Science.gov

Sample records for assaying pet targets

  1. CPTAC Assay Portal: a repository of targeted proteomic assays

    SciTech Connect

    Whiteaker, Jeffrey R.; Halusa, Goran; Hoofnagle, Andrew N.; Sharma, Vagisha; MacLean, Brendan; Yan, Ping; Wrobel, John; Kennedy, Jacob; Mani, DR; Zimmerman, Lisa J.; Meyer, Matthew R.; Mesri, Mehdi; Rodriguez, Henry; Abbateillo, Susan E.; Boja, Emily; Carr, Steven A.; Chan, Daniel W.; Chen, Xian; Chen, Jing; Davies, Sherri; Ellis, Matthew; Fenyo, David; Hiltket, Tara; Ketchum, Karen; Kinsinger, Christopher; Kuhn, Eric; Liebler, Daniel; Lin, De; Liu, Tao; Loss, Michael; MacCoss, Michael; Qian, Weijun; Rivers, Robert; Rodland, Karin D.; Ruggles, Kelly; Scott, Mitchell; Smith, Richard D.; Thomas, Stefani N.; Townsend, Reid; Whiteley, Gordon; Wu, Chaochao; Zhang, Hui; Zhang, Zhen; Paulovich, Amanda G.

    2014-06-27

    To address these issues, the Clinical Proteomic Tumor Analysis Consortium (CPTAC) of the National Cancer Institute (NCI) has launched an Assay Portal (http://assays.cancer.gov) to serve as a public repository of well-characterized quantitative, MS-based, targeted proteomic assays. The purpose of the CPTAC Assay Portal is to facilitate widespread adoption of targeted MS assays by disseminating SOPs, reagents, and assay characterization data for highly characterized assays. A primary aim of the NCI-supported portal is to bring together clinicians or biologists and analytical chemists to answer hypothesis-driven questions using targeted, MS-based assays. Assay content is easily accessed through queries and filters, enabling investigators to find assays to proteins relevant to their areas of interest. Detailed characterization data are available for each assay, enabling researchers to evaluate assay performance prior to launching the assay in their own laboratory.

  2. Predictive Assay For Cancer Targets

    SciTech Connect

    Suess, A; Nguyen, C; Sorensen, K; Montgomery, J; Souza, B; Kulp, K; Dugan, L; Christian, A

    2005-09-19

    Early detection of cancer is a key element in successful treatment of the disease. Understanding the particular type of cancer involved, its origins and probable course, is also important. PhIP (2-amino-1-methyl-6 phenylimidazo [4,5-b]pyridine), a heterocyclic amine produced during the cooking of meat at elevated temperatures, has been shown to induce mammary cancer in female, Sprague-Dawley rats. Tumors induced by PhIP have been shown to contain discreet cytogenetic signature patterns of gains and losses using comparative genomic hybridization (CGH). To determine if a protein signature exists for these tumors, we are analyzing expression levels of the protein products of the above-mentioned tumors in combination with a new bulk protein subtractive assay. This assay produces a panel of antibodies against proteins that are either on or off in the tumor. Hybridization of the antibody panel onto a 2-D gel of tumor or control protein will allow for identification of a distinct protein signature in the tumor. Analysis of several gene databases has identified a number of rat homologs of human cancer genes located in these regions of gain and loss. These genes include the oncogenes c-MYK, ERBB2/NEU, THRA and tumor suppressor genes EGR1 and HDAC3. The listed genes have been shown to be estrogen-responsive, suggesting a possible link between delivery of bio-activated PhIP to the cell nucleus via estrogen receptors and gene-specific PhIP-induced DNA damage, leading to cell transformation. All three tumors showed similar silver staining patterns compared to each other, while they all were different than the control tissue. Subsequent screening of these genes against those from tumors know to be caused by other agents may produce a protein signature unique to PhIP, which can be used as a diagnostic to augment optical and radiation-based detection schemes.

  3. Electroplated targets for production of unique PET radionuclides

    NASA Astrophysics Data System (ADS)

    Bui, V.; Sheh, Y.; Finn, R.; Francesconi, L.; Cai, S.; Schlyer, D.; Wieland, B.

    1995-12-01

    The past decade has witnessed the applications of positron emission tomography (PET) evolving from a purely research endeavor to a procedure which has specific clinical applications in the areas of cardiology, neurology and oncology. The growth of PET has been facilitated by developments in both medical instrumentation and radiopharmaceutical chemistry efforts. Included in this latter effort has been the low energy accelerator production and processing of unique PET radionuclides appropriate for the radiolabeling of biomolecules, i.e. monoclonal antibodies and peptides. The development and application of electroplated targets of antimony and copper for the production of iodine-124 and gallium-66 respectively, utilizing the Memorial Sloan-Kettering Cancer Center (MSKCC) cyclotron are examples of target design and development applicable to many medical accelerators.

  4. Electroplating targets for production of unique PET radionuclides

    SciTech Connect

    Bui, V.; Sheh, Y.; Finn, R.

    1994-12-31

    The past decade has witnessed the applications of Positron Emission Tomography (PET) evolving from a purely research endeavour to a procedure which has specific clinical applications in the areas of cardiology, neurology and oncology. The growth of PET has been facilitated by developments in medical instrumentation and radiopharmaceutical chemistry efforts. Included in this latter effort has been the low energy accelerator production and processing of unique PET radionuclides appropriate for the radiolabeling of biomolecules i.e. monoclonal antibodies and pepetides. The development and application of electroplated targets of antimony and copper for the production of iodine-124 and gallium-66 respectively, utilizing the Memorial Sloan-Kettering Cancer Center cyclotron are examples of target design and development applicable to many medical accelerators.

  5. Thresholding in PET images of static and moving targets

    NASA Astrophysics Data System (ADS)

    Yaremko, Brian; Riauka, Terence; Robinson, Don; Murray, Brad; Alexander, Abraham; McEwan, Alexander; Roa, Wilson

    2005-12-01

    Continued therapeutic gain in the treatment of non-small-cell lung cancer (NSCLC) will depend upon our ability to escalate the dose to the primary tumour while minimizing normal tissue toxicity. Both these objectives are facilitated by the accurate definition of a target volume that is as small as possible. To this end, both tumour immobilizations via deep inspiratory breath-hold, along with positron emission tomography (PET), have emerged as two promising approaches. Though PET is an excellent means of defining the general location of a tumour focus, its ability to define exactly the geometric extent of such a focus strongly depends upon selection of an appropriate image threshold. However, in clinical practice, the image threshold is typically not chosen according to consistent, well-established criteria. This study explores the relationship between image threshold and the resultant PET-defined volume using a series of F-18 radiotracer-filled hollow spheres of known internal volumes, both static and under oscillatory motion. The effects of both image threshold and tumour motion on the resultant PET image are examined. Imaging data are further collected from a series of simulated gated PET acquisitions in order to test the feasibility of a patient-controlled gating mechanism during deep inspiratory breath-hold. This study illustrates quantitatively considerable variability in resultant PET-defined tumour volumes depending upon numerous factors, including image threshold, size of the lesion, the presence of tumour motion and the scanning protocol. In this regard, when using PET in treatment planning for NSCLC, the radiation oncologist must select the image threshold very carefully to avoid either under-dosing the tumour or overdosing normal tissues.

  6. Microfluidic integration for automated targeted proteomic assays.

    PubMed

    Hughes, Alex J; Lin, Robert K C; Peehl, Donna M; Herr, Amy E

    2012-04-17

    A dearth of protein isoform-based clinical diagnostics currently hinders advances in personalized medicine. A well-organized protein biomarker validation process that includes facile measurement of protein isoforms would accelerate development of effective protein-based diagnostics. Toward scalable protein isoform analysis, we introduce a microfluidic "single-channel, multistage" immunoblotting strategy. The multistep assay performs all immunoblotting steps: separation, immobilization of resolved proteins, antibody probing of immobilized proteins, and all interim wash steps. Programmable, low-dispersion electrophoretic transport obviates the need for pumps and valves. A three-dimensional bulk photoreactive hydrogel eliminates manual blotting. In addition to simplified operation and interfacing, directed electrophoretic transport through our 3D nanoporous reactive hydrogel yields superior performance over the state-of-the-art in enhanced capture efficiency (on par with membrane electroblotting) and sparing consumption of reagents (ca. 1 ng antibody), as supported by empirical and by scaling analyses. We apply our fully integrated microfluidic assay to protein measurements of endogenous prostate specific antigen isoforms in (i) minimally processed human prostate cancer cell lysate (1.1 pg limit of detection) and (ii) crude sera from metastatic prostate cancer patients. The single-instrument functionality establishes a scalable microfluidic framework for high-throughput targeted proteomics, as is relevant to personalized medicine through robust protein biomarker verification, systematic characterization of new antibody probes for functional proteomics, and, more broadly, to characterization of human biospecimen repositories. PMID:22474344

  7. Some target assay uncertainties for passive neutron coincidence counting

    SciTech Connect

    Ensslin, N.; Langner, D.G.; Menlove, H.O.; Miller, M.C.; Russo, P.A.

    1990-01-01

    This paper provides some target assay uncertainties for passive neutron coincidence counting of plutonium metal, oxide, mixed oxide, and scrap and waste. The target values are based in part on past user experience and in part on the estimated results from new coincidence counting techniques that are under development. The paper summarizes assay error sources and the new coincidence techniques, and recommends the technique that is likely to yield the lowest assay uncertainty for a given material type. These target assay uncertainties are intended to be useful for NDA instrument selection and assay variance propagation studies for both new and existing facilities. 14 refs., 3 tabs.

  8. A solid target system with remote handling of irradiated targets for PET cyclotrons.

    PubMed

    Siikanen, J; Tran, T A; Olsson, T G; Strand, S-E; Sandell, A

    2014-12-01

    A solid target system was developed for a PET cyclotron. The system is compatible with many different target materials in the form of foils and electroplated/sputtered targets which makes it useful for production of a wide variety of different PET radionuclides. The target material is manually loaded into the system. Remote handling of irradiated target material is managed with a pneumatic piston and a vacuum technique which allows the targets to be dropped into a shielded transport container. To test the target performance, proton irradiations (12.8 MeV, 45 μA) of monoisotopic yttrium foils (0.64 mm, direct water cooling) were performed to produce 89Zr. The yields were 2200±200 MBq (1 h, n=13) and 6300±65 MBq (3 h, n=3). PMID:25265518

  9. Multimodality PET/MRI agents targeted to activated macrophages.

    PubMed

    Tu, Chuqiao; Ng, Thomas S C; Jacobs, Russell E; Louie, Angelique Y

    2014-02-01

    The recent emergence of multimodality imaging, particularly the combination of PET and MRI, has led to excitement over the prospect of improving detection of disease. Iron oxide nanoparticles have become a popular platform for the fabrication of PET/MRI probes owing to their advantages of high MRI detection sensitivity, biocompatibility, and biodegradability. In this article, we report the synthesis of dextran-coated iron oxide nanoparticles (DIO) labeled with the positron emitter (64)Cu to generate a PET/MRI probe, and modified with maleic anhydride to increase the negative surface charge. The modified nanoparticulate PET/MRI probe (MDIO-(64)Cu-DOTA) bears repetitive anionic charges on the surface that facilitate recognition by scavenger receptor type A (SR-A), a ligand receptor found on activated macrophages but not on normal vessel walls. MDIO-(64)Cu-DOTA has an average iron oxide core size of 7-8 nm, an average hydrodynamic diameter of 62.7 nm, an r1 relaxivity of 16.8 mM(-1) s(-1), and an r 2 relaxivity of 83.9 mM(-1) s(-1) (37 °C, 1.4 T). Cell studies confirmed that the probe was nontoxic and was specifically taken up by macrophages via SR-A. In comparison with the nonmodified analog, the accumulation of MDIO in macrophages was substantially improved. These characteristics demonstrate the promise of MDIO-(64)Cu-DOTA for identification of vulnerable atherosclerotic plaques via the targeting of macrophages. PMID:24166283

  10. Fully Bayesian Analysis of High-throughput Targeted Metabolomics Assays

    EPA Science Inventory

    High-throughput metabolomic assays that allow simultaneous targeted screening of hundreds of metabolites have recently become available in kit form. Such assays provide a window into understanding changes to biochemical pathways due to chemical exposure or disease, and are usefu...

  11. Avian-specific real-time PCR assay for authenticity control in farm animal feeds and pet foods.

    PubMed

    Pegels, Nicolette; González, Isabel; García, Teresa; Martín, Rosario

    2014-01-01

    A highly sensitive TaqMan real-time PCR assay targeting the mitochondrial 12S rRNA gene was developed for detection of an avian-specific DNA fragment (68bp) in farm animal and pet feeds. The specificity of the assay was verified against a wide representation of animal and plant species. Applicability assessment of the avian real-time PCR was conducted through representative analysis of two types of compound feeds: industrial farm animal feeds (n=60) subjected to extreme temperatures, and commercial dog and cat feeds (n=210). Results obtained demonstrated the suitability of the real-time PCR assay to detect the presence of low percentages of highly processed avian material in the feed samples analysed. Although quantification results were well reproducible under the experimental conditions tested, an accurate estimation of the target content in feeds is impossible in practice. Nevertheless, the method may be useful as an alternative tool for traceability purposes within the framework of feed control. PMID:24001810

  12. PNA-FISH assays for early targeted bacteraemia treatment.

    PubMed

    Parcell, B J; Orange, G V

    2013-11-01

    PNA-FISH S. aureus/CNS and GNR Traffic Light assays were compared with standard culture methods for identifying bacteraemia in 156 blood cultures from 131 patients. Results correlated with final culture results in 153 cultures. Retrospective case note review revealed that earlier targeted treatment would have occurred in 10.7% of cases. PMID:24055387

  13. Pet-1 Switches Transcriptional Targets Postnatally to Regulate Maturation of Serotonin Neuron Excitability

    PubMed Central

    Wyler, Steven C.; Spencer, W. Clay; Green, Noah H.; Rood, Benjamin D.; Crawford, LaTasha; Craige, Caryne; Gresch, Paul; McMahon, Douglas G.; Beck, Sheryl G.

