Real-time Labeling Research Articles

Using Algorithms on Smart Transducer: An IEEE Standard Perspective

This paper describes the role of algorithm development in the IEEE standard 21451.001. The purpose is to promote and enhance the computational capabilities of smart transducers and facilitate the flow from sensor raw data processing to sensor knowledge fusion. A case study on real-time segmentation and labeling is conducted to demonstrate the feasibility of the first layer algorithm. The key role of the IEEE 21451.001 standard is to standardize the data structure for user-defined application code and pattern learning algorithms and regulate the protocols for data sharing and exchange. The considerations for the standard development are illustrated and discussed. The foreseeable impact of the standard for algorithms is highlighted as well.

Real time movement labelling of mobile event data

In this paper we introduce an advanced platform to label mobile event data with significant subscriber locations in real time. The presented platform is divided into two sections – the learning section and the real-time processing section. During the real-time processing step, we enrich live event streams with significant locations calculated in the learning step using stream and call detail record data. We validate our system by comparing a sample of subscribers' calculated locations with actual locations and give state benchmarks for minimum event counts. The validation confirms that the platform works within desired deviation levels from real locations. The accuracy strongly depends on the event count that we can take into account. Finally, we simulate a real-world scenario and measure the real-time labelling performance of our system. The results of this simulation confirm that our event labelling platform performs sufficiently well to process real event streams for millions of subscribers in real time.

MRI Visualization of CSF Flow

The ability to non-invasively establish patency of cerebral spinal fluid (CSF) flow between adjacent central nervous system (CNS) compartments is of importance in the diagnosis and treatment of patients with various areas of CSF flow obstruction. Recent advances in magnetic resonance imaging (MRI) technology allows for real-time magnetic labeling of CSF to directly visualize flow through different compartments.

Gambogic Acid Combined With CdTe QDs For Leukemia Cancer Cells Inhibition and Their Bio-Safety For Rat Brain

The development of new anticancer agents typically involves long research cycles, high cost and low efficiency and so on. There has been a growing public interest in the use of complementary and alternative medicine for cancer treatment and drug discovery. The emergence of nanotechnology and nanomaterials has greatly stimulated research in drug delivery and optimization. As a good fluorescence probe and simultaneously as potential drug carrier, Cadmium-Tellurium quantum dots (CdTe QDs) can optimize the use and a new potential therapy method of some drug-active compounds. Based on these considerations, we have explored the possibility to connect cysteamine (Cys) modified CdTe QDs (Cys-CdTe) to gambogic acid (GA). These functional QDs were studied for multi-functional drug delivery to investigate the synergetic effect of these composites between GA and QDs for human leukemia caner cells K562 and their drug-resistant cell line K562/A02 in vitro. Also the neurotoxicity of different drug systems for Sprague Dawley (SD) rat brain was analyzed by in vivo real-time microdialysis. The nanocomposites of GA loaded CdTe QDs were prepared and the drug releasing of GA-CdTe nanocomposites was performed at the medium of pH 6.0 and 7.4 in vitro. The cytotoxicity of GA and the nanocomposites for K562 and K562/A02 cells was studied by MTT, cell morphology, and flow cytometry. The labeling and imaging of CdTe QDs and the nanocomposites for cancer cells was performed by laser confocal fluorescence microscopy. Different drug systems were respectively injected into SD rat by vena caudalis and cerebrospinal fluid was collected by microdialysis in vivo. The neurotoxicity of different drug systems for rat brain was investigated by the changes of amino acid content in the cerebrospinal fluid. The Cys modified CdTe QDs showed the good fluorescence characteristics, with the wide excitation spectrum and narrow emission spectrum. The CdTe QDs could label in human leukemia cancer cells not only for K562 but also K562/A02 , which had the potential application in the cell imaging. Following the electrostatic attraction, surface adsorption et al, these CdTe QDs had the good capability of drug delivery with the high drug-loading and envelopment capacity. Also these GA-CdTe nanocomposites showed the pH sensitivity for the drug release and improved the drug's un-dissolvability. The GA-CdTe nanocomposites also had the fluorescence characteristics for labeling the different kinds of leukemia cancer cells. Gambogic acid play an important role in cancer therapy and also showed the sensitivity for the multidrug resistance (MDR) of K562/A02 cells in this study. The results indicated that GA-CdTe nanocomposites could significantly enhance the drug accumulation to improve the cytotoxicity and considerably overcome the multidrug resistance. GA-CdTe nanocomposites induced the arrest of G0/G1 phase to promote the cells apoptosis. The combination between GA and CdTe QDs can optimize the new potential therapeutic method for GA with the real time labeling and tracing during the disease therapy. The neurotoxicity of GA and GA-CdTe nanocomposites for rat brain was firstly explored by using the microdialysis in vivo. The results indicated that GA had the serious neurotoxicity through the concentration changes of amino acids for the rat brain. The concentration of some excitatory amino acids was remarkably increased. For the GA-CdTe nanocomposites based on CdTe QDs, the side effect of GA was visibly cut down, and the time to cause the neurotoxicity was apparently shortened. These nanocomposites own the better biocompatibility and bio-safety for the relevant cancer treatment in vitro and in vivo. This raises the promising possibility of the application of these fluorescent nanocomposites based on CdTe QDs for the target cancer therapy. The combination of GA with Cys-CdTe QDs can optimize the use and new potential multi-mode therapy of cancers. This may shed new light to exploit the potential application of the active compounds from Traditional Chinese Medicine (TCM). Disclosures:No relevant conflicts of interest to declare.

