supports the development of new reconstruction and analysis algorithms through restricting the axial field of view to any set of rings covering a region of interest and thus performing fully 3D reconstruction and corrections using real data significantly faster. All the software is available as open source with the accompanying wiki-page and test data.

Visualization of small lesions in rat cartilage by means of laboratory-based x-ray phase contrast imaging

NASA Astrophysics Data System (ADS)

Marenzana, Massimo; Hagen, Charlotte K.; Das Neves Borges, Patricia; Endrizzi, Marco; Szafraniec, Magdalena B.; Ignatyev, Konstantin; Olivo, Alessandro

2012-12-01

Being able to quantitatively assess articular cartilage in three-dimensions (3D) in small rodent animal models, with a simple laboratory set-up, would prove extremely important for the development of pre-clinical research focusing on cartilage pathologies such as osteoarthritis (OA). These models are becoming essential tools for the development of new drugs for OA, a disease affecting up to 1/3 of the population older than 50 years for which there is no cure except prosthetic surgery. However, due to limitations in imaging technology, high-throughput 3D structural imaging has not been achievable in small rodent models, thereby limiting their translational potential and their efficiency as research tools. We show that a simple laboratory system based on coded-aperture x-ray phase contrast imaging (CAXPCi) can correctly visualize the cartilage layer in slices of an excised rat tibia imaged both in air and in saline solution. Moreover, we show that small, surgically induced lesions are also correctly detected by the CAXPCi system, and we support this finding with histopathology examination. Following these successful proof-of-concept results in rat cartilage, we expect that an upgrade of the system to higher resolutions (currently underway) will enable extending the method to the imaging of mouse cartilage as well. From a technological standpoint, by showing the capability of the system to detect cartilage also in water, we demonstrate phase sensitivity comparable to other lab-based phase methods (e.g. grating interferometry). In conclusion, CAXPCi holds a strong potential for being adopted as a routine laboratory tool for non-destructive, high throughput assessment of 3D structural changes in murine articular cartilage, with a possible impact in the field similar to the revolution that conventional microCT brought into bone research.

Fuzzy Logic-based expert system for evaluating cake quality of freeze-dried formulations.

PubMed

Trnka, Hjalte; Wu, Jian X; Van De Weert, Marco; Grohganz, Holger; Rantanen, Jukka

2013-12-01

Freeze-drying of peptide and protein-based pharmaceuticals is an increasingly important field of research. The diverse nature of these compounds, limited understanding of excipient functionality, and difficult-to-analyze quality attributes together with the increasing importance of the biosimilarity concept complicate the development phase of safe and cost-effective drug products. To streamline the development phase and to make high-throughput formulation screening possible, efficient solutions for analyzing critical quality attributes such as cake quality with minimal material consumption are needed. The aim of this study was to develop a fuzzy logic system based on image analysis (IA) for analyzing cake quality. Freeze-dried samples with different visual quality attributes were prepared in well plates. Imaging solutions together with image analytical routines were developed for extracting critical visual features such as the degree of cake collapse, glassiness, and color uniformity. On the basis of the IA outputs, a fuzzy logic system for analysis of these freeze-dried cakes was constructed. After this development phase, the system was tested with a new screening well plate. The developed fuzzy logic-based system was found to give comparable quality scores with visual evaluation, making high-throughput classification of cake quality possible. © 2013 Wiley Periodicals, Inc. and the American Pharmacists Association.