    2016-01-01

    Newborn neurons enter an extended maturation stage, during which they acquire excitability characteristics crucial for development of presynaptic and postsynaptic connectivity. In contrast to earlier specification programs, little is known about the regulatory mechanisms that control neuronal maturation. The Pet-1 ETS (E26 transformation-specific) factor is continuously expressed in serotonin (5-HT) neurons and initially acts in postmitotic precursors to control acquisition of 5-HT transmitter identity. Using a combination of RNA sequencing, electrophysiology, and conditional targeting approaches, we determined gene expression patterns in maturing flow-sorted 5-HT neurons and the temporal requirements for Pet-1 in shaping these patterns for functional maturation of mouse 5-HT neurons. We report a profound disruption of postmitotic expression trajectories in Pet-1−/− neurons, which prevented postnatal maturation of 5-HT neuron passive and active intrinsic membrane properties, G-protein signaling, and synaptic responses to glutamatergic, lysophosphatidic, and adrenergic agonists. Unexpectedly, conditional targeting revealed a postnatal stage-specific switch in Pet-1 targets from 5-HT synthesis genes to transmitter receptor genes required for afferent modulation of 5-HT neuron excitability. 5-HT1a autoreceptor expression depended transiently on Pet-1, thus revealing an early postnatal sensitive period for control of 5-HT excitability genes. Chromatin immunoprecipitation followed by sequencing revealed that Pet-1 regulates 5-HT neuron maturation through direct gene activation and repression. Moreover, Pet-1 directly regulates the 5-HT neuron maturation factor Engrailed 1, which suggests Pet-1 orchestrates maturation through secondary postmitotic regulatory factors. The early postnatal switch in Pet-1 targets uncovers a distinct neonatal stage-specific function for Pet-1, during which it promotes maturation of 5-HT neuron excitability. SIGNIFICANCE STATEMENT The

  14. Pet Food Palatability Evaluation: A Review of Standard Assay Techniques and Interpretation of Results with a Primary Focus on Limitations

    PubMed Central

    Aldrich, Gregory C.; Koppel, Kadri

    2015-01-01

    Simple Summary Palatability of pet foods is typically measured using a single-bowl or a two-bowl test. While these tests give a general understanding of the liking or preference of one food over another, opportunities exist for further method development. Abstract The pet food industry continues to grow steadily as a result of new innovative products. Quality control and product development tests for pet foods are typically conducted through palatability testing with dogs and cats. Palatability is the measure of intake of a food that indicates acceptance or the measure of preference of one food over another. Pet food palatability is most commonly measured using a single-bowl or a two-bowl assay. While these tests answer some questions about the animals’ perception of the food, there are many limitations as well. This review addresses some of these limitations and indicates opportunities for future research. PMID:26479136

  15. Lung cancer biomarkers, targeted therapies and clinical assays

    PubMed Central

    Ersek, Jennifer L.; Kim, Edward S.

    2015-01-01

    Until recently, the majority of genomic cancer research has been in discovery and validation; however, as our knowledge of tumor molecular profiling improves, the idea of genomic application in the clinic becomes increasingly tangible, paralleled with the drug development of newer targeted therapies. A number of profiling methodologies exist to identify biomarkers found within the patient (germ-line DNA) and tumor (somatic DNA). Subsequently, commercially available clinical assays to test for both germ-line and somatic alterations that are prognostic and/or predictive of disease outcome, toxicity or treatment response have significantly increased. This review aims to summarize clinically relevant cancer biomarkers that serve as targets for therapy and their potential relationship to lung cancer. In order to realize the full potential of genomic cancer medicine, it is imperative that clinicians understand these intricate molecular pathways, the therapeutic implication of mutations within these pathways, and the availability of clinical assays to identify such biomarkers. PMID:26629419

  16. Validation of a 4D-PET Maximum Intensity Projection for Delineation of an Internal Target Volume

    SciTech Connect

    Callahan, Jason; Kron, Tomas; Schneider-Kolsky, Michal; Dunn, Leon; Thompson, Mick; Siva, Shankar; Aarons, Yolanda; Binns, David; Hicks, Rodney J.

    2013-07-15

    Purpose: The delineation of internal target volumes (ITVs) in radiation therapy of lung tumors is currently performed by use of either free-breathing (FB) {sup 18}F-fluorodeoxyglucose-positron emission tomography-computed tomography (FDG-PET/CT) or 4-dimensional (4D)-CT maximum intensity projection (MIP). In this report we validate the use of 4D-PET-MIP for the delineation of target volumes in both a phantom and in patients. Methods and Materials: A phantom with 3 hollow spheres was prepared surrounded by air then water. The spheres and water background were filled with a mixture of {sup 18}F and radiographic contrast medium. A 4D-PET/CT scan was performed of the phantom while moving in 4 different breathing patterns using a programmable motion device. Nine patients with an FDG-avid lung tumor who underwent FB and 4D-PET/CT and >5 mm of tumor motion were included for analysis. The 3 spheres and patient lesions were contoured by 2 contouring methods (40% of maximum and PET edge) on the FB-PET, FB-CT, 4D-PET, 4D-PET-MIP, and 4D-CT-MIP. The concordance between the different contoured volumes was calculated using a Dice coefficient (DC). The difference in lung tumor volumes between FB-PET and 4D-PET volumes was also measured. Results: The average DC in the phantom using 40% and PET edge, respectively, was lowest for FB-PET/CT (DCAir = 0.72/0.67, DCBackground 0.63/0.62) and highest for 4D-PET/CT-MIP (DCAir = 0.84/0.83, DCBackground = 0.78/0.73). The average DC in the 9 patients using 40% and PET edge, respectively, was also lowest for FB-PET/CT (DC = 0.45/0.44) and highest for 4D-PET/CT-MIP (DC = 0.72/0.73). In the 9 lesions, the target volumes of the FB-PET using 40% and PET edge, respectively, were on average 40% and 45% smaller than the 4D-PET-MIP. Conclusion: A 4D-PET-MIP produces volumes with the highest concordance with 4D-CT-MIP across multiple breathing patterns and lesion sizes in both a phantom and among patients. Freebreathing PET/CT consistently

  17. Novel Bispecific PSMA/GRPr Targeting Radioligands with Optimized Pharmacokinetics for Improved PET Imaging of Prostate Cancer.

    PubMed

    Liolios, C; Schäfer, M; Haberkorn, U; Eder, M; Kopka, K

    2016-03-16

    A new series of bispecific radioligands (BRLs) targeting prostate-specific membrane antigen (PSMA) and gastrin releasing peptide receptor (GRPr), both expressed on prostate cancer cells, was developed. Their design was based on the bombesin (BN) analogue, H2N-PEG2-[d-Tyr(6),β-Ala(11),Thi(13),Nle(14)]BN(6-14), which binds to GRPr with high affinity and specificity, and the peptidomimetic urea-based pseudoirreversible inhibitor of PSMA, Glu-ureido-Lys. The two pharmacophores were coupled through copper(I)-catalyzed azide-alkyne cycloaddition to the bis(tetrafluorophenyl) ester of the chelating agent HBED-CC via amino acid linkers made of positively charged His (H) and negatively charged Glu (E): -(HE)n- (n = 0-3). The BRLs were labeled with (68)Ga, and their preliminary pharmacological properties were evaluated in vitro (competitive and time kinetic binding assays) on prostate cancer (PC-3, LNCaP) and rat pancreatic (AR42J) cell lines and in vivo by biodistribution and small animal PET imaging studies in both normal and tumor-bearing mice. The IC50/Ki values determined for all BRLs essentially matched those of the respective monomers. The maximal cellular uptake of the BLRs was observed between 20 and 30 min. The BRLs showed a synergistic ability in vivo by targeting both PSMA (LNCaP) and GRPr (PC-3) positive tumors, whereas the charged -(HE)n- (n = 1-3) linkers significantly reduced the kidney and spleen uptake. The bispecific (PSMA and GRPr) targeting ability and optimized pharmacokinetics of the compounds developed in this study could lead to their future application in clinical practice as more sensitive radiotracers for noninvasive imaging of prostate cancer (PCa) by PET/CT and PET/MRI. PMID:26726823

  18. Development of a Targeted Urine Proteome Assay for Kidney Diseases

    PubMed Central

    Cantley, Lloyd G.; Colangelo, Christopher M.; Stone, Kathryn L.; Chung, Lisa; Belcher, Justin; Abbott, Thomas; Cantley, Jennifer L.; Williams, Kenneth R.; Parikh, Chirag R.

    2016-01-01

    Human urine is the least invasive and most readily available bio fluid whose proteome has been shown to change in response to disease or drug treatment. Urine is thus very amenable to quantitative proteomics and is a logical sample choice for identifying protein biomarkers for kidney diseases. In this study potential biomarkers were identified initially by using a multi-proteomics workflow to compare urine proteomes of kidney transplant patients who exhibited immediate versus delayed graft function. To comprehensively interrogate the urine proteome two “bottom up”, mass spectrometric-based discovery approaches, iTRAQ and Label Free Quantitation (LFQ), were complemented by Differential Fluorescence Gel Electrophoresis (DIGE) analyses of intact urine proteins from kidney transplant recipients who received a deceased donor kidney. Differentially expressed proteins in the two patient groups were identified, and corresponding stable isotope–labeled internal peptide standard (SIS) peptides were synthesized for scheduled multiple reaction monitoring (MRM). The Targeted Urine Proteome Assay (TUPA) was then developed by identifying those peptides for which there were at least 2 transitions for which interference in a urine matrix across 156 MRM runs was less than 30%. This resulted in a final assay that monitors 224 peptides corresponding to 167 quantifiable proteins. PMID:26220717

  19. Bimodal Thrombus Imaging: Simultaneous PET/MR Imaging with a Fibrin-targeted Dual PET/MR Probe—Feasibility Study in Rat Model

    PubMed Central

    Uppal, Ritika; Catana, Ciprian; Ay, Ilknur; Benner, Thomas; Sorensen, A. Gregory

    2011-01-01

    Purpose: To image thrombus by using magnetic resonance (MR) imaging and positron emission tomography (PET) simultaneously in a rat arterial thrombus model with a dual PET/MR probe. Materials and Methods: Animal studies were approved by the institutional animal use committee. A dual PET/MR probe was synthesized by means of partial exchange of gadolinium for copper 64 (64Cu) in the fibrin-targeted MR probe EP-2104R. A preformed 25-mm thrombus was injected into the right internal carotid artery of a rat. Imaging was performed with a clinical 3.0-T MR imager with an MR-compatible human PET imager. Rats (n = 5) were imaged prior to and after systemic administration of the dual probe by using simultaneous PET/MR. The organ distribution of 64Cu and gadolinium was determined ex vivo (n = 8), 2 hours after injection by using well counting and inductively coupled plasma mass spectrometry, respectively. Signal intensity ratios (SIRs) between the thrombus-containing and contralateral vessel were computed from PET images and MR data before and after probe administration. Results: The dual probe was synthesized with greater than 98% radiochemical purity. Thrombus enhancement was observed in all five animals at both MR (SIR[postprobe]/SIR[preprobe] = 1.71 ± 0.35, P = .0053) and PET (SIR = 1.85 ± 0.48, P = .0087) after injection of the dual PET/MR probe. Ex vivo analysis at 2 hours after injection showed the highest 64Cu and gadolinium concentrations, after the excretory organs (kidney and liver), to be in the thrombus. Conclusion: A fibrin-targeted dual PET/MR probe enables simultaneous, direct MR and PET imaging of thrombus. © RSNA, 2010 PMID:21177389

  20. Ligand-binding assays for cyanobacterial neurotoxins targeting cholinergic receptors.

    PubMed

    Aráoz, Rómulo; Vilariño, Natalia; Botana, Luis M; Molgó, Jordi

    2010-07-01

    Toxic cyanobacterial blooms are a threat to public health because of the capacity of some cyanobacterial species to produce potent hepatotoxins and neurotoxins. Cyanobacterial neurotoxins are involved in the rapid death of wild and domestic animals by targeting voltage gated sodium channels and cholinergic synapses, including the neuromuscular junction. Anatoxin-a and its methylene homologue homoanatoxin-a are potent agonists of nicotinic acetylcholine receptors. Since the structural determination of anatoxin-a, several mass spectrometry-based methods have been developed for detection of anatoxin-a and, later, homoanatoxin-a. Mass spectrometry-based techniques provide accuracy, precision, selectivity, sensitivity, reproducibility, adequate limit of detection, and structural and quantitative information for analyses of cyanobacterial anatoxins from cultured and environmental cyanobacterial samples. However, these physicochemical techniques will only detect known toxins for which toxin standards are commercially available, and they require highly specialized laboratory personnel and expensive equipment. Receptor-based assays are functional methods that are based on the mechanism of action of a class of toxins and are thus, suitable tools for survey of freshwater reservoirs for cyanobacterial anatoxins. The competition between cyanobacterial anatoxins and a labelled ligand for binding to nicotinic acetylcholine receptors is measured radioactively or non-radioactively providing high-throughput screening formats for routine detection of this class of neurotoxins. The mouse bioassay is the method of choice for marine toxin monitoring, but has to be replaced by fully validated functional methods. In this paper we review the ligand-binding assays developed for detection of cyanobacterial and algal neurotoxins targeting the nicotinic acetylcholine receptors and for high-throughput screening of novel nicotinic agents. PMID:20238109

  1. From anatomical to biological target volumes: the role of PET in radiation treatment planning

    PubMed Central

    Schinagl, D A X; Kaanders, J H A M; Oyen, W J G

    2006-01-01

    Progress in radiation oncology requires a re-evaluation of the methods of target volume delineation beyond anatomical localization. New molecular imaging techniques for tumour visualisation such as positron emission tomography (PET) provide insight into tumour characteristics and can be complementary to the anatomical data of computed tomography or magnetic resonance imaging. In this review, three issues are discussed: First, can PET identify a tumour more accurately? Second, can biological tumour characteristics be visualised? Third, can intratumoural heterogeneity of these characteristics be identified? PMID:17114062

  2. Integrating respiratory-gated PET-based target volume delineation in liver SBRT planning, a pilot study

    PubMed Central

    2014-01-01

    Background To assess the feasibility and benefit of integrating four-dimensional (4D) Positron Emission Tomography (PET) – computed tomography (CT) for liver stereotactic body radiation therapy (SBRT) planning. Methods 8 patients with 14 metastases were accrued in the study. They all underwent a non-gated PET and a 4D PET centered on the liver. The same CT scan was used for attenuation correction, registration, and considered the planning CT for SBRT planning. Six PET phases were reconstructed for each 4D PET. By applying an individualized threshold to the 4D PET, a Biological Internal Target Volume (BITV) was generated for each lesion. A gated Planning Target Volume (PTVg) was created by adding 3 mm to account for set-up margins. This volume was compared to a manual Planning Target Volume (PTV) delineated with the help of a semi-automatic Biological Target Volume (BTV) obtained from the non-gated exam. A 5 mm radial and a 10 mm craniocaudal margins were applied to account for tumor motion and set-up margins to create the PTV. Results One undiagnosed liver metastasis was discovered thanks to the 4D PET. The semi-automatic BTV were significantly smaller than the BITV (p = 0.0031). However, after applying adapted margins, 4D PET allowed a statistically significant decrease in the PTVg as compared to the PTV (p = 0.0052). Conclusions In comparison to non-gated PET, 4D PET may better define the respiratory movements of liver targets and improve SBRT planning for liver metastases. Furthermore, non respiratory-gated PET exams can both misdiagnose liver metastases and underestimate the real internal target volumes. PMID:24885897

  3. Targeting Angiogenesis Using a C-Type Atrial Natriuretic Factor–Conjugated Nanoprobe and PET

    PubMed Central

    Liu, Yongjian; Pressly, Eric D.; Abendschein, Dana R.; Hawker, Craig J.; Woodard, Geoffrey E.; Woodard, Pamela K.; Welch, Michael J.