Ion sensing (EIS) real-time quantitative monitorization of isothermal DNA amplification

Field-effect-based devices are becoming a basic structural element in a new generation of microbiosensors. Reliable molecular characterization of DNA and/or RNA is of paramount importance for disease diagnostics and to follow up alterations in gene expression profiles. The use of such devices for point-of-need diagnostics has been hindered by the need of standard or real-time PCR amplification procedures. The present work focuses on the development of a tantalum pentoxide (Ta2O5) based sensor for the real-time label free detection of DNA amplification via loop mediated isothermal amplification (LAMP) allowing for quantitative analysis of the cMYC proto-oncogene. The strategy based on the field effect sensor was tested within a range of 1×108–1011 copies of target DNA, and a linear relationship between the log copy number of the initial template DNA and threshold time was observed allowing for a semi-quantitative analysis of DNA template. The concept offers many of the advantages of isothermal quantitative real-time DNA amplification in a label free approach and may pave the way to point-of-care quantitative molecular analysis focused on ease of use and low cost.

Synergetic effect of functional cadmium–tellurium quantum dots conjugated with gambogic acid for HepG2 cell-labeling and proliferation inhibition

We prepared and studied novel fluorescent nanocomposites based on gambogic acid (GA) and cadmium–tellurium (CdTe) quantum dots (CdTe QDs) modified with cysteamine for purpose of cancer cell labeling and combined treatment. The nanocomposites were denoted as GA-CdTe. Characterization results indicated that the CdTe QDs can readily bind onto cell plasma membranes and then be internalized into cancer cells for real-time labeling and tracing of human liver hepatocellular carcinoma cell line (HepG2) cells. GA-CdTe significantly enhanced drug accumulation in HepG2 cells and inhibited cancer cell proliferation. GA-CdTe nanocomposites also improved the drug action of GA molecules in HepG2 cells and induced the G2/M phase arrest of the cancer cell cycle, promoting cell apoptosis. Given the sensitive, pH-triggered release of GA-CdTe, the side effects of GA anticancer agents on normal cells/tissues in the blood circulation markedly decreased. Efficient drug release and accumulation in target tumor cells were also facilitated. Thus, the fluorescent GA-CdTe offered a new strategy for potential multimode cancer therapy and provided new channels for research into naturally-active compounds extracted from traditional Chinese medicinal plants.

