Charge-coupled Device Imaging System Research Articles

A Morphing Point-to-Point Displacement Control Based on Long Short-Term Memory for a Coplanar XXY Stage.

In this study, visual recognition with a charge-coupled device (CCD) image feedback control system was used to record the movement of a coplanar XXY stage. The position of the stage is fedback through the image positioning method, and the positioning compensation of the stage is performed by the image compensation control parameter. The image resolution was constrained and resulted in an average positioning error of the optimized control parameter of 6.712 µm, with the root mean square error being 2.802 µm, and the settling time being approximately 7 s. The merit of a long short-term memory (LSTM) deep learning model is that it can identify long-term dependencies and sequential state data to determine the next control signal. As for improving the positioning performance, LSTM was used to develop a training model for stage motion with an additional dial indicator with an accuracy of 1 μm being used to record the XXY position information. After removing the assisting dial indicator, a new LSTM-based XXY feedback control system was subsequently constructed to reduce the positioning error. In other words, the morphing control signals are dependent not only on time, but also on the iterations of the LSTM learning process. Point-to-point commanded forward, backward and repeated back-and-forth repetitive motions were conducted. Experimental results revealed that the average positioning error achieved after using the LSTM model was 2.085 µm, with the root mean square error being 2.681 µm, and a settling time of 2.02 s. With the assistance of LSTM, the stage exhibited a higher control accuracy and less settling time than did the CCD imaging system according to three positioning indices.

Kinetic Properties of CO2 Hydrate Formation in the Wax-Containing System at Different Concentrations

Precipitation of wax crystals and hydrate formation during deep-sea multiphase pipeline transportation would limit pipeline transportation flow and even trigger accidents. It is necessary to study the hydrate formation behavior in the wax-containing system. This paper utilized a mixture of number 60 Kunlun paraffin wax and number 2 industrial white oil to simulate the wax-containing system, and the CO2 hydrate was formed in the wax-containing system. The morphology of CO2 hydrate formation was captured by the microscope and the charge-coupled device imaging system. In this study, the influences of the wax crystal concentrations on the induction time and cumulative gas consumption of hydrate formation were investigated, and the effects of wax crystal precipitation on the CO2 hydrate nucleation mechanism were discussed. The experimental results indicated that, for the wax crystal concentrations of 1.50 and 2.50 wt %, the induction time of CO2 hydrate formation in the wax-containing system was reduced by 29.23 and 69.23% compared to that in the pure water system, respectively. However, for the wax crystal concentration of 0.50 wt %, the induction time of CO2 hydrate formation in the wax-containing system was increased by 130.77% compared to that in the pure water system. The reason for this was that the precipitation of wax crystals at high concentrations increased the chance of heterogeneous nucleation and the probability of heterogeneous nucleation increased with the increase of wax crystal concentrations. However, for the wax crystals at low concentrations, it not only lowered the chance of cross nucleation but also increased the mass transfer resistance for gas diffusion, thereby resulting in the increase of the induction time of hydrate formation. It was also found that, with the increase of the wax crystal concentrations, the cumulative gas consumption during hydrate formation decreased gradually. When the wax crystal concentration increased to 2.50 wt %, the cumulative gas consumption was reduced by 32.96% compared to that of the pure water system. The experimental results obtained in this study were helpful for understanding the kinetic characteristics of hydrate formation in the wax-containing system, which is meaningful for the pipeline flow assurance, safety, and economic security of pipeline transportation as well as the development of carbon capture and storage.

Gas Absorption and Mass Transfer in a Pore-Array Intensified Tube-in-Tube Microchannel

A pore-array intensified tube-in-tube microchannel (PA-TMC), which is characterized by high throughput and low pressure drop, was developed as a gas–liquid contactor. The sulfite oxidation method was used to determine the oxygen efficiency (φ) and volumetric mass transfer coefficient (kLa) of PA-TMC, and the mass transfer amount per unit energy (e) was calculated by using the pressure drop. The effects of structural and operating parameters were investigated systematically, and the two-phase flow behavior was monitored by using a charge-coupled device imaging system. The results indicated that the gas absorption efficiency and mass transfer performance of the PA-TMC were improved with increasing pore number, flow rate, and number of helical coil turns and decreasing pore size, row number, annular size, annular length, and surface tension. The φ, e and kLa of PA-TMC could reach 31.3%, 1.73 × 10−4 mol/J, and 7.0 s−1, respectively. The Sherwood number was correlated with the investigated parameters to guide the design of PA-TMC in gas absorption and mass transfer processes.