    2014-01-01

    Sensitive, specific, and noninvasive detection of angiogenesis would be helpful in discovering new strategies for the treatment of cardiovascular diseases. Recently, we reported the 64Cu-labeled C-type atrial natriuretic factor (CANF) fragment for detecting the upregulation of natriuretic peptide clearance receptor (NPR-C) with PET on atherosclerosis-like lesions in an animal model. However, it is unknown whether NPR-C is present and overexpressed during angiogenesis. The goal of this study was to develop a novel CANF-integrated nanoprobe to prove the presence of NPR-C and offer sensitive detection with PET during development of angiogenesis in mouse hind limb. Methods We prepared a multifunctional, core-shell nanoparticle consisting of DOTA chelators attached to a poly(methyl methacrylate) core and CANF-targeting moieties attached to poly(ethylene glycol) chain ends in the shell of the nanoparticle. Labeling of this nanoparticle with 64Cu yielded a high-specific-activity nanoprobe for PET imaging NPR-C receptor in a mouse model of hind limb ischemia–induced angiogenesis. Histology and immunohistochemistry were performed to assess angiogenesis development and NPR-C localization. Results 15O-H2O imaging showed blood flow restoration in the previously ischemic hind limb, consistent with the development of angiogenesis. The targeted DOTA-CANF-comb nanoprobe showed optimized pharmacokinetics and biodistribution. PET imaging demonstrated significantly higher tracer accumulation for the targeted DOTA-CANF-comb nanoprobe than for either the CANF peptide tracer or the nontargeted control nanoprobe (P < 0.05, both). Immunohistochemistry confirmed NPR-C upregulation in the angiogenic lesion with colocalization in both endothelial and smooth muscle cells. PET and immunohistochemistry competitive receptor blocking verified the specificity of the targeted nanoprobe to NPR-C receptor. Conclusion As evidence of its translational potential, this customized DOTA

  4. Impact of Manual and Automated Interpretation of Fused PET/CT Data on Esophageal Target Definitions in Radiation Planning

    SciTech Connect

    Hong, Theodore S. Killoran, Joseph H.; Mamede, Marcelo; Mamon, Harvey J.

    2008-12-01

    Purpose: We compare CT-only based esophageal tumor definition with two PET/CT based methods: (1) manual contouring and (2) a semiautomated method based on specific thresholds. Methods and Materials: Patients with esophageal cancer treated at Brigham and Women's Hospital from 2003 to 2006 were identified. CT-based tumor volumes were compared with manual PET/CT-based volumes and semiautomated PET-based tumor volumes. Differences were scored as (1) minor if the superior or inferior extent of the primary tumor (or both) differed by 1-2 cm and (2) major if the difference was > 2 cm or if different noncontiguous nodal regions were identified as being grossly involved. Results: Comparing CT-based gross tumor volumes (GTVs) to manually defined PET/CT-based GTVs, use of PET changed volumes for 21 of 25 (84%) patients: 12 patients (48%) exhibited minor differences, whereas for 9 patients (36%), the differences were major. For 4 (16%) patients, the major difference was due to discrepancy in celiac or distant mediastinal lymph node involvement. Use of automated PET volumes changed the manual PET length in 14 patients (56%): 8 minor and 6 major. Conclusions: The use of PET/CT in treatment planning for esophageal cancer can affect target definition. Two PET-based techniques can also produce significantly different tumor volumes in a large percentage of patients. Further investigations to clarify the optimal use of PET/CT data in treatment planning are warranted.

  5. PET radioligands targeting the brain GABAA /benzodiazepine receptor complex.

    PubMed

    Andersson, Jan D; Halldin, Christer

    2013-01-01

    The development of positron emission tomography radioligands for the GABAA /benzodiazepine receptor complex (GABAA receptor) labeled with (11) C and (18) F is examined. The review covers labeling strategies as well as brief biological evaluations of radioligands. In addition, we assess the special considerations that must be taken during a development program for radioligands targeting the GABAA receptor and explore some of the challenges that lie ahead. PMID:24285326

  6. Preclinical Study on GRPR-Targeted (68)Ga-Probes for PET Imaging of Prostate Cancer.

    PubMed

    Sun, Yao; Ma, Xiaowei; Zhang, Zhe; Sun, Ziyan; Loft, Mathias; Ding, Bingbing; Liu, Changhao; Xu, Liying; Yang, Meng; Jiang, Yuxin; Liu, Jianfeng; Xiao, Yuling; Cheng, Zhen; Hong, Xuechuan

    2016-08-17

    Gastrin-releasing peptide receptor (GRPR) targeted positron emission tomography (PET) is a highly promising approach for imaging of prostate cancer (PCa) in small animal models and patients. Developing a GRPR-targeted PET probe with excellent in vivo performance such as high tumor uptake, high contrast, and optimal pharmacokinetics is still very challenging. Herein, a novel bombesin (BBN) analogue (named SCH1) based on JMV594 peptide modified with an 8-amino octanoic acid spacer (AOC) was thus designed and conjugated with the metal chelator 1,4,7-triazacyclononane,1-glutaric acid-4,7-acetic acid (NODAGA). The resulting NODAGA-SCH1 was then radiolabeled with (68)Ga and evaluated for PET imaging of PCa. Compared with (68)Ga-NODAGA-JMV594 probe, (68)Ga-NODAGA-SCH1 exhibited excellent PET/CT imaging properties on PC-3 tumor-bearing nude mice, such as high tumor uptake (5.80 ± 0.42 vs 3.78 ± 0.28%ID/g, 2 h) and high tumor/muscle contrast (16.6 ± 1.50 vs 8.42 ± 0.61%ID/g, 2 h). Importantly, biodistribution data indicated a relatively similar accumulation of (68)Ga-NODAGA-SCH1 was observed in the liver (4.21 ± 0.42%ID/g) and kidney (3.41 ± 0.46%ID/g) suggesting that the clearance is through both the kidney and the liver. Overall, (68)Ga-NODAGA-SCH1 showed promising in vivo properties and is a promising candidate for translation into clinical PET-imaging of PCa patients. PMID:27399868

  7. Full in-beam PET measurements of 62 MeV protons onto a PMMA target

    NASA Astrophysics Data System (ADS)

    Sportelli, G.; Straub, K.; Aiello, M.; Attanasi, F.; Belcari, N.; Camarlinghi, N.; Cirrone, G. A. P.; Cuttone, G.; Ferretti, S.; Marino, N.; Nicolosi, D.; Romano, F.; Rosso, V.; Del Guerra, A.

    2013-08-01

    Positron emission tomography (PET) is a valuable technique to monitor in situ and non-invasively the particle range in ion beam therapy exploiting the beta+ activity produced in nuclear interactions along the beam path within the target volume. Due to the high random rates and dead-time losses induced by the particle spills, as of to date data are usually acquired during beam pauses or after the irradiation. We have developed a new PET prototype with a faster photon discrimination component that reduces the front-end dead time, and a modularized acquisition system that parallelizes the sensitive detector area, so as to enable data acquisition also during therapeutic irradiation (full in-beam measurement). The PET system has been able to sustain the single photon count rates and acquire coincidences during the beam, in conditions of sub-clinical beam currents. A study on the paralyzation conditions and dead time losses under different beam currents is presented and the feasibility of a full in-beam PET scanner is discussed.

  8. Combined {sup 18}F-FDG-PET/CT Imaging in Radiotherapy Target Delineation for Head-and-Neck Cancer

    SciTech Connect

    Guido, Alessandra; Fuccio, Lorenzo; Rombi, Barbara; Castellucci, Paolo; Cecconi, Agnese; Bunkheila, Feisal; Fuccio, Chiara; Spezi, Emiliano; Angelini, Anna Lisa; Barbieri, Enza

    2009-03-01

    Purpose: To evaluate the effect of the use of {sup 18}F-fluorodeoxyglucose (FDG)-positron emission tomography (PET)/computed tomography (CT) in radiotherapy target delineation for head-and-neck cancer compared with CT alone. Methods and Materials: A total of 38 consecutive patients with head-and-neck cancer were included in this study. The primary tumor sites were as follow: 20 oropharyngeal tumors, 4 laryngeal tumors, 2 hypopharyngeal tumors, 2 paranasal sinuses tumors, 9 nasopharyngeal tumors, and 1 parotid gland tumor. The FDG-PET and CT scans were performed with a dedicated PET/CT scanner in one session and then fused. Subsequently, patients underwent treatment planning CT with intravenous contrast enhancement. The radiation oncologist defined all gross tumor volumes (GTVs) using both the PET/CT and CT scans. Results: In 35 (92%) of 38 cases, the CT-based GTVs were larger than the PET/CT-based GTVs. The average total GTV from the CT and PET/CT scans was 34.54 cm{sup 3} (range, 3.56-109) and 29.38 cm{sup 3} (range, 2.87-95.02), respectively (p < 0.05). Separate analyses of the difference between the CT- and PET/CT-based GTVs of the primary tumor compared with the GTVs of nodal disease were not statistically significant. The comparison between the PET/CT-based and CT-based boost planning target volumes did not show a statistically significant difference. All patients were alive at the end of the follow-up period (range, 3-38 months). Conclusion: GTVs, but not planning target volumes, were significantly changed by the implementation of combined PET/CT. Large multicenter studies are needed to ascertain whether combined PET/CT in target delineation can influence the main clinical outcomes.

  9. Uterotrophic and Hershberger assays for endocrine disruption properties of plastic food contact materials polypropylene (PP) and polyethylene terephthalate (PET).

    PubMed

    Chung, Bu Young; Kyung, Minji; Lim, Seong Kwang; Choi, Seul Min; Lim, Duck Soo; Kwack, Seung Jun; Kim, Hyung Sik; Lee, Byung-Mu

    2013-01-01

    Plasticizers or plastic materials such as phthalates, bisphenol-A (BPA), and styrene are widely used in the plastic industry and are suspected endocrine-disrupting chemicals (EDC). Although plastic materials such as polypropylene (PP) and polyethylene terephthalate (PET) are not EDC and are considered to be safe, their potential properties as EDC have not been fully investigated. In this study, plastic samples eluted from plastic food containers (PP or PET) were investigated in Sprague-Dawley rats using Hershberger and uterotrophic assays. In the Hershberger assay, 6-wk-old castrated male rats were orally treated for 10 consecutive days with plastic effluent at 3 different doses (5 ml/kg) or vehicle control (corn oil, 1 ml/100 g) to determine the presence of both anti-androgenic and androgenic effects. Testosterone (0.4 mg/ml/kg) was subcutaneously administered for androgenic evaluation as a positive control, whereas testosterone (0.4 mg/ml/kg) and flutamide (3 mg/kg/day) were administered to a positive control group for anti-androgenic evaluation. The presence of any anti-androgenic or androgenic activities of plastic effluent was not detected. Sex accessory tissues such as ventral prostate or seminal vesicle showed no significant differences in weight between treated and control groups. For the uterotrophic assay, immature female rats were treated with plastic effluent at three different doses (5 ml/kg), with vehicle control (corn oil, 1 ml/100 g), or with ethinyl estradiol (3 μg/kg/d) for 3 d. There were no significant differences between test and control groups in vagina or uterine weight. Data suggest that effluents from plastic food containers do not appear to produce significant adverse effects according to Hershberger and uterotrophic assays. PMID:23862761

  10. High performance ZnO:Al films deposited on PET substrates using facing target sputtering

    NASA Astrophysics Data System (ADS)

    Guo, Tingting; Dong, Guobo; Gao, Fangyuan; Xiao, Yu; Chen, Qiang; Diao, Xungang

    2013-10-01

    ZnO:Al (ZAO) thin films have been deposited on flexible PET substrates using a plasma damage-free facing target sputtering system at room temperature. The structure, surface morphology, electrical and optical properties were investigated as a function of working power. All the samples have a highly preferred orientation of the c-axis perpendicular to the PET substrate and have a high quality surface. With increased working power, the carrier concentration changes slightly, the mobility increases at the beginning and decreases after it reaches a maximum value, in line with electrical conductivity. The figure of merit has been significantly improved with increasing of the working power. Under the optimized condition, the lowest resistivity of 1.3 × 10-3 Ω cm with a sheet resistance of 29 Ω/□ and the relative visible transmittance above 93% in the visible region were obtained.

  11. Comparison of quantitative PCR assays for Escherichia coli targeting ribosomal RNA and single copy genes

    EPA Science Inventory

    Aims: Compare specificity and sensitivity of quantitative PCR (qPCR) assays targeting single and multi-copy gene regions of Escherichia coli. Methods and Results: A previously reported assay targeting the uidA gene (uidA405) was used as the basis for comparing the taxono...

  12. A survey of yeast genomic assays for drug and target discovery

    PubMed Central

    Smith, Andrew M.; Ammar, Ron; Nislow, Corey; Giaever, Guri

    2010-01-01

    Over the past decade, the development and application of chemical genomic assays using the model organism Saccharomyces cerevisiae has provided powerful methods to identify the mechanism of action of known drugs and novel small molecules in vivo. These assays identify drug target candidates, genes involved in buffering drug target pathways and also help to define the general cellular response to small molecules. In this review, we examine current yeast chemical genomic assays and summarize the potential applications of each approach. PMID:20546776

  13. Quantitative imaging of protein targets in the human brain with PET

    NASA Astrophysics Data System (ADS)

    Gunn, Roger N.; Slifstein, Mark; Searle, Graham E.; Price, Julie C.