Real‐time cell viability assays using a new anthracycline derivative DRAQ7®

The exclusion of charged fluorescent dyes by intact cells has become a well-established assay for determining viability of cells. In search for a noninvasive fluorescent probe capable of long-term monitoring of cell death in real-time, we evaluated a new anthracycline derivative DRAQ7. The novel probe does not penetrate the plasma membrane of living cells but when the membrane integrity is compromised, it enters and binds readily to nuclear DNA to report cell death. It proved to be nontoxic to a panel of cancer cell lines grown continuously for up to 72 h and did not induce any detectable DNA damage signaling when analyzed using laser scanning microscopy and flow cytometry. The DRAQ7 provided a sensitive, real-time readout of cell death induced by a variety of stressors such as hypoxia, starvation, and drug-induced cytotoxicity. The overall responses to anticancer agents and resulting pharmacological dose-response profiles were not affected by the growth of tumor cells in the presence DRAQ7. Moreover, we for the first time introduced a near real-time microflow cytometric assay based on combination of DRAQ7 and mitochondrial inner membrane potential (ΔΨ(m) ) sensitive probe TMRM. We provide evidence that this low-dosage, real-time labeling procedure provides multiparameter and kinetic fingerprint of anticancer drug action.

Dynamic Analysis of Drug-Induced Cytotoxicity Using Chip-Based Dielectrophoretic Cell Immobilization Technology

Quantification of programmed and accidental cell death provides useful end-points for the anticancer drug efficacy assessment. Cell death is, however, a stochastic process. Therefore, the opportunity to dynamically quantify individual cellular states is advantageous over the commonly employed static, end-point assays. In this work, we describe the development and application of a microfabricated, dielectrophoretic (DEP) cell immobilization platform for the real-time analysis of cancer drug-induced cytotoxicity. Microelectrode arrays were designed to generate weak electro-thermal vortices that support efficient drug mixing and rapid cell immobilization at the delta-shape regions of strong electric field formed between the opposite microelectrodes. We applied this technology to the dynamic analysis of hematopoietic tumor cells that represent a particular challenge for real-time imaging due to their dislodgement during image acquisition. The present study was designed to provide a comprehensive mechanistic rationale for accelerated cell-based assays on DEP chips using real-time labeling with cell permeability markers. In this context, we provide data on the complex behavior of viable vs dying cells in the DEP fields and probe the effects of DEP fields upon cell responses to anticancer drugs and overall bioassay performance. Results indicate that simple DEP cell immobilization technology can be readily applied for the dynamic analysis of investigational drugs in hematopoietic cancer cells. This ability is of particular importance in studying the outcome of patient derived cancer cells, when exposed to therapeutic drugs, as these cells are often rare and difficult to collect, purify and immobilize.

Real-Time Label Free Analysis of Native Bacterial Cells Binding to Carbohydrate Microarray

We present a novel experimental system based on glycoconjugates microarray and oblique-incidence reflectivity difference (OI-RD) microscopy to analyze binding of the onco-pathogen Helicobacter pylori to surface immobilized carbohydrates. The results confirm the recognized high-affinity binding of H. pylori BabA to the fucosylated Leb antigen, and suggest the presence of additional Leb binding adhesins not previously detected. OI-RD microscopy is a recently developed method for analyses of label-free biomolecules binding to immobilized targets by measuring small changes in phase and amplitude of a reflected optical wave from a solid surface due to the reaction of a solution-phase probe with the target. Besides applications of OI-RD in DNA hybridization, antigen-antibody interactions and small molecule libraries screening, we apply OI-RD microscopy to the binding analyses of whole bacterial cells to their host receptors presented on surfaces. Glycoconjugates microarrays consisting of Lea-HSA, Leb-HSA, Lex-HSA, and Ley-HSA were probed with H. pylori, H. pylori BabA deletion mutant and recombinant BabA protein. H. pylori demonstrated specific binding to Leb antigen, but did not bind to the closely related Lea, Lex or Ley antigens. The specificity in binding to the Leb antigen and not to Lea, Lex or Ley antigens was faithfully reproduced by recombinant BabA protein. Besides, H. pylori BabA deletion mutant does not express BabA and does not bind to Leb-HSA in solution, also demonstrated specific albeit reduced binding to Leb antigen. These results suggest that some H. pylori strains exhibit a complementary Leb-binding activity (independent of BabA) that has not been previously detected. The OI-RD technology with real-time analyses of whole cell bacterial binding to multivalently presented receptors in solid phase combined with magnified sensitivity for weaker adherence properties, offers new possibilities for identification of discrete attachment mechanisms essential for the infectious.