Luciferase Complementation Assay for Protein‐Protein Interactions in Plants

Constitutive and dynamic protein-protein interactions are fundamental to all aspects of cellular processes. Compared to other techniques measuring protein-protein interactions in plants, the luciferase complementation assay has a number of advantages: it detects plant protein-protein interactions in real time, requires little hands-on manipulation of samples, is highly quantitative, has extremely low background, and can be easily scaled up for high-throughput interactome studies. Here, we describe a protocol that includes two alternate data collection methods to quantify luminescence results based on Agrobacterium-mediated transient luciferase expression in Nicotiana benthamiana. One data collection method employs a charge-coupled device imaging system that allows the interactions to be presented as images, and the other employs a luminometer, which enables the assay to be conducted in a 96-well plate. Technical parameters related to frequently encountered problems and common errors, presented here, are important for performing this assay successfully. © 2018 by John Wiley & Sons, Inc.

An improved adaptive preprocessing method for TDI CCD images

In order to achieve high quality images with time-delayed integration (TDI) charge-coupled device (CCD) imaging system, an improved adaptive preprocessing method is proposed with functions of both denoising and edge enhancement. It is a weighted average filter integrating the average filter and the improved range filter. The weighted factors are deduced in terms of a cost function, which are adjustable to different images. To validate the proposed method, extensive tests are carried out on a developed TDI CCD imaging system. The experimental results confirm that this preprocessing method can fulfill the noise removal and edge sharpening simultaneously, which can play an important role in remote sensing field.

Noise Model of a Multispectral TDI CCD Imaging System and Its Parameter Estimation of Piecewise Weighted Least Square Fitting

The time-delayed integration (TDI) charge-coupled device (CCD) imaging technology is widely used in remote sensing field and accurate estimation of noise level is essential to assure good performance of denoising. Therefore, in order to achieve images with high signal-to-noise ratio by the TDI CCD sensor, the noise model of the imaging system is investigated. First, the design of the imaging circuit system is presented, and the main noises of the system are analyzed, such as the TDI CCD sensor, amplifiers, and so on. Then a theoretical noise model of the imaging system is established as a priori information. An improved least square fitting method of parameter estimation is proposed. To validate the theoretical model and the proposed method, extensive tests are carried out to obtain multiple uniform images on a developed TDI CCD imaging system. The experiment results confirm that a precise noise model can be achieved by this method and it verifies the theoretical model, meanwhile. It can be used for noise removal and aided design of imaging systems to acquire high-quality images. It is meaningful for remote sensing to obtain more targets of interest.

Laser Ablation Transient Plasma Structures Expansion in Vacuum

The dynamics of transient plasmas generated by high-fluence nanosecond laser ablation has been investigated by means of a fast intensified charge-coupled device imaging system. The recorded pictures reveal a specific splitting process of the plasma plume into fast and slow components.

An Optimized Telomerase-Specific Lentivirus for Optical Imaging of Tumors

Advances in medical imaging techniques, such as ultrasound, computed tomography, magnetic resonance imaging, and positron emission tomography, have made great progress in detecting tumors. However, these imaging techniques are unable to differentiate malignant tumors from benign ones. Recently developed optical imaging of tumors in small animals provides a useful method to distinguish malignant tumors from their surrounding normal tissues. Human telomerase reverse transcriptase (hTERT) is normally inactivated in most somatic cells, whereas it is commonly reactivated in many cancer cells. In this study, we constructed a lentiviral vector that expresses green fluorescent protein (GFP) driven by an optimized hTERT promoter to create a noninvasive tumor-specific imaging methodology. The activity of this optimized hTERT promoter was found to be equal to the activity of SV40 and cytomegalovirus promoters. In vitro experiments showed that GFP was only expressed in telomerase-positive tumor cells infected with this lentivirus, whereas there was no GFP expression in telomerase-negative tumor cells or normal somatic cells. We also found that subcutaneous telomerase-positive tumors could be visualized 24 hours after an intratumoral injection with this lentivirus by using a charge-coupled device (CCD) camera. In contrast, telomerase-negative tumors could not be imaged after an intratumoral injection even for 30 days. These results suggest that infection with lentivirus containing this optimized hTERT promoter might be a useful diagnostic tool for the real-time visualization of macroscopically invisible tumor tissues using a highly sensitive CCD imaging system.