    2015-11-01

    PET imaging of proteins in the human brain with high affinity radiolabelled molecules has a history stretching back over 30 years. During this period the portfolio of protein targets that can be imaged has increased significantly through successes in radioligand discovery and development. This portfolio now spans six major categories of proteins; G-protein coupled receptors, membrane transporters, ligand gated ion channels, enzymes, misfolded proteins and tryptophan-rich sensory proteins. In parallel to these achievements in radiochemical sciences there have also been significant advances in the quantitative analysis and interpretation of the imaging data including the development of methods for image registration, image segmentation, tracer compartmental modeling, reference tissue kinetic analysis and partial volume correction. In this review, we analyze the activity of the field around each of the protein targets in order to give a perspective on the historical focus and the possible future trajectory of the field. The important neurobiology and pharmacology is introduced for each of the six protein classes and we present established radioligands for each that have successfully transitioned to quantitative imaging in humans. We present a standard quantitative analysis workflow for these radioligands which takes the dynamic PET data, associated blood and anatomical MRI data as the inputs to a series of image processing and bio-mathematical modeling steps before outputting the outcome measure of interest on either a regional or parametric image basis. The quantitative outcome measures are then used in a range of different imaging studies including tracer discovery and development studies, cross sectional studies, classification studies, intervention studies and longitudinal studies. Finally we consider some of the confounds, challenges and subtleties that arise in practice when trying to quantify and interpret PET neuroimaging data including motion artifacts

  14. Quantitative imaging of protein targets in the human brain with PET.

    PubMed

    Gunn, Roger N; Slifstein, Mark; Searle, Graham E; Price, Julie C

    2015-11-21

    PET imaging of proteins in the human brain with high affinity radiolabelled molecules has a history stretching back over 30 years. During this period the portfolio of protein targets that can be imaged has increased significantly through successes in radioligand discovery and development. This portfolio now spans six major categories of proteins; G-protein coupled receptors, membrane transporters, ligand gated ion channels, enzymes, misfolded proteins and tryptophan-rich sensory proteins. In parallel to these achievements in radiochemical sciences there have also been significant advances in the quantitative analysis and interpretation of the imaging data including the development of methods for image registration, image segmentation, tracer compartmental modeling, reference tissue kinetic analysis and partial volume correction. In this review, we analyze the activity of the field around each of the protein targets in order to give a perspective on the historical focus and the possible future trajectory of the field. The important neurobiology and pharmacology is introduced for each of the six protein classes and we present established radioligands for each that have successfully transitioned to quantitative imaging in humans. We present a standard quantitative analysis workflow for these radioligands which takes the dynamic PET data, associated blood and anatomical MRI data as the inputs to a series of image processing and bio-mathematical modeling steps before outputting the outcome measure of interest on either a regional or parametric image basis. The quantitative outcome measures are then used in a range of different imaging studies including tracer discovery and development studies, cross sectional studies, classification studies, intervention studies and longitudinal studies. Finally we consider some of the confounds, challenges and subtleties that arise in practice when trying to quantify and interpret PET neuroimaging data including motion artifacts

  15. Targeting MT1-MMP as an ImmunoPET-Based Strategy for Imaging Gliomas

    PubMed Central

    Oteo, M.; Romero, E.; Cámara, J. A.; de Martino, A.; Arroyo, A. G.; Morcillo, M. Á.; Squatrito, M.; Martinez-Torrecuadrada, J. L.; Mulero, F.

    2016-01-01

    vivo validation showed high-specific-contrast imaging of MT1-MMP positive GBM tumors and provided strong evidence for utility of MT1-MMP-targeted immunoPET as an alternate to nonspecific imaging of GBM. PMID:27462980

  16. Site-specifically labeled CA19.9-targeted immunoconjugates for the PET, NIRF, and multimodal PET/NIRF imaging of pancreatic cancer

    PubMed Central

    Houghton, Jacob L.; Zeglis, Brian M.; Abdel-Atti, Dalya; Aggeler, Robert; Sawada, Ritsuko; Agnew, Brian J.; Scholz, Wolfgang W.; Lewis, Jason S.

    2015-01-01

    Molecular imaging agents for preoperative positron emission tomography (PET) and near-infrared fluorescent (NIRF)-guided delineation of surgical margins could greatly enhance the diagnosis, staging, and resection of pancreatic cancer. PET and NIRF optical imaging offer complementary clinical applications, enabling the noninvasive whole-body imaging to localize disease and identification of tumor margins during surgery, respectively. We report the development of PET, NIRF, and dual-modal (PET/NIRF) imaging agents, using 5B1, a fully human monoclonal antibody that targets CA19.9, a well-established pancreatic cancer biomarker. Desferrioxamine (DFO) and/or a NIRF dye (FL) were conjugated to the heavy-chain glycans of 5B1, using a robust and reproducible site-specific (ss) labeling methodology to generate three constructs (ssDFO-5B1, ssFL-5B1, and ssdual-5B1) in which the immunoreactivity was not affected by the conjugation of either label. Each construct was evaluated in a s.c. xenograft model, using CA19.9-positive (BxPC3) and -negative (MIAPaCa-2) human pancreatic cancer cell lines. Each construct showed exceptional uptake and contrast in antigen-positive tumors with negligible nonspecific uptake in antigen-negative tumors. Additionally, the dual-modal construct was evaluated in an orthotopic murine pancreatic cancer model, using the human pancreatic cancer cell line, Suit-2. The ssdual-5B1 demonstrated a remarkable capacity to delineate metastases and to map the sentinel lymph nodes via tandem PET-computed tomography (PET/CT) and NIRF imaging. Fluorescence microscopy, histopathology, and autoradiography were performed on representative sections of excised tumors to visualize the distribution of the constructs within the tumors. These imaging tools have tremendous potential for further preclinical research and for clinical translation. PMID:26668398

  17. Performance Assessment PCR-Based Assays Targeting Bacteroidales Genetic Markers of Bovine Fecal Pollution▿

    PubMed Central

    Shanks, Orin C.; White, Karen; Kelty, Catherine A.; Hayes, Sam; Sivaganesan, Mano; Jenkins, Michael; Varma, Manju; Haugland, Richard A.

    2010-01-01

    There are numerous PCR-based assays available to characterize bovine fecal pollution in ambient waters. The determination of which approaches are most suitable for field applications can be difficult because each assay targets a different gene, in many cases from different microorganisms, leading to variation in assay performance. We describe a performance evaluation of seven end-point PCR and real-time quantitative PCR (qPCR) assays reported to be associated with either ruminant or bovine feces. Each assay was tested against a reference collection of DNA extracts from 247 individual bovine fecal samples representing 11 different populations and 175 fecal DNA extracts from 24 different animal species. Bovine-associated genetic markers were broadly distributed among individual bovine samples ranging from 39 to 93%. Specificity levels of the assays spanned 47.4% to 100%. End-point PCR sensitivity also varied between assays and among different bovine populations. For qPCR assays, the abundance of each host-associated genetic marker was measured within each bovine population and compared to results of a qPCR assay targeting 16S rRNA gene sequences from Bacteroidales. Experiments indicate large discrepancies in the performance of bovine-associated assays across different bovine populations. Variability in assay performance between host populations suggests that the use of bovine microbial source-tracking applications will require a priori characterization at each watershed of interest. PMID:20061457

  18. {sup 11}C-methionine PET improves the target volume delineation of meningiomas treated with stereotactic fractionated radiotherapy

    SciTech Connect

    Grosu, Anca-Ligia . E-mail: anca-ligia.grosu@lrz.tum.de; Weber, Wolfgang A.; Astner, Sabrina T.; Adam, Markus; Krause, Bernd J.; Schwaiger, Markus; Molls, Michael; Nieder, Carsten

    2006-10-01

    Purpose: To evaluate the role of {sup 11}C-methionine positron emission tomography (MET-PET) in target volume delineation for meningiomas and to determine the interobserver variability. Methods and Materials: Two independent observers performed treatment planning in 10 patients according to a prospective written protocol. In the first step, they used coregistered computed tomography (CT) and magnetic resonance imaging (MRI). In the second step, MET-PET was added to CT/MRI (image fusion based on mutual information). Results: The correlation between gross tumor volume (GTVs) delineated by the two observers based on CT/MRI was r = 0.855 (Spearman's correlation coefficient, p = 0.002) and r = 0.988 (p = 0.000) when MET-PET/CT/MRI were used. The number of patients with agreement in more then 80% of the outlined volume increased with the availability of MET-PET from 1 in 10 to 5 in 10. The median volume of intersection between the regions delineated by two observers increased significantly from 69% (from the composite volume) to 79%, by the addition of MET-PET (p = 0.005). The information of MET-PET was useful to delineate GTV in the area of cavernous sinus, orbit, and base of the skull. Conclusions: The hypothesis-generating findings of potential normal tissue sparing and reduced interobserver variability provide arguments for invasive studies of the correlation between MET-PET images and histologic tumor extension and for prospective trials of target volume delineation with CT/MRI/MET-PET image fusion.

  19. A rapid screening assay for identifying mycobacteria targeted nanoparticle antibiotics.

    PubMed

    Donnellan, Samantha; Tran, Lang; Johnston, Helinor; McLuckie, Joyce; Stevenson, Karen; Stone, Vicki

    2016-08-01

    Antibiotic resistance is a serious problem. Nanotechnology offers enormous potential in medicine, yet there is limited knowledge regarding the toxicity of nanoparticles (NP) for mycobacterial species that cause serious human diseases (e.g. tuberculosis (TB) and leprosy). Mycobacterial diseases are a major global health problem; TB caused by Mycobacterium tuberculosis (Mtb) kills up to 2 million people annually and there are over 200 000 leprosy cases each year caused by Mycobacterium leprae (M. leprae). Few drugs are effective against these mycobacteria and increasing antibiotic resistance exacerbates the problem. As such, alternative therapies are urgently needed but most current assays used to assess the effectiveness of therapeutics against mycobacteria are slow and expensive. This study aimed to develop a rapid, low-cost assay which can be used for screening the antimicrobial properties of compounds against pathogenic mycobacteria and to assess the toxicity of three NP (silver [Ag], copper oxide [Cu(II)O], and zinc oxide [ZnO]) against a green fluorescent protein reporter strain of Mycobacterium avium subspecies paratuberculosis, a slow growing, pathogenic mycobacterial species causing paratuberculosis in ruminants. Fluorescence was used to monitor mycobacterial growth over time, with NP concentrations of 6.25-100 μg/mL tested for up to 7 days, and a method of data analysis was designed to permit comparison between results. Mycobacterial sensitivity to the NP was found to be NP composition specific and toxicity could be ranked in the following order: Ag > Cu(II)O > ZnO. PMID:26618564

  20. Very Late Antigen-4 (α4β1 Integrin) Targeted PET Imaging of Multiple Myeloma

    PubMed Central

    Soodgupta, Deepti; Hurchla, Michelle A.; Jiang, Majiong; Zheleznyak, Alexander; Weilbaecher, Katherine N.; Anderson, Carolyn J.; Tomasson, Michael H.; Shokeen, Monica

    2013-01-01

    Biomedical imaging techniques such as skeletal survey and 18F-fluorodeoxyglucose (FDG)/Positron Emission Tomography (PET) are frequently used to diagnose and stage multiple myeloma (MM) patients. However, skeletal survey has limited sensitivity as it can detect osteolytic lesions only after 30–50% cortical bone destruction, and FDG is a marker of cell metabolism that has limited sensitivity for intramedullary lesions in MM. Targeted, and non-invasive novel probes are needed to sensitively and selectively image the unique molecular signatures and cellular processes associated with MM. Very late antigen-4 (VLA-4; also called α4β1 integrin) is over-expressed on MM cells, and is one of the key mediators of myeloma cell adhesion to the bone marrow (BM) that promotes MM cell trafficking and drug resistance. Here we describe a proof-of-principle, novel molecular imaging strategy for MM tumors using a VLA-4 targeted PET radiopharmaceutical, 64Cu-CB-TE1A1P-LLP2A. Cell uptake studies in a VLA-4-positive murine MM cell line, 5TGM1, demonstrated receptor specific uptake (P<0.0001, block vs. non-block). Tissue biodistribution at 2 h of 64Cu-CB-TE1A1P-LLP2A in 5TGM1 tumor bearing syngeneic KaLwRij mice demonstrated high radiotracer uptake in the tumor (12±4.5%ID/g), and in the VLA-4 rich organs, spleen (8.8±1.0%ID/g) and marrow (11.6±2.0%ID/g). Small animal PET/CT imaging with 64Cu-CB-TE1A1P-LLP2A demonstrated high uptake in the 5TGM1 tumors (SUV 6.6±1.1). There was a 3-fold reduction in the in vivo tumor uptake in the presence of blocking agent (2.3±0.4). Additionally, 64Cu-CB-TE1A1P-LLP2A demonstrated high binding to the human MM cell line RPMI-8226 that was significantly reduced in the presence of the cold targeting agent. These results provide pre-clinical evidence that VLA-4-targeted imaging using 64Cu-CB-TE1A1P-LLP2A is a novel approach to imaging MM tumors. PMID:23409060

  1. Method for nondestructive fuel assay of laser fusion targets

    DOEpatents

    Farnum, Eugene H.; Fries, R. Jay

    1976-01-01

    A method for nondestructively determining the deuterium and tritium content of laser fusion targets by counting the x rays produced by the interaction of tritium beta particles with the walls of the microballoons used to contain the deuterium and tritium gas mixture under high pressure. The x rays provide a direct measure of the tritium content and a means for calculating the deuterium content using the initial known D-T ratio and the known deuterium and tritium diffusion rates.

  2. Improved receptor analysis in PET using a priori information from in vitro binding assays

    NASA Astrophysics Data System (ADS)

    Litton, J.-E.; Hall, H.; Blomqvist, G.

    1997-08-01

    An accurate determination of non-specific binding is required for the analysis of in vitro and in vivo receptor binding data. For some radioligands the non-specific binding is of the same magnitude as the specific binding. Furthermore, in vitro measurements have shown that the non-specific binding can be different in different brain regions. If this is the case in a PET study for determining and , a correction for the non-specific binding has to be applied. The aim of the present communication is to present a means for determining corrected and with Scatchard analysis using in vitro binding studies. The influence of non-specific binding on the free and specifically bound radioligand is expressed with the aid of a correction factor, which can be calculated from measurable quantities. Introduction of the corrected free and specifically bound radioligand should give binding parameters closer to reality than previously obtained results.

  3. The Cellular Thermal Shift Assay: A Novel Biophysical Assay for In Situ Drug Target Engagement and Mechanistic Biomarker Studies.

    PubMed

    Martinez Molina, Daniel; Nordlund, Pär

    2016-01-01

    A drug must engage its intended target to achieve its therapeutic effect. However, conclusively measuring target engagement (TE) in situ is challenging. This complicates preclinical development and is considered a key factor in the high rate of attrition in clinical trials. Here, we discuss a recently developed, label-free, biophysical assay, the cellular thermal shift assay (CETSA), which facilitates the direct assessment of TE in cells and tissues at various stages of drug development. CETSA also reveals biochemical events downstream of drug binding and therefore provides a promising means of establishing mechanistic biomarkers. The implementation of proteome-wide CETSA using quantitative mass spectrometry represents a novel strategy for defining off-target toxicity and polypharmacology and for identifying downstream mechanistic biomarkers. The first year of CETSA applications in the literature has focused on TE studies in cell culture systems and has confirmed the broad applicability of CETSA to many different target families. The next phase of CETSA applications will likely encompass comprehensive animal and patient studies, and CETSA will likely serve as a very valuable tool in many stages of preclinical and clinical drug development. PMID:26566155

  4. Evaluation of (68)Ga- and (177)Lu-DOTA-PEG4-LLP2A for VLA-4-Targeted PET Imaging and Treatment of Metastatic Melanoma.