Sox10- Venus mice: a new tool for real-time labeling of neural crest lineage cells and oligodendrocytes

BackgroundWhile several mouse strains have recently been developed for tracing neural crest or oligodendrocyte lineages, each strain has inherent limitations. The connection between human SOX10 mutations and neural crest cell pathogenesis led us to focus on the Sox10 gene, which is critical for neural crest development. We generated Sox10-Venus BAC transgenic mice to monitor Sox10 expression in both normal development and in pathological processes.ResultsTissue fluorescence distinguished neural crest progeny cells and oligodendrocytes in the Sox10-Venus mouse embryo. Immunohistochemical analysis confirmed that Venus expression was restricted to cells expressing endogenous Sox10. Time-lapse imaging of various tissues in Sox10-Venus mice demonstrated that Venus expression could be visualized at the single-cell level in vivo due to the intense, focused Venus fluorescence. In the adult Sox10-Venus mouse, several types of mature and immature oligodendrocytes along with Schwann cells were clearly labeled with Venus, both before and after spinal cord injury.ConclusionsIn the newly-developed Sox10-Venus transgenic mouse, Venus fluorescence faithfully mirrors endogenous Sox10 expression and allows for in vivo imaging of live cells at the single-cell level. This Sox10-Venus mouse will thus be a useful tool for studying neural crest cells or oligodendrocytes, both in development and in pathological processes.

Proteomic Signatures Uncover Hydrogen Peroxide and Nitric Oxide Cross-Talk Signaling Network in Citrus Plants

Hydrogen peroxide (H(2)O(2)) and nitric oxide ((•)NO) elicit numerous processes in plants. However, our knowledge of H(2)O(2) and (•)NO-responsive proteins is limited. The present study aimed to identify proteins whose accumulation levels were regulated by these signaling molecules in citrus leaves. To address this question, hydroponically grown citrus plants were treated by incubating their roots in the presence of H(2)O(2) or the (•)NO donor, sodium nitroprusside (SNP). Both treatments induced H(2)O(2) and (•)NO production in leaves, indicating occurrence of oxidative and nitrosative stress conditions. However, treated plants maintained their normal physiological status. The vascular system was shown to be involved in the H(2)O(2) and (•)NO systemic signaling as evidenced by real-time labeling of the two molecules. Comparative proteomic analysis identified a number of proteins whose accumulation levels were altered by treatments. They were mainly involved in photosynthesis, defense and energy. More than half of them were commonly modulated by both treatments, indicating a strong overlap between H(2)O(2) and (•)NO responses. Using a redox proteomic approach, several proteins were also identified as being carbonylation targets of H(2)O(2) and SNP. The analysis reveals an interlinked H(2)O(2) and (•)NO proteins network allowing a deeper understanding of oxidative and nitrosative signaling in plants.

UPAINTing the plasma membrane

Using real-time labeling with fluorescent ligands, researchers track thousands of endogenous proteins at the surface of living cells.