T helper17 Cells Are Sufficient But Not Necessary to Induce Acute Graft-Versus-Host Disease

T helper (Th)1 cells were considered responsible for the induction of graft-versus-host disease (GVHD), but recently the concept has been challenged. Th17 cells play a critical role in mediating autoimmune diseases, but their role in the pathogenesis of GVHD remains unclear. Herein we compare the ability of in vitro generated Th1 and Th17 cells from C57BL/6 mice to induce GVHD in lethally irradiated BALB/c recipients. Allogeneic Th17 cells had superior expansion and infiltration capabilities in GVHD target organs, which correlated with their increased pathogenicity when compared with naïve or Th1 controls. Th17 cells caused no pathology in the syngeneic recipients, indicating that antigen-activation was required for their pathogenicity. Polarized Th17 cells could not maintain their phenotype in vivo as they produced a significant amount of interferon (IFN)-gamma after being transplanted into allogeneic recipients; however, IFN-gamma was not required for Th17 cell-induced GVHD. Further, we evaluated the pathogenesis of Th17 cells in GVHD by using polyclonal nonprimed CD4T cells in a clinically relevant allogeneic bone marrow transplantation (BMT) setting. We found that disruption of Th17-differentiation alone by targeting RORgammat (Th17-specific transcription factor) had no significant effect on GVHD development. We conclude that Th17 cells are sufficient but not necessary to induce GVHD.

Genetically encoded optical probes for imaging cellular signaling pathways

The intracellular signaling can be monitored in vivo in living cells by genetically encoded intracellular fluorescent and bioluminescent probes or indicators, which include second messengers, protein phosphorylation, protein conformational changes, protein-protein interactions, and protein localizations. These probes are of general use not only for fundamental biological studies, but also for assay and screening of possible pharmaceutical or toxic chemicals that inhibit or facilitate cellular signaling pathways. In this review, two examples of such indicators were briefly introduced. First, a genetically encoded fluorescent indicator was described for the detection and characterization of estrogen agonists and antagonists. The indicator was named SCCoR (single cell-coactivator recruitment). The high sensitivity of the present indicator made it possible to distinguish between estrogen strong and weak agonists in a dose-dependent fashion, immediately after adding a ligand to live cells. Discrimination of agonists from antagonists was efficiently achieved using the indicator. The approach described here can be applied to develop biosensors for other hormone receptors as well. Another example herein is a genetically encoded bioluminescent indicator for monitoring the nuclear trafficking of target proteins in vitro and in vivo. 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.

High-throughput sensing and noninvasive imaging of protein nuclear transport by using reconstitution of split Renilla luciferase.

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.

A cell-based microarrayed compound screening format for identifying agonists of G-protein-coupled receptors

The identification of agonist and antagonist leads for G-protein-coupled receptors (GPCRs) is of critical importance to the pharmaceutical and biotechnology industries. We report on the utilization of a novel, high-density, well-less screening platform known as microarrayed compound screening (μARCS) that tests 8640 compounds in the footprint of a standard microtiter plate for the identification of novel agonists for a specific G-protein-coupled receptor. Although receptors coupled to the Gα q protein can readily be assessed by fluorescence-based Ca 2+ release measurements, many GPCRs that are coupled to Gα s or Gα i/o proteins are not amenable to functional evaluation in such a high-throughput manner. In this study, the human dopamine D 4.4 receptor, which normally couples through the Gα i/o protein to inhibit adenylate cyclase and to reduce levels of intracellular cAMP, was coupled to intracellular Ca 2+ release by stably coexpressing this receptor with a chimeric G αqo5 protein in HEK-293 cells. In μARCS format, the cells expressing D 4.4 receptor and Gα qo5 protein were preloaded with fluo-4, cast into a 1% agarose gel, placed above the compound sheets, and imaged successively using a ViewLux charge-coupled device imaging system. Dopamine and other agonists evoked an increase in fluorescence response that appeared as bright spots in a time- and concentration-dependent manner. Utilizing this technology, a library of 260,000 compounds was rapidly screened and led to the identification of several novel agonists. These agonists were further characterized using a fluorometric imaging plate reader assay. Excellent confirmation rates coupled with enhanced efficiency and throughput enable μARCS to serve as an alternative platform for the screening and identification of novel GPCR agonists.

Chemiluminescent-based methods to detect subpicomole levels of c-type cytochromes

A variety of luminol-based substrates and either an autoradiographic film or a charge-coupled device (CCD) imaging system were used for chemiluminescence detection of c-type cytochromes. The Pierce Femto peroxidase substrate was at least 10 times more sensitive when using film than the highly cited 3,3 ′,5,5 ′-tetramethylbenzidine (benzidine derivative) staining method and 50 times more sensitive when using a CCD imaging system. Film or CCD imaging result in highly sensitive and quantitative signals. The quantitative detection of c-type cytochromes from single colonies or from less than 1 ml of a bacterial culture is possible.