    PubMed

    Beaino, Wissam; Nedrow, Jessie R; Anderson, Carolyn J

    2015-06-01

    Malignant melanoma is a highly aggressive cancer, and the incidence of this disease is increasing worldwide at an alarming rate. Despite advances in the treatment of melanoma, patients with metastatic disease still have a poor prognosis and low survival rate. New strategies, including targeted radiotherapy, would provide options for patients who become resistant to therapies such as BRAF inhibitors. Very late antigen-4 (VLA-4) is expressed on melanoma tumor cells in higher levels in more aggressive and metastatic disease and may provide an ideal target for drug delivery and targeted radiotherapy. In this study, we evaluated (177)Lu- and (68)Ga-labeled DOTA-PEG4-LLP2A as a VLA-4-targeted radiotherapeutic with a companion PET agent for diagnosis and monitoring metastatic melanoma treatment. DOTA-PEG4-LLP2A was synthesized by solid-phase synthesis. The affinity of (177)Lu- and (68)Ga-labeled DOTA-PEG4-LLP2A to VLA-4 was determined in B16F10 melanoma cells by saturation binding and competitive binding assays, respectively. Biodistribution of the LLP2A conjugates was determined in C57BL/6 mice bearing B16F10 subcutaneous tumors, while PET/CT imaging was performed in subcutaneous and metastatic models. (177)Lu-DOTA-PEG4-LLP2A showed high affinity to VLA-4 with a Kd of 4.1 ± 1.5 nM and demonstrated significant accumulation in the B16F10 melanoma tumor after 4 h (31.5 ± 7.8%ID/g). The tumor/blood ratio of (177)Lu-DOTA-PEG4-LLP2A was highest at 24 h (185 ± 26). PET imaging of metastatic melanoma with (68)Ga-DOTA-PEG4-LLP2A showed high uptake in sites of metastases and correlated with bioluminescence imaging of the tumors. These data demonstrate that (177)Lu-DOTA-PEG4-LLP2A has potential as a targeted therapeutic for treating melanoma as well as other VLA-4-expressing tumors. In addition, (68)Ga-DOTA-PEG4-LLP2A is a readily translatable companion PET tracer for imaging of metastatic melanoma. PMID:25919487

  5. Quantification of Human Kallikrein-Related Peptidases in Biological Fluids by Multiplatform Targeted Mass Spectrometry Assays.

    PubMed

    Karakosta, Theano D; Soosaipillai, Antoninus; Diamandis, Eleftherios P; Batruch, Ihor; Drabovich, Andrei P

    2016-09-01

    Human kallikrein-related peptidases (KLKs) are a group of 15 secreted serine proteases encoded by the largest contiguous cluster of protease genes in the human genome. KLKs are involved in coordination of numerous physiological functions including regulation of blood pressure, neuronal plasticity, skin desquamation, and semen liquefaction, and thus represent promising diagnostic and therapeutic targets. Until now, quantification of KLKs in biological and clinical samples was accomplished by enzyme-linked immunosorbent assays (ELISA). Here, we developed multiplex targeted mass spectrometry assays for the simultaneous quantification of all 15 KLKs. Proteotypic peptides for each KLK were carefully selected based on experimental data and multiplexed in single assays. Performance of assays was evaluated using three different mass spectrometry platforms including triple quadrupole, quadrupole-ion trap, and quadrupole-orbitrap instruments. Heavy isotope-labeled synthetic peptides with a quantifying tag were used for absolute quantification of KLKs in sweat, cervico-vaginal fluid, seminal plasma, and blood serum, with limits of detection ranging from 5 to 500 ng/ml. Analytical performance of assays was evaluated by measuring endogenous KLKs in relevant biological fluids, and results were compared with selected ELISAs. The multiplex targeted proteomic assays were demonstrated to be accurate, reproducible, sensitive, and specific alternatives to antibody-based assays. Finally, KLK4, a highly prostate-specific protein and a speculated biomarker of prostate cancer, was unambiguously detected and quantified by immunoenrichment-SRM assay in seminal plasma and blood serum samples from individuals with confirmed prostate cancer and negative biopsy. Mass spectrometry revealed exclusively the presence of a secreted isoform and thus unequivocally resolved earlier disputes about KLK4 identity in seminal plasma. Measurements of KLK4 in either 41 seminal plasma or 58 blood serum samples

  6. DNA Sequence Signatures for Rapid Detection of Six Target Bacterial Pathogens Using PCR Assays

    PubMed Central

    Nagamine, Kenjiro; Hung, Guo-Chiuan; Li, Bingjie; Lo, Shyh-Ching

    2015-01-01

    Using Streptococcus pyogenes as a model, we previously established a stepwise computational workflow to effectively identify species-specific DNA signatures that could be used as PCR primer sets to detect target bacteria with high specificity and sensitivity. In this study, we extended the workflow for the rapid development of PCR assays targeting Enterococcus faecalis, Enterococcus faecium, Clostridium perfringens, Clostridium difficile, Clostridium tetani, and Staphylococcus aureus, which are of safety concern for human tissue intended for transplantation. Twenty-one primer sets that had sensitivity of detecting 5–50 fg DNA from target bacteria with high specificity were selected. These selected primer sets can be used in a PCR array for detecting target bacteria with high sensitivity and specificity. The workflow could be widely applicable for the rapid development of PCR-based assays for a wide range of target bacteria, including those of biothreat agents. PMID:26279626

  7. DNA Sequence Signatures for Rapid Detection of Six Target Bacterial Pathogens Using PCR Assays.

    PubMed

    Nagamine, Kenjiro; Hung, Guo-Chiuan; Li, Bingjie; Lo, Shyh-Ching

    2015-01-01

    Using Streptococcus pyogenes as a model, we previously established a stepwise computational workflow to effectively identify species-specific DNA signatures that could be used as PCR primer sets to detect target bacteria with high specificity and sensitivity. In this study, we extended the workflow for the rapid development of PCR assays targeting Enterococcus faecalis, Enterococcus faecium, Clostridium perfringens, Clostridium difficile, Clostridium tetani, and Staphylococcus aureus, which are of safety concern for human tissue intended for transplantation. Twenty-one primer sets that had sensitivity of detecting 5-50 fg DNA from target bacteria with high specificity were selected. These selected primer sets can be used in a PCR array for detecting target bacteria with high sensitivity and specificity. The workflow could be widely applicable for the rapid development of PCR-based assays for a wide range of target bacteria, including those of biothreat agents. PMID:26279626

  8. Evaluation of the migration of mutagens/carcinogens from PET bottles into mineral water by Tradescantia/micronuclei test, Comet assay on leukocytes and GC/MS.

    PubMed

    Biscardi, D; Monarca, S; De Fusco, R; Senatore, F; Poli, P; Buschini, A; Rossi, C; Zani, C

    2003-01-20

    This study monitored the release of mutagenic/carcinogenic compounds into mineral water (natural and carbonated) from polyethylene terephthalate (PET) bottles, using a plant mutagenicity test which reveals micronuclei formation in Tradescantia pollen cells (Trad/MCN test), a DNA damage assay (Comet assay) on human leukocytes and gas chromatography/mass spectrometry (GC/MS) for the characterisation of migrants. The water samples were collected at a bottling plant and stored in PET bottles for a period ranging from 1 to 12 months. Every month some samples were randomly collected and lyophilised, the residual powders were extracted with organic solvents and then analysed by GC/MS and tested for DNA damage in human leukocytes, or reconstituted with distilled water to obtain concentrates for the exposure of Tradescantia inflorescences. Micronuclei increase in pollen was found only in natural mineral water stored for 2 months. DNA-damaging activity was found in many of the natural and carbonated water samples. Spring water was negative in the plant micronuclei test and the Comet assay, whereas distributed spring water showed DNA-damaging effects, suggesting a possible introduction of genotoxins through the distribution pipelines. GC/MS analysis showed the presence in mineral water of di(2-ethylhexyl)phthalate, a nongenotoxic hepatocarcinogenic plasticizer, after 9 months of storage in PET bottles. PMID:12526902

  9. Characteristics of AZO thin films prepared at various Al target input current deposited on PET substrate

    NASA Astrophysics Data System (ADS)

    Kim, Yun-Hae; Park, Chang-Wook; Lee, Jin-Woo; Lee, Dong Myung

    2015-03-01

    Transparent conductive oxide is a thin film to be used in numerous applications throughout the industry in general. Transparent electrode materials used in these industries are in need of light transmittance with excellent high and low electrical characteristics, substances showing the most excellent physical properties while satisfying all the characteristics such as indium tin oxide film. However, reserves of indium are very small, there is an environmental pollution problem. So the study of zinc oxide (ZnO) is actively carried out in an alternative material. This study analyzed the characteristics by using a direct current (DC) magnetron sputtering system. The electric and optical properties of these films were studied by Hall measurement and optical spectroscopy, respectively. When the Al target input current is 2 mA and 4 mA, it demonstrates about 80% transmittance in the range of the visible spectrum. Also, when Al target input current was 6 mA, sheet resistance was the smallest on PET substrate. The minimum resistivity is 3.96×10-3 ohm/sq.

  10. Sensitive, simultaneous quantitation of two unlabeled DNA targets using a magnetic nanoparticle-enzyme sandwich assay.

    PubMed

    Zhang, Yue; Pilapong, Chalermchai; Guo, Yuan; Ling, Zhenlian; Cespedes, Oscar; Quirke, Philip; Zhou, Dejian

    2013-10-01

    We report herein the development of a simple, sensitive colorimetric magnetic nanoparticle (MNP)-enzyme-based DNA sandwich assay that is suitable for simultaneous, label-free quantitation of two DNA targets down to 50 fM level. It can also effectively discriminate single-nucleotide polymorphisms (SNPs) in genes associated with human cancers (KRAS codon 12/13 SNPs). This assay uses a pair of specific DNA probes, one being covalently conjugated to an MNP for target capture and the other being linked to an enzyme for signal amplification, to sandwich a DNA target, allowing for convenient magnetic separation and subsequent efficient enzymatic signal amplification for high sensitivity. Careful optimization of the MNP surfaces and assay conditions greatly reduced the background, allowing for sensitive, specific detection of as little as 5 amol (50 fM in 100 μL) of target DNA. Moreover, this sensor is robust, it can effectively discriminate cancer-specific SNPs against the wild-type noncancer target, and it works efficiently in 10% human serum. Furthermore, this sensor can simultaneously quantitate two different DNA targets by using two pairs of unique capture- and signal-DNA probes specific for each target. This general, simple, and sensitive DNA sensor appears to be well-suited for a wide range of genetics-based biosensing and diagnostic applications. PMID:23971744

  11. Sensitive, Simultaneous Quantitation of Two Unlabeled DNA Targets Using a Magnetic Nanoparticle–Enzyme Sandwich Assay

    PubMed Central

    2013-01-01

    We report herein the development of a simple, sensitive colorimetric magnetic nanoparticle (MNP)–enzyme-based DNA sandwich assay that is suitable for simultaneous, label-free quantitation of two DNA targets down to 50 fM level. It can also effectively discriminate single-nucleotide polymorphisms (SNPs) in genes associated with human cancers (KRAS codon 12/13 SNPs). This assay uses a pair of specific DNA probes, one being covalently conjugated to an MNP for target capture and the other being linked to an enzyme for signal amplification, to sandwich a DNA target, allowing for convenient magnetic separation and subsequent efficient enzymatic signal amplification for high sensitivity. Careful optimization of the MNP surfaces and assay conditions greatly reduced the background, allowing for sensitive, specific detection of as little as 5 amol (50 fM in 100 μL) of target DNA. Moreover, this sensor is robust, it can effectively discriminate cancer-specific SNPs against the wild-type noncancer target, and it works efficiently in 10% human serum. Furthermore, this sensor can simultaneously quantitate two different DNA targets by using two pairs of unique capture- and signal-DNA probes specific for each target. This general, simple, and sensitive DNA sensor appears to be well-suited for a wide range of genetics-based biosensing and diagnostic applications. PMID:23971744

  12. SU-E-CAMPUS-I-06: Y90 PET/CT for the Instantaneous Determination of Both Target and Non-Target Absorbed Doses Following Hepatic Radioembolization

    SciTech Connect

    Pasciak, A; Kao, J

    2014-06-15

    Purpose The process of converting Yttrium-90 (Y90) PET/CT images into 3D absorbed dose maps will be explained. The simple methods presented will allow the medical physicst to analyze Y90 PET images following radioembolization and determine the absorbed dose to tumor, normal liver parenchyma and other areas of interest, without application of Monte-Carlo radiation transport or dose-point-kernel (DPK) convolution. Methods Absorbed dose can be computed from Y90 PET/CT images based on the premise that radioembolization is a permanent implant with a constant relative activity distribution after infusion. Many Y90 PET/CT publications have used DPK convolution to obtain 3D absorbed dose maps. However, this method requires specialized software limiting clinical utility. The Local Deposition method, an alternative to DPK convolution, can be used to obtain absorbed dose and requires no additional computer processing. Pixel values from regions of interest drawn on Y90 PET/CT images can be converted to absorbed dose (Gy) by multiplication with a scalar constant. Results There is evidence that suggests the Local Deposition method may actually be more accurate than DPK convolution and it has been successfully used in a recent Y90 PET/CT publication. We have analytically compared dose-volume-histograms (DVH) for phantom hot-spheres to determine the difference between the DPK and Local Deposition methods, as a function of PET scanner point-spread-function for Y90. We have found that for PET/CT systems with a FWHM greater than 3.0 mm when imaging Y90, the Local Deposition Method provides a more accurate representation of DVH, regardless of target size than DPK convolution. Conclusion Using the Local Deposition Method, post-radioembolization Y90 PET/CT images can be transformed into 3D absorbed dose maps of the liver. An interventional radiologist or a Medical Physicist can perform this transformation in a clinical setting, allowing for rapid prediction of treatment efficacy by

  13. Evaluation of 64Cu-Labeled Acridinium Cation: A PET Radiotracer Targeting Tumor Mitochondria

    PubMed Central

    Zhou, Yang; Kim, Young-Seung; Shi, Jiyun; Jacobson, Orit; Chen, Xiaoyuan; Liu, Shuang

    2011-01-01

    This report presents the synthesis and evaluations of 64Cu(DO3A-xy-ACR) (DO3A-xy-ACR = 2,6-bis(dimethylamino)-10-(4-((4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl)methyl)benzyl)acridin-10-ium) as a radiotracer for imaging tumors in athymic nude mice bearing U87MG glioma xenografts by PET (positron emission tomography). The biodistribution data suggested that 64Cu(DO3A-xy-ACR) was excreted mainly through the renal system with >65% of injected radioactivity being recovered from urine samples at 1 h post-injection (p.i.). The tumor uptake of 64Cu(DO3A-xy-ACR) was 1.07 ± 0.23, 1.58 ± 0.55, 2.71 ± 0.66, 3.47 ± 1.19, and 3.52 ± 1.72 %ID/g at 0.5, 1, 2, 4 and 24 h p.i., respectively. 64Cu(DO3A-xy-ACR) had very high liver uptake (31.90 ± 3.98, 24.95 ± 5.64, 15.20 ± 4.29, 14.09 ± 6.82, and 8.18 ± 1.27 %ID/g at 0.5, 1, 2, 4 and 24 h p.i., respectively) with low tumor/liver ratios. MicroPET studies showed that the tumors were clearly visualized as early as 30 min p.i. in the glioma-bearing mouse administered with 64Cu(DO3A-xy-ACR). The high liver radioactivity accumulation was also seen. 64Cu(DO3A-xy-ACR) had a relatively high metabolic stability during excretion via both renal and hepatobiliary routes; but it was completely decomposed in the liver homogenate. We explored the localization mechanism of Cu(DO3A-xy-ACR) using both U87MG human glioma and the cultured primary U87MG glioma cells. The results from the cellular staining assays showed that 64Cu(DO3A-xy-ACR) is able to localize in the mitochondria of living U87MG glioma cells due to the enhanced negative mitochondrial potential as compared to normal cells. Although 64Cu(DO3A-xy-ACR) is not an ideal PET radiotracer for tumor imaging due to its high liver uptake, the results from this study strongly suggest that 64Cu-labeled acridinium cations are indeed able to localize in the energized mitochondria of tumor cells. PMID:21413736