Abstract 1497: Aldehyde dehydrogenase 1A1 expression is altered in ovarian cancer and is not associated with stem cells

Abstract Ovarian cancer (OvCa) is the leading cause of death from gynecologic malignancies. Tumors with serous histology are the most common and have the poorest prognosis among ovarian cancers. We identified Aldehyde dehydrogenase 1A1 (ALDH1) as an autoantigen in infertility patients, where infertility is an epidemiologic risk factor for OvCa. Aldehyde dehydrogenases belong to a family of detoxifying enzymes that may be related to the aggressiveness of various cancers. In breast and liver cancer, the cytosolic isoform, ALDH1, appears to be associated with cancer stem cells (CSC) and its high expression in breast tumors correlates with poor prognosis. Furthermore, ALDH1 expression is implicated in increased cell proliferation (aggressive behavior) in a lung cancer cell line. In contrast, studies show that ALDH1 is significantly higher in early stages of OvCa and high ALDH1expression is correlated with an increased OvCa patient survival. We hypothesized that ALDH1 expression differs in normal ovary and ovarian tumors and, unlike breast cancer, is not associated with CSCs and an aggressive OvCa phenotype. Methods: We compared ALDH1 with CD133 & CD44 (established CSC markers) mRNA and protein expression using quantitative Real-Time PCR (qPCR) and double label immunohistochemistry (IHC) in human normal ovary and tumors (benign and malignant) with serous histology. The percent of ALDH1 stained cells in at least 3 sections/ ovary were counted using unbiased StereoInvestigator cell counting software. The co-expression of ALDH1 and CD133/CD44 was assessed in malignant tumors by double labeled IHC and flow cytometry. Results: ALDH1 mRNA expression was significantly reduced (p<0.05) in malignant ovarian tumors compared to normal ovaries and benign ovarian tumors. ALDH1 was predominantly expressed in stromal fibroblasts along with surface epithelium in normal ovary. In malignant OvCa, ALDH1 was associated with tumor cells while in benign tumors it was associated with both tumor and normal ovarian stroma. Cells expressing ALDH1 were more numerous in normal ovaries (26.53 ± 2.69%; n=5; p<0.01) and serous benign tumors (22.89 ± 1.78%; n=3; p<0.05) compared to serous carcinomas (13.36 ± 6.23 %; n=5). ALDH1 was rarely co-expressed in CSCs. Conclusions: We compare for the first time, ALDH1 expression and differences in localization in normal and tumor ovaries and lack of ALDH1 co-expression in putative ovarian CSCs. Interestingly, ALDH1 mRNA and protein expression in benign tumors was intermediate between normal ovary and malignant tumors. ALDH1 expression was higher in non-CSCs and was significantly lower in malignant tumors suggesting OvCa differs from breast and liver cancer with regard to ALDH1 expression. Significance: Thus expression of ALDH1 may be advantageous in OvCa and loss of ALDH1 expression might be involved in malignant transformation. Support: NIH R01AI055060, Ovarian Cancer Survivors Network. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1497.

Real-Time Labeling of MPEG-2 Compressed Video

Digital mass recording devices for video data will enter the consumer market soon. Service providers are reluctant to offer services in digital form because of their fears for unrestricted duplication and dissemination. Therefore adequate copy protection systems should be developed, most likely based on labeling techniques. In this paper we propose two different techniques for the real-time labeling of digital video. Both methods embed the label information directly into an MPEG compressed video bitstream. The first method embeds the label by changing variable length codes in the bitstream. The second method discards some of the high frequency DCT-coefficients of the bitstream to embed the label. The first technique is computationally less expensive than the second one, however, the second one is more robust against attacks to remove the label. We show that the label can still be extracted after MPEG re-encoding at a lower bit-rate.

Hidden Markov mesh random field models in image analysis

This paper addresses the image modeling problem under the assumption that images can be represented by third-order, hidden Markov mesh random field models. The range of applications of the techniques described hereafter comprises the restoration of binary images, the modeling and compression of image data, as well as the segmentation of gray-level or multi-spectral images, and image sequences under the short-range motion hypothesis. We outline coherent approaches to both the problems of image modeling (pixel labeling) and estimation of model parameters (learning). We derive a real-time labeling algorithm-based on a maximum, marginal a posteriori probability criterion-for a hidden third-order Markov mesh random field model. Our algorithm achieves minimum time and space complexities simultaneously, and we describe what we believe to be the most appropriate data structures to implement it. Critical aspects of the computer simulation of a real-time implementation are discussed, down to the computer code level...