Application of Micro Arrayed Compound Screening (μARCS) to Identify Inhibitors of Caspase-3

Micro Arrayed Compound Screening (microARCS) is a miniaturized ultra-high-throughput screening platform developed at Abbott Laboratories. In this format, 8,640 discrete compounds are spotted and dried onto a polystyrene sheet, which has the same footprint as a 96-well plate. A homogeneous time-resolved fluorescence assay format (LANCE) was applied to identify the inhibitors of caspase-3 using a peptide substrate labeled with a fluorescent europium chelate and a dabcyl quencher. The caspase-3 enzyme was cast into a thin agarose gel, which was placed on a sheet containing test compounds. A second gel containing caspase substrate was then laid above the enzyme gel to initiate the reaction. Caspase-3 cleaves the substrate and separates the europium from the quencher, giving rise to a time-resolved fluorescent signal, which was detected using a ViewLux charge-coupled device imaging system. Potential inhibitors of caspase-3 appeared as dark spots on a bright fluorescent background. Results from the microARCS assay format were compared to those from a conventional 96-well plate-screening format.

Visualization of advanced human prostate cancer lesions in living mice by a targeted gene transfer vector and optical imaging.

Non-invasive imaging and transcriptional targeting can improve the safety of therapeutic approaches in cancer. Here we demonstrate the ability to identify metastases in a human-prostate cancer model, employing a prostate-specific adenovirus vector (AdPSE-BC-luc) and a charge-coupled device-imaging system. AdPSE-BC-luc, which expresses firefly luciferase from an enhanced prostate-specific antigen promoter, restricted expression in the liver but produced robust signals in prostate tumors. In fact, expression was higher in advanced, androgen-independent tumors than in androgen-dependent lesions. Repetitive imaging over a three-week period after AdPSE-BC-luc injection into tumor-bearing mice revealed that the virus could locate and illuminate metastases in the lung and spine. Systemic injection of low doses of AdPSE-BC-luc illuminated lung metastasis. These results demonstrate the potential use of a non-invasive imaging modality in therapeutic and diagnostic strategies to manage prostate cancer.

Novel technique for low-jitter dual-laser synchronization in a thin film deposition system

The need for precise laser pulse synchronization in a dual-laser ablation system to optimize the quality of the deposited thin films has been previously demonstrated. We present, in this article, a novel technique for the synchronization of an excimer and a CO2 laser with synchronization having a temporal fluctuation (jitter) of less than ±14 ns. This is several times better than the best precision of temporal synchronization possible using traditional electronic techniques and is crucial for the application of dual-laser ablation in the manufacturing of thin films. Evidence for reproducibility in the ablation of targets using this system is presented by analyzing the initial stages of the ablated plasma using a time-gated charge coupled device imaging system.

Factors affecting quantitation of DNA bands in gels using a charge-coupled device imaging system.

Quantitation of DNA bands separated in polyacrylamide or agarose gels was tested under a variety of conditions to examine key factors contributing to the ability to obtain quantitative data. Variations tested included comparison of different fluorescent stains (ethidium bromide and GelStar stain), and variations of other parameters relating to the electrophoretic separation, e.g. gel and sample geometry, mode of staining, and mode of excitation of stains. The results showed that linear results were seen for a similar 20-30-fold range of DNA concentrations with both stains tested and that it is critical to standardize separations to obtain consistent results. Variations in separation and detection could lead to relatively large changes in fluorescent signals seen for similar amounts of DNA.

Alis, a state-of-the-art optical observation network for the exploration of polar atmospheric processes

Abstract An optical group at the Swedish Institute of Space Physics in Kiruna, Sweden has been developing the ALIS (Auroral Large Imaging System) multi-station optical observing network which makes it possible to obtain composite monochromatic 2-D images over a fairly wide field-of-view (FOV), and more interestingly, a CT (Computed Tomography) image set for the retrieval of 3-D structure of aurora by adjusting vergence angles of cameras to a common volume. National Institute of Polar Research, Japan is collaborating in observation and analysis. At the moment, the network has 6 stations separated from each other by about 50 km. Each station houses a monochromatic CCD (Charge Coupled Device) imaging system mounted on the steerable azimuth/elevation drive along with a house keeping unit and supervising computer linked to the control center via a telephone line. Altitude profiles of luminosity for stable arc and aurora vortex at 557.7nm and recently at 427.8nm are analysed by the algebraic reconstruction technique and compared with sophisticated numerical modelling of auroral emission rate. Conjunctions with satellites and radars are now intensively explored towards comprehensive understanding of the formation and electrodynamics of aurora. Imaging of polar stratospheric clouds is also attempted in relation to arctic environmental studies.

Offset-free black restoration and simplified correlated double sampling for linear CCDs

Black reference restoration and simple correlated double sampling techniques for a charge coupled device imaging system are discussed. A feedback loop which includes the entire video circuitry and analogue to digital conversion is used. The chief advantage of the technique is the elimination of offset errors.

Charge-coupled device imaging system for precision beam steering in laser communications

A CCD imaging system has been developed for precision laser beam pointing. The system contains a high frame rate CCD camera operated in windowing mode under command of an embedded processor. A laser pointing and tracking update rate of 2 kHz has been achieved.