  14. Activity, assay and target data curation and quality in the ChEMBL database.

    PubMed

    Papadatos, George; Gaulton, Anna; Hersey, Anne; Overington, John P

    2015-09-01

    The emergence of a number of publicly available bioactivity databases, such as ChEMBL, PubChem BioAssay and BindingDB, has raised awareness about the topics of data curation, quality and integrity. Here we provide an overview and discussion of the current and future approaches to activity, assay and target data curation of the ChEMBL database. This curation process involves several manual and automated steps and aims to: (1) maximise data accessibility and comparability; (2) improve data integrity and flag outliers, ambiguities and potential errors; and (3) add further curated annotations and mappings thus increasing the usefulness and accuracy of the ChEMBL data for all users and modellers in particular. Issues related to activity, assay and target data curation and integrity along with their potential impact for users of the data are discussed, alongside robust selection and filter strategies in order to avoid or minimise these, depending on the desired application. PMID:26201396

  15. Automated Radiation Targeting in Head-and-Neck Cancer Using Region-Based Texture Analysis of PET and CT Images

    SciTech Connect

    Yu Huan; Caldwell, Curtis Mah, Katherine; Poon, Ian; Balogh, Judith; MacKenzie, Robert; Khaouam, Nader; Tirona, Romeo

    2009-10-01

    Purpose: A co-registered multimodality pattern analysis segmentation system (COMPASS) was developed to automatically delineate the radiation targets in head-and-neck cancer (HNC) using both {sup 18}F-fluoro-deoxy glucose-positron emission tomography (PET) and computed tomography (CT) images. The performance of the COMPASS was compared with the results of existing threshold-based methods and radiation oncologist-drawn contours. Methods and Materials: The COMPASS extracted texture features from corresponding PET and CT voxels. Using these texture features, a decision-tree-based K-nearest-neighbor classifier labeled each voxel as either 'normal' or 'abnormal.' The COMPASS was applied to the PET/CT images of 10 HNC patients. Automated segmentation results were validated against the manual segmentations of three radiation oncologists using the volume, sensitivity, and specificity. The performance of the COMPASS was compared with three PET-based threshold methods: standard uptake value of 2.5, 50% maximal intensity, and signal/background ratio. Results: The tumor delineations of the COMPASS were both quantitatively and qualitatively more similar to those of the radiation oncologists than the delineations from the other methods. The specificity was 95% {+-} 2%, 84% {+-} 9%, 98% {+-} 3%, and 96% {+-} 4%, and the sensitivity was 90% {+-} 12%, 93% {+-} 10%, 48% {+-} 20%, and 68% {+-} 25% for the COMPASS, for a standard uptake value of 2.5, 50% maximal intensity, and signal/background ratio, respectively. The COMPASS distinguished HNC from adjacent normal tissues with high physiologic uptake and consistently defined tumors with large variability in {sup 18}F-fluoro-deoxy glucose uptake, which are often problematic with the threshold-based methods. Conclusion: Automated segmentation using texture analysis of PET/CT images has the potential to provide accurate delineation of HNC. This could lead to reduced interobserver variability, reduced uncertainty in target delineation

  16. Target-driven self-assembly of stacking deoxyribonucleic acids for highly sensitive assay of proteins.

    PubMed

    Cao, Ya; Chen, Weiwei; Han, Peng; Wang, Zhuxin; Li, Genxi

    2015-08-26

    In this paper, we report a new signal amplification strategy for highly sensitive and enzyme-free method to assay proteins based on the target-driven self-assembly of stacking deoxyribonucleic acids (DNA) on an electrode surface. In the sensing procedure, binding of target protein with the aptamer probe is used as a starting point for a scheduled cycle of DNA hairpin assembly, which consists of hybridization, displacement and target regeneration. Following numbers of the assembly repeats, a great deal of DNA duplexes can accordingly be formed on the electrode surface, and then switch on a succeeding propagation of self-assembled DNA concatemers that provide further signal enhancement. In this way, each target binding event can bring out two cascaded DNA self-assembly processes, namely, stacking DNA self-assembly, and therefore can be converted into remarkably intensified electrochemical signals by associating with silver nanoparticle-based readout. Consequently, highly sensitive detection of target proteins can be achieved. Using interferon-gamma as a model, the assay method displays a linear range from 1 to 500 pM with a detection limit of 0.57 pM, which is comparable or even superior to other reported amplified assays. Moreover, the proposed method eliminates the involvement of any enzymes, thereby enhancing the feasibility in clinical diagnosis. PMID:26347164

  17. Chloroplast envelope protein targeting fidelity is independent of cytosolic components in dual organelle assays

    PubMed Central

    Kriechbaumer, Verena; Abell, Ben M.

    2012-01-01

    The general mechanisms of intracellular protein targeting are well established, and depend on a targeting sequence in the protein, which is recognized by a targeting factor. Once a membrane protein is delivered to the correct organelle its targeting sequence can be recognized by receptors and a translocase, leading to membrane insertion. However, the relative contribution of each step for generating fidelity and efficiency of the overall process has not been systematically addressed. Here, we use tail-anchored (TA) membrane proteins in cell-free competitive targeting assays to chloroplasts to show that targeting can occur efficiently and with high fidelity in the absence of all cytosolic components, suggesting that chloroplast envelope protein targeting is primarily dependent on events at the outer envelope. Efficiency of targeting was increased by the addition of complete cytosol, and by Hsp70 or Hsp90, depending on the protein, but none of these cytosolic components influenced the fidelity of targeting. Our results suggest that the main role of targeting factors in chloroplast localization is to increase targeting efficiency by maintaining recognition competency at the outer envelope. PMID:22783268

  18. Establishment of two quantitative nested qPCR assays targeting the human EPO transgene.

    PubMed

    Neuberger, E W I; Perez, I; Le Guiner, C; Moser, D; Ehlert, T; Allais, M; Moullier, P; Simon, P; Snyder, R O

    2016-04-01

    For ethical and safety reasons it is critical to develop easily implemented assays with high sensitivity and specificity for gene doping surveillance. Two nested quantitative real-time PCR (qPCR) assays were developed that target the human EPO (hEPO) cDNA sequence in a circular form, representative of recombinant adeno-associated viral (rAAV) vector genomes found in vivo. Through an interlaboratory evaluation, the assays were validated and utilized in an in vitro blinded study. These assays are specific and extremely sensitive with a limit of detection (LOD) of 1 copy of circular plasmid DNA and a limit of quantification (LOQ) of 10 to 20 copies in the presence of 500 ng of human genomic DNA (hgDNA) extracted from WBCs. Additionally, using the two nested qPCR assays in a non-human primate study, where macaques were injected intramuscularly with a rAAV8 vector harboring a promoterless hEPO cDNA sequence, the viral vector was detected 8 to 14 weeks post-injection in macaque WBCs. The high sensitivity of the nested qPCR approach along with the capability of quantifying target DNA, make this approach a reliable tool for gene doping surveillance and the monitoring of exogenous DNA sequences. PMID:26752352

  19. Commercially available antibodies can be applied in quantitative multiplexed peptide immunoaffinity enrichment targeted mass spectrometry assays

    PubMed Central

    Schoenherr, Regine M.; Zhao, Lei; Ivey, Richard G.; Voytovich, Uliana J.; Kennedy, Jacob; Yan, Ping; Lin, Chenwei; Whiteaker, Jeffrey R.; Paulovich, Amanda G.

    2016-01-01

    Immunoaffinity enrichment of peptides coupled to multiple reaction monitoring-mass spectrometry (immuno-MRM) enables highly specific, sensitive, and precise quantification of peptides and post-translational modifications. Major obstacles to developing a large number of immuno-MRM assays are the poor availability of monoclonal antibodies (mAbs) validated for immunoaffinity enrichment of peptides and the cost and lead time of developing the antibodies de novo. Although many thousands of mAbs are commercially offered, few have been tested for application to immunoaffinity enrichment of peptides. In this study we tested the success rate of using commercially available mAbs for peptide immuno-MRM assays. We selected 105 commercial mAbs (76 targeting non-modified “pan” epitopes, 29 targeting phosphorylation) to proteins associated with the DNA damage response network. We found that 8 of the 76 pan (11%) and 5 of the 29 phospho-specific mAbs (17%) captured tryptic peptides (detected by LC-MS/MS) of their protein targets from human cell lysates. Seven of these mAbs were successfully used to configure and analytically characterize immuno-MRM assays. By applying selection criteria upfront, the results indicate that a screening success rate of up to 24% is possible, establishing the feasibility of screening a large number of catalog antibodies to provide readily-available assay reagents. PMID:27094115

  20. Platelet hexosaminidase a enzyme assay effectively detects carriers missed by targeted DNA mutation analysis.

    PubMed

    Nakagawa, Sachiko; Zhan, Jie; Sun, Wei; Ferreira, Jose Carlos; Keiles, Steven; Hambuch, Tina; Kammesheidt, Anja; Mark, Brian L; Schneider, Adele; Gross, Susan; Schreiber-Agus, Nicole

    2012-01-01

    Biochemical testing of hexosaminidase A (HexA) enzyme activity has been available for decades and has the ability to detect almost all Tay-Sachs disease (TSD) carriers, irrespective of ethnic background. This is increasingly important, as the gene pool of those who identify as Ashkenazi Jewish is diversifying. Here we describe the analysis of a cohort of 4,325 individuals arising from large carrier screening programs and tested by the serum and/or platelet HexA enzyme assays and by targeted DNA mutation analysis. Our results continue to support the platelet assay as a highly effective method for TSD carrier screening, with a low inconclusive rate and the ability to detect possible disease-causing mutation carriers that would have been missed by targeted DNA mutation analysis. Sequence analysis performed on one such platelet assay carrier, who had one non-Ashkenazi Jewish parent, identified the amino acid change Thr259Ala (A775G). Based on crystallographic modeling, this change is predicted to be deleterious, as threonine 259 is positioned proximal to the HexA alpha subunit active site and helps to stabilize key residues therein. Accordingly, if individuals are screened for TSD in broad-based programs by targeted molecular testing alone, they must be made aware that there is a more sensitive and inexpensive test available that can identify additional carriers. Alternatively, the enzyme assays can be offered as a first tier test, especially when screening individuals of mixed or non-Jewish ancestry. PMID:23430931

  1. Meat Species Identification using Loop-mediated Isothermal Amplification Assay Targeting Species-specific Mitochondrial DNA

    PubMed Central

    2014-01-01

    Meat source fraud and adulteration scandals have led to consumer demands for accurate meat identification methods. Nucleotide amplification assays have been proposed as an alternative method to protein-based assays for meat identification. In this study, we designed Loop-mediated isothermal amplification (LAMP) assays targeting species-specific mitochondrial DNA to identify and discriminate eight meat species; cattle, pig, horse, goat, sheep, chicken, duck, and turkey. The LAMP primer sets were designed and the target genes were discriminated according to their unique annealing temperature generated by annealing curve analysis. Their unique annealing temperatures were found to be 85.56±0.07℃ for cattle, 84.96±0.08℃ for pig, and 85.99±0.05℃ for horse in the BSE-LAMP set (Bos taurus, Sus scrofa domesticus and Equus caballus); 84.91±0.11℃ for goat and 83.90±0.11℃ for sheep in the CO-LAMP set (Capra hircus and Ovis aries); and 86.31±0.23℃ for chicken, 88.66±0.12℃ for duck, and 84.49±0.08℃ for turkey in the GAM-LAMP set (Gallus gallus, Anas platyrhynchos and Meleagris gallopavo). No cross-reactivity was observed in each set. The limits of detection (LODs) of the LAMP assays in raw and cooked meat were determined from 10 pg/μL to 100 fg/μL levels, and LODs in raw and cooked meat admixtures were determined from 0.01% to 0.0001% levels. The assays were performed within 30 min and showed greater sensitivity than that of the PCR assays. These novel LAMP assays provide a simple, rapid, accurate, and sensitive technology for discrimination of eight meat species. PMID:26761677

  2. Optimal de novo design of MRM experiments for rapid assay development in targeted proteomics.

    PubMed

    Bertsch, Andreas; Jung, Stephan; Zerck, Alexandra; Pfeifer, Nico; Nahnsen, Sven; Henneges, Carsten; Nordheim, Alfred; Kohlbacher, Oliver

    2010-05-01

    Targeted proteomic approaches such as multiple reaction monitoring (MRM) overcome problems associated with classical shotgun mass spectrometry experiments. Developing MRM quantitation assays can be time consuming, because relevant peptide representatives of the proteins must be found and their retention time and the product ions must be determined. Given the transitions, hundreds to thousands of them can be scheduled into one experiment run. However, it is difficult to select which of the transitions should be included into a measurement. We present a novel algorithm that allows the construction of MRM assays from the sequence of the targeted proteins alone. This enables the rapid development of targeted MRM experiments without large libraries of transitions or peptide spectra. The approach relies on combinatorial optimization in combination with machine learning techniques to predict proteotypicity, retention time, and fragmentation of peptides. The resulting potential transitions are scheduled optimally by solving an integer linear program. We demonstrate that fully automated construction of MRM experiments from protein sequences alone is possible and over 80% coverage of the targeted proteins can be achieved without further optimization of the assay. PMID:20201589

  3. SU-E-I-81: Targeting of HER2-Expressing Tumors with Dual PET-MR Imaging Probes

    SciTech Connect

    Xu, P; Peng, Y; Sun, M; Yang, X

    2015-06-15

    Purpose: The detection of human epidermal growth factor receptor type 2 (HER2) expression in malignant tumors provides important information influencing patient management. Radionuclide in vivo imaging of HER2 may permit the detection of HER2 in both primary tumors and metastases by a single noninvasive procedure. Trastuzumab, effective in about 15 % of women with breast cancer, downregulates signalling through the Akt/PI3K and MAPK pathways.These pathways modulate metabolism which can be monitored by positron emission tomography (PET) and magnetic resonance imaging (MRI). Methods: The relationship between response of HER2 overexpressing tumours and changes in imaging PET or SPECT and MRI will be examined by a integrated bimodal imaging probe.Small (7 kDa) high-affinity anti-HER2 Affibody molecules and KCCYSL targeting peptide may be suitable tracers for visualization of HER2-expressing tumors. Peptide-conjugated iron oxide nanoparticles (Fe3O4 NPs) as MRI imaging and CB-TE2A as PET imaging are integrated into a single synthetic molecule in the HER2 positive cancer. Results: One of targeted contrast bimodal imaging probe agents was synthesized and evaluated to target HER2-expressing tumors in a HER2 positive rat model. We will report the newest results regarding the development of bimodal imaging probes. Conclusion: The preliminary results of the bimodal imaging probe presents high correlation of MRI signal and PET imaging intensity in vivo. This unique feature can hardly be obtained by single model contrast agents. It is envisioned that this bimodal agents can hold great potential for accurate detection of HER2-expressing tumors which are critical for clinical management of the disease.

  4. A high-affinity [18F]-labeled phosphoramidate peptidomimetic PSMA-targeted inhibitor for PET imaging of prostate cancer

    PubMed Central

    Ganguly, Tanushree; Dannoon, Shorouk; Hopkins, Mark R.; Murphy, Stephanie; Cahaya, Hendry; Blecha, Joseph E.; Jivan, Salma; Drake, Christopher R.; Barinka, Cyril; Jones, Ella F.; VanBrocklin, Henry F.; Berkman, Clifford E.

    2015-01-01

    Introduction In this study, a structurally modified phosphoramidate scaffold, with improved prostate-specific membrane antigen (PSMA) avidity, stability and in vivo characteristics, as a PET imaging agent for prostate cancer (PCa), was prepared and evaluated. Methods p-Fluorobenzoyl-aminohexanoate and 2-(3-hydroxypropyl)glycine were introduced into the PSMA-targeting scaffold yielding phosphoramidate 5. X-ray crystallography was performed on the PSMA/5 complex. [18F]5 was synthesized, and cell uptake and internalization studies were conducted in PSMA(+) LNCaP and CWR22Rv1 cells and PSMA(−) PC-3 cells. In vivo PET imaging and biodistribution studies were performed at 1 and 4 h post injection in mice bearing CWR22Rv1 tumor, with or without blocking agent. Results The crystallographic data showed interaction of the p-fluorobenzoyl group with an arene-binding cleft on the PSMA surface. In vitro studies revealed elevated uptake of [18F]5 in PSMA(+) cells (2.2% in CWR22Rv1 and 12.1% in LNCaP) compared to PSMA(−) cells (0.08%) at 4 h. In vivo tumor uptake of 2.33% ID/g and tumor-to-blood ratio of 265:1 was observed at 4 h. Conclusions We have successfully synthesized, radiolabeled and evaluated a new PSMA-targeted PET agent. The crystal structure of the PSMA/5 complex highlighted the interactions within the arene-binding cleft contributing to the overall complex stability. The high target uptake and rapid non-target clearance exhibited by [18F]5 in PSMA(+) xenografts substantiates its potential use for PET imaging of PCa. Advances in Knowledge The only FDA-approved imaging agent for PCa, Prostascint®, targets PSMA but suffers from inherent shortcomings. The data acquired in this manuscript confirmed that our new generation of [18F]-labeled PSMA inhibitor exhibited promising in vivo performance as a PET imaging agent for PCa and is well-positioned for subsequent clinical trials. Implications for Patient Care Our preliminary data demonstrate that this tracer possesses

  5. Multiplex PCR Assay Targeting a Diguanylate Cyclase-Encoding Gene, cgcA, To Differentiate Species within the Genus Cronobacter

    PubMed Central

    Carter, L.; Lindsey, L. A.; Grim, C. J.; Sathyamoorthy, V.; Jarvis, K. G.; Gopinath, G.; Lee, C.; Sadowski, J. A.; Trach, L.; Pava-Ripoll, M.; McCardell, B. A.; Tall, B. D.

    2013-01-01

    In a comparison to the widely used Cronobacter rpoB PCR assay, a highly specific multiplexed PCR assay based on cgcA, a diguanylate cyclase gene, that identified all of the targeted six species among 305 Cronobacter isolates was designed. This assay will be a valuable tool for identifying suspected Cronobacter isolates from food-borne investigations. PMID:23144142

  6. Design, synthesis and evaluation of (18)F-labeled bradykinin B1 receptor-targeting small molecules for PET imaging.

    PubMed

    Zhang, Zhengxing; Kuo, Hsiou-Ting; Lau, Joseph; Jenni, Silvia; Zhang, Chengcheng; Zeisler, Jutta; Bénard, François; Lin, Kuo-Shyan

    2016-08-15

    Two fluorine-18 ((18)F) labeled bradykinin B1 receptor (B1R)-targeting small molecules, (18)F-Z02035 and (18)F-Z02165, were synthesized and evaluated for imaging with positron emission tomography (PET). Z02035 and Z02165 were derived from potent antagonists, and showed high binding affinity (0.93±0.44 and 2.80±0.50nM, respectively) to B1R. (18)F-Z02035 and (18)F-Z02165 were prepared by coupling 2-[(18)F]fluoroethyl tosylate with their respective precursors, and were obtained in 10±5 (n=4) and 22±14% (n=3), respectively, decay-corrected radiochemical yield with >99% radiochemical purity. (18)F-Z02035 and (18)F-Z02165 exhibited moderate lipophilicity (LogD7.4=1.10 and 0.59, respectively), and were stable in mouse plasma. PET imaging and biodistribution studies in mice showed that both tracers enabled visualization of the B1R-positive HEK293T::hB1R tumor xenografts with better contrast than control B1R-negative HEK293T tumors. Our data indicate that small molecule antagonists can be used as pharmacophores for the design of B1R-targeting PET tracers. PMID:27390067

  7. FRET- and PET-based sensing in a single material: expanding the dynamic range of an ultra-sensitive nitroaromatic explosives assay.

    PubMed

    Wang, Ying; La, Anthony; Brückner, Christian; Lei, Yu

    2012-10-11

    A polyethylenimine polymer derivatized with pyrene moieties suitable for the fluorescence-based detection of nitroaromatic explosives (NAC) in aqueous systems is described. The system exhibits an exceptionally wide dynamic sensing range of 7 orders of magnitude (from 33 ppt to 225 ppm TNT or Tetryl). This broad range was achieved by the combination of FRET and PET sensing mechanisms in a single material. The sensing material is suitable for a paper strip assay. Simplicity, selectivity, and the wide dynamic range suggest this material for explosives detection in the field. PMID:22935771

  8. A High Throughput Assay for Screening Host Restriction Factors and Antivirals Targeting Influenza A Virus

    PubMed Central

    Wang, Lingyan; Li, Wenjun; Li, Shitao

    2016-01-01

    Influenza A virus (IAV) is a human respiratory pathogen that causes seasonal epidemics and occasional global pandemics with devastating levels of morbidity and mortality. Currently approved treatments against influenza are losing effectiveness, as new viral strains are often refractory to conventional treatments. Thus, there is an urgent need to find new therapeutic targets with which to develop novel antiviral drugs. The common strategy to discover new drug targets and antivirals is high throughput screening. However, most current screenings for IAV rely on the engineered virus carrying a reporter, which prevents the application to newly emerging wild type flu viruses, such as 2009 pandemic H1N1 flu. Here we developed a simple and sensitive screening assay for wild type IAV by quantitatively analyzing viral protein levels using a Dot Blot Assay in combination with the LI-COR Imaging System (DBALIS). We first validated DBALIS in overexpression and RNAi assays, which are suitable methods for screening host factors regulating viral infection. More importantly, we also validated and initiated drug screening using DBALIS. A pilot compound screening identified a small molecule that inhibited IAV infection. Taken together, our method represents a reliable and convenient high throughput assay for screening novel host factors and antiviral compounds. PMID:27375580

  9. A new genotoxicity assay based on p53 target gene induction.

    PubMed

    Zerdoumi, Y; Kasper, E; Soubigou, F; Adriouch, S; Bougeard, G; Frebourg, T; Flaman, J-M

    2015-08-01

    The p53 tumor suppressor protein has emerged as a universal sensor of genotoxic stress that regulates the transcription of numerous genes required for appropriate cellular response to DNA damage. Therefore, transcriptional induction of p53 target genes can be considered as a global and early indicator of genotoxic stress. By performing expression microarrays and RNA-Seq analysis on wild-type and mutant TP53 human lymphocytes respectively derived from controls and Li-Fraumeni patients and exposed to different classes of genotoxic agents, we first determined a common p53-dependent transcriptional signature of DNA damage. We then derived a simple and fast assay based on the exposure of wild-type TP53 lymphocytes to physical or chemical agents and on the quantitative measurement of selected p53 target gene transcriptional induction. The specificity of the p53 genotoxicity assay can easily be demonstrated by performing the same experiment in control lymphocytes with heterozygous TP53 mutations, which compromise responses to DNA damage. This assay allowed us to show that most of the drugs commonly used in cancer treatment, except the microtubule poisons, are highly genotoxic. The p53 genotoxicity assay should facilitate the measurement of the genotoxic effects of chemical and physical agents and the identification of drugs that are not genotoxic and do not expose patients to the risk of secondary malignancies, especially those with a constitutional defect in response to DNA damage, such as patients with Li-Fraumeni syndrome. PMID:26232255

  10. A High Throughput Assay for Screening Host Restriction Factors and Antivirals Targeting Influenza A Virus.

    PubMed

    Wang, Lingyan; Li, Wenjun; Li, Shitao

    2016-01-01

    Influenza A virus (IAV) is a human respiratory pathogen that causes seasonal epidemics and occasional global pandemics with devastating levels of morbidity and mortality. Currently approved treatments against influenza are losing effectiveness, as new viral strains are often refractory to conventional treatments. Thus, there is an urgent need to find new therapeutic targets with which to develop novel antiviral drugs. The common strategy to discover new drug targets and antivirals is high throughput screening. However, most current screenings for IAV rely on the engineered virus carrying a reporter, which prevents the application to newly emerging wild type flu viruses, such as 2009 pandemic H1N1 flu. Here we developed a simple and sensitive screening assay for wild type IAV by quantitatively analyzing viral protein levels using a Dot Blot Assay in combination with the LI-COR Imaging System (DBALIS). We first validated DBALIS in overexpression and RNAi assays, which are suitable methods for screening host factors regulating viral infection. More importantly, we also validated and initiated drug screening using DBALIS. A pilot compound screening identified a small molecule that inhibited IAV infection. Taken together, our method represents a reliable and convenient high throughput assay for screening novel host factors and antiviral compounds. PMID:27375580

  11. CD146-targeted immunoPET and NIRF Imaging of Hepatocellular Carcinoma with a Dual-Labeled Monoclonal Antibody

    PubMed Central

    Hernandez, Reinier; Sun, Haiyan; England, Christopher G.; Valdovinos, Hector F.; Ehlerding, Emily B.; Barnhart, Todd E.; Yang, Yunan; Cai, Weibo

    2016-01-01

    Overexpression of CD146 has been correlated with aggressiveness, recurrence rate, and poor overall survival in hepatocellular carcinoma (HCC) patients. In this study, we set out to develop a CD146-targeting probe for high-contrast noninvasive in vivo positron emission tomography (PET) and near-infrared fluorescence (NIRF) imaging of HCCs. YY146, an anti-CD146 monoclonal antibody, was employed as a targeting molecule to which we conjugated the zwitterionic near-infrared fluorescence (NIRF) dye ZW800-1 and the chelator deferoxamine (Df). This enabled labeling of Df-YY146-ZW800 with 89Zr and its subsequent detection using PET and NIRF imaging, all without compromising antibody binding properties. Two HCC cell lines expressing high (HepG2) and low (Huh7) levels of CD146 were employed to generate subcutaneous (s.c.) and orthotopic xenografts in athymic nude mice. Sequential PET and NIRF imaging performed after intravenous injection of 89Zr-Df-YY146-ZW800 into tumor-bearing mice unveiled prominent and persistent uptake of the tracer in HepG2 tumors that peaked at 31.65 ± 7.15 percentage of injected dose per gram (%ID/g; n=4) 72 h post-injection. Owing to such marked accumulation, tumor delineation was successful by both PET and NIRF, which facilitated the fluorescence image-guided resection of orthotopic HepG2 tumors, despite the relatively high liver background. CD146-negative Huh7 and CD146-blocked HepG2 tumors exhibited significantly lower 89Zr-Df-YY146-ZW800 accretion (6.1 ± 0.5 and 8.1 ± 1.0 %ID/g at 72 h p.i., respectively; n=4), demonstrating the CD146-specificity of the tracer in vivo. Ex vivo biodistribution and immunofluorescent staining corroborated the accuracy of the imaging data and correlated tracer uptake with in situ CD146 expression. Overall, 89Zr-Df-YY146-ZW800 showed excellent properties as a PET/NIRF imaging agent, including high in vivo affinity and specificity for CD146-expressing HCC. CD146-targeted molecular imaging using dual-labeled YY146

  12. CD146-targeted immunoPET and NIRF Imaging of Hepatocellular Carcinoma with a Dual-Labeled Monoclonal Antibody.

    PubMed

    Hernandez, Reinier; Sun, Haiyan; England, Christopher G; Valdovinos, Hector F; Ehlerding, Emily B; Barnhart, Todd E; Yang, Yunan; Cai, Weibo

    2016-01-01

    Overexpression of CD146 has been correlated with aggressiveness, recurrence rate, and poor overall survival in hepatocellular carcinoma (HCC) patients. In this study, we set out to develop a CD146-targeting probe for high-contrast noninvasive in vivo positron emission tomography (PET) and near-infrared fluorescence (NIRF) imaging of HCCs. YY146, an anti-CD146 monoclonal antibody, was employed as a targeting molecule to which we conjugated the zwitterionic near-infrared fluorescence (NIRF) dye ZW800-1 and the chelator deferoxamine (Df). This enabled labeling of Df-YY146-ZW800 with (89)Zr and its subsequent detection using PET and NIRF imaging, all without compromising antibody binding properties. Two HCC cell lines expressing high (HepG2) and low (Huh7) levels of CD146 were employed to generate subcutaneous (s.c.) and orthotopic xenografts in athymic nude mice. Sequential PET and NIRF imaging performed after intravenous injection of (89)Zr-Df-YY146-ZW800 into tumor-bearing mice unveiled prominent and persistent uptake of the tracer in HepG2 tumors that peaked at 31.65 ± 7.15 percentage of injected dose per gram (%ID/g; n=4) 72 h post-injection. Owing to such marked accumulation, tumor delineation was successful by both PET and NIRF, which facilitated the fluorescence image-guided resection of orthotopic HepG2 tumors, despite the relatively high liver background. CD146-negative Huh7 and CD146-blocked HepG2 tumors exhibited significantly lower (89)Zr-Df-YY146-ZW800 accretion (6.1 ± 0.5 and 8.1 ± 1.0 %ID/g at 72 h p.i., respectively; n=4), demonstrating the CD146-specificity of the tracer in vivo. Ex vivo biodistribution and immunofluorescent staining corroborated the accuracy of the imaging data and correlated tracer uptake with in situ CD146 expression. Overall, (89)Zr-Df-YY146-ZW800 showed excellent properties as a PET/NIRF imaging agent, including high in vivo affinity and specificity for CD146-expressing HCC. CD146-targeted molecular imaging using dual

  13. Detection of miRNA Targets in High-throughput Using the 3'LIFE Assay.

    PubMed

    Wolter, Justin M; Kotagama, Kasuen; Babb, Cody S; Mangone, Marco

    2015-01-01

    Luminescent Identification of Functional Elements in 3'UTRs (3'LIFE) allows the rapid identification of targets of specific miRNAs within an array of hundreds of queried 3'UTRs. Target identification is based on the dual-luciferase assay, which detects binding at the mRNA level by measuring translational output, giving a functional readout of miRNA targeting. 3'LIFE uses non-proprietary buffers and reagents, and publically available reporter libraries, making genome-wide screens feasible and cost-effective. 3'LIFE can be performed either in a standard lab setting or scaled up using liquid handling robots and other high-throughput instrumentation. We illustrate the approach using a dataset of human 3'UTRs cloned in 96-well plates, and two test miRNAs, let-7c and miR-10b. We demonstrate how to perform DNA preparation, transfection, cell culture and luciferase assays in 96-well format, and provide tools for data analysis. In conclusion 3'LIFE is highly reproducible, rapid, systematic, and identifies high confidence targets. PMID:26066857

  14. An EGFR Targeted PET Imaging Probe for the Detection of Colonic Adenocarcinomas in the Setting of Colitis

    PubMed Central

    Turker, N. Selcan; Heidari, Pedram; Kucherlapati, Raju; Kucherlapati, Melanie; Mahmood, Umar

    2014-01-01

    Colorectal cancer is a serious complication associated with inflammatory bowel disease, often indistinguishable by screening with conventional FDG PET probes. We have developed an alternative EGFR-targeted PET imaging probe that may be used to overcome this difficulty, and successfully assessed its utility for neoplastic lesion detection in preclinical models. Cetuximab F(ab′)2 fragments were enzymatically generated, purified, and DOTA-conjugated. Radiolabeling was performed with 67Ga for cell based studies and 64Cu for in vivo imaging. Competitive binding studies were performed on CT26 cells to assess affinity (KD) and receptors per cell (Bmax). In vivo imaging using the EGFR targeted PET probe and 18F FDG was performed on CT26 tumor bearing mice in both control and dextran sodium sulfate (DSS) induced colitis settings. Spontaneous adenomas in genetically engineered mouse (GEM) models of colon cancer were additionally imaged. The EGFR imaging agent was generated with high purity (> 98%), with a labeling efficiency of 60 ± 5% and ≥99% radiochemical purity. The KD was 6.6 ± 0.7 nM and the Bmax for CT26 cells was 3.3 ± 0.1 × 106 receptors/cell. Target to background ratios (TBR) for CT26 tumors compared to colonic uptake demonstrated high values for both 18F-FDG (3.95 ± 0.13) and the developed 64Cu-DOTA-cetuximab-F(ab′)2 probe (4.42 ± 0.11) in control mice. The TBR for the EGFR targeted probe remained high (3.78 ± 0.06) in the setting of colitis, while for 18F FDG, this was markedly reduced (1.54 ± 0.08). Assessment of the EGFR targeted probe in the GEM models demonstrated a correlation between radiotracer uptake in spontaneous colonic lesions and the EGFR staining level ex vivo. A clinically translatable PET imaging probe was successfully developed to assess EGFR. The imaging agent can detect colonic tumors with a high TBR for detection of in situ lesions in the setting of colitis, and opens the possibility for a new approach for screening high

  15. Improved target volume definition for fractionated stereotactic radiotherapy in patients with intracranial meningiomas by correlation of CT, MRI, and [{sup 68}Ga]-DOTATOC-PET

    SciTech Connect

    Milker-Zabel, Stefanie . E-mail: stefanie_milker-zabel@med.uni-heidelberg.de; Zabel-du Bois, Angelika; Henze, Marcus; Huber, Peter; Schulz-Ertner, Daniela; Hoess, Angelika; Haberkorn, Uwe; Debus, Juergen

    2006-05-01

    Purpose: To evaluate the influence of {sup 68}-Ga-labeled DOTA ( )-D-Phe ({sup 1})-Tyr ({sup 3})-Octreotide positron emission tomography ([{sup 68}Ga]-DOTATOC-PET) for target definition for fractionated stereotactic radiotherapy (FSRT) as a complementary modality to computed tomography (CT) and magnetic resonance imaging (MRI). Because meningiomas show a high expression of somatostatin receptor subtype 2, somatostatin analogs such as DOTATOC offer the possibility of receptor-targeted imaging. Patients and Methods: Twenty-six patients received stereotactic CT, MRI, and [{sup 68}Ga]-DOTATOC-PET as part of their treatment planning. Histology was: World Health Organization (WHO) Grade 1 61.5%, WHO Grade 2 7.7%, WHO Grade 3 3.9%, and undetermined 26.9%. Six patients received radiotherapy as primary treatment, 2 after subtotal resection; 17 patients were treated for recurrent disease. Dynamic PET scans were acquired before radiotherapy over 60 min after intravenous injection of 156 {+-} 29 MBq [{sup 68}Ga]-DOTATOC. These PET images were imported in the planning software for FSRT. Planning target volume (PTV)-I outlined on CT and contrast-enhanced MRI was compared with PTV-II outlined on PET. PTV-III was defined with CT, MRI, and PET and was actually used for radiotherapy treatment. Results: PTV-III was smaller than PTV-I in 9 patients, the same size in 7 patients, and larger in 10 patients. Median PTV-I was 49.6 cc, median PTV-III was 57.2 cc. In all patients [{sup 68}Ga]-DOTATOC-PET delivered additional information concerning tumor extension. PTV-III was significantly modified based on DOTATOC-PET data in 19 patients. In 1 patient no tumor was exactly identified on CT/MRI but was visible on PET. Conclusion: These data demonstrate that [{sup 68}Ga]-DOTATOC-PET improves target definition for FSRT in patients with intracranial meningiomas. Radiation targeting with fused DOTATOC-PET, CT, and MRI resulted in significant alterations in target definition in 73%.

  16. Targeted RNA Sequencing Assay to Characterize Gene Expression and Genomic Alterations.

    PubMed

    Martin, Dorrelyn P; Miya, Jharna; Reeser, Julie W; Roychowdhury, Sameek

    2016-01-01

    RNA sequencing (RNAseq) is a versatile method that can be utilized to detect and characterize gene expression, mutations, gene fusions, and noncoding RNAs. Standard RNAseq requires 30 - 100 million sequencing reads and can include multiple RNA products such as mRNA and noncoding RNAs. We demonstrate how targeted RNAseq (capture) permits a focused study on selected RNA products using a desktop sequencer. RNAseq capture can characterize unannotated, low, or transiently expressed transcripts that may otherwise be missed using traditional RNAseq methods. Here we describe the extraction of RNA from cell lines, ribosomal RNA depletion, cDNA synthesis, preparation of barcoded libraries, hybridization and capture of targeted transcripts and multiplex sequencing on a desktop sequencer. We also outline the computational analysis pipeline, which includes quality control assessment, alignment, fusion detection, gene expression quantification and identification of single nucleotide variants. This assay allows for targeted transcript sequencing to characterize gene expression, gene fusions, and mutations. PMID:27585245

  17. Development of a multi-target TaqMan assay to detect eastern equine encephalitis virus variants in mosquitoes.

    PubMed

    Armstrong, Philip M; Prince, Nicholanna; Andreadis, Theodore G

    2012-10-01

    Disease outbreaks caused by eastern equine encephalitis virus (EEEV; Togaviridae, Alphavirus) may be prevented by implementing effective surveillance and intervention strategies directed against the mosquito vector. Methods for EEEV detection in mosquitoes include a real-time reverse transcriptase PCR technique (TaqMan assay), but we report its failure to detect variants isolated in Connecticut in 2011, due to a single base-pair mismatch in the probe-binding site. To improve the molecular detection of EEEV, we developed a multi-target TaqMan assay by adding a second primer/probe set to provide redundant targets for EEEV detection. The multi-target TaqMan assay had similar performance characteristics to the conventional assay, but also detected newly-evolving strains of EEEV. The approach described here increases the reliability of the TaqMan assay by creating back-up targets for virus detection without sacrificing sensitivity or specificity. PMID:22835151

  18. Expediting SRM assay development for large-scale targeted proteomics experiments

    SciTech Connect

    Wu, Chaochao; Shi, Tujin; Brown, Joseph N.; He, Jintang; Gao, Yuqian; Fillmore, Thomas L.; Shukla, Anil K.; Moore, Ronald J.; Camp, David G.; Rodland, Karin D.; Qian, Weijun; Liu, Tao; Smith, Richard D.

    2014-10-03

    Due to their high sensitivity and specificity, targeted proteomics measurements, e.g. selected reaction monitoring (SRM), are becoming increasingly popular for biological and translational applications. Selection of optimal transitions and optimization of collision energy (CE) are important assay development steps for achieving sensitive detection and accurate quantification; however, these steps can be labor-intensive, especially for large-scale applications. Herein, we explored several options for accelerating SRM assay development evaluated in the context of a relatively large set of 215 synthetic peptide targets. We first showed that HCD fragmentation is very similar to CID in triple quadrupole (QQQ) instrumentation, and by selection of top six y fragment ions from HCD spectra, >86% of top transitions optimized from direct infusion on QQQ instrument are covered. We also demonstrated that the CE calculated by existing prediction tools was less accurate for +3 precursors, and a significant increase in intensity for transitions could be obtained using a new CE prediction equation constructed from the present experimental data. Overall, our study illustrates the feasibility of expediting the development of larger numbers of high-sensitivity SRM assays through automation of transitions selection and accurate prediction of optimal CE to improve both SRM throughput and measurement quality.

  19. Fluorescence assay for the detection of adherent Candida yeasts to target cells in microtest plates.

    PubMed

    Borg-von Zepelin, M; Wagner, T

    1995-01-01

    We describe an assay based on photometric analysis for the measurement of adherence of Candida species to epithelial target cells (Vero cell line). Adherent Candida cells were detected by staining the cells with the fluorescent dye Calcofluor white (CFW), which binds to chitin and glucan in the yeasts. The tests were performed on microtest plates, which were analysed automatically by fluorescence plate readers. The assay is based on the following steps: (i) coating of the microtest plates with target cells (e.g. Vero cells); (ii) infection with Candida: (iii) staining of Candida with CFW; (iv) rinsing to remove non-adherent Candida cells and unbound dye; (v) detection of adherent fluorescent Candida cells. The test was able to detect 4 x 10(4) cells ml-1. The standard deviation was +/- 8%. Day-to-day variation was +/- 10% at most. The adherence of strains of different Candida species was assayed by a standard procedure. The results confirmed the order of adherence, with C. albicans ranking first, followed by C. tropicalis, C. parapsilosis and C. glabrata. PMID:8569807

  20. Expediting SRM Assay Development for Large-Scale Targeted Proteomics Experiments

    PubMed Central

    2015-01-01

    Because of its high sensitivity and specificity, selected reaction monitoring (SRM)-based targeted proteomics has become increasingly popular for biological and translational applications. Selection of optimal transitions and optimization of collision energy (CE) are important assay development steps for achieving sensitive detection and accurate quantification; however, these steps can be labor-intensive, especially for large-scale applications. Herein, we explored several options for accelerating SRM assay development evaluated in the context of a relatively large set of 215 synthetic peptide targets. We first showed that HCD fragmentation is very similar to that of CID in triple quadrupole (QQQ) instrumentation and that by selection of the top 6 y fragment ions from HCD spectra, >86% of the top transitions optimized from direct infusion with QQQ instrumentation are covered. We also demonstrated that the CE calculated by existing prediction tools was less accurate for 3+ precursors and that a significant increase in intensity for transitions could be obtained using a new CE prediction equation constructed from the present experimental data. Overall, our study illustrated the feasibility of expediting the development of larger numbers of high-sensitivity SRM assays through automation of transition selection and accurate prediction of optimal CE to improve both SRM throughput and measurement quality. PMID:25145539

  1. Expediting SRM assay development for large-scale targeted proteomics experiments

    SciTech Connect

    Wu, Chaochao; Shi, Tujin; Brown, Joseph N.; He, Jintang; Gao, Yuqian; Fillmore, Thomas L.; Shukla, Anil K.; Moore, Ronald J.; Camp, David G.; Rodland, Karin D.; Qian, Weijun; Liu, Tao; Smith, Richard D.

    2014-08-22

    Due to their high sensitivity and specificity, targeted proteomics measurements, e.g. selected reaction monitoring (SRM), are becoming increasingly popular for biological and translational applications. Selection of optimal transitions and optimization of collision energy (CE) are important assay development steps for achieving sensitive detection and accurate quantification; however, these steps can be labor-intensive, especially for large-scale applications. Herein, we explored several options for accelerating SRM assay development evaluated in the context of a relatively large set of 215 synthetic peptide targets. We first showed that HCD fragmentation is very similar to CID in triple quadrupole (QQQ) instrumentation, and by selection of top six y fragment ions from HCD spectra, >86% of top transitions optimized from direct infusion on QQQ instrument are covered. We also demonstrated that the CE calculated by existing prediction tools was less accurate for +3 precursors, and a significant increase in intensity for transitions could be obtained using a new CE prediction equation constructed from the present experimental data. Overall, our study illustrates the feasibility of expediting the development of larger numbers of high-sensitivity SRM assays through automation of transitions selection and accurate prediction of optimal CE to improve both SRM throughput and measurement quality.

  2. Expediting SRM assay development for large-scale targeted proteomics experiments

    DOE PAGESBeta

    Wu, Chaochao; Shi, Tujin; Brown, Joseph N.; He, Jintang; Gao, Yuqian; Fillmore, Thomas L.; Shukla, Anil K.; Moore, Ronald J.; Camp, David G.; Rodland, Karin D.; et al

    2014-08-22

    Due to their high sensitivity and specificity, targeted proteomics measurements, e.g. selected reaction monitoring (SRM), are becoming increasingly popular for biological and translational applications. Selection of optimal transitions and optimization of collision energy (CE) are important assay development steps for achieving sensitive detection and accurate quantification; however, these steps can be labor-intensive, especially for large-scale applications. Herein, we explored several options for accelerating SRM assay development evaluated in the context of a relatively large set of 215 synthetic peptide targets. We first showed that HCD fragmentation is very similar to CID in triple quadrupole (QQQ) instrumentation, and by selection ofmore » top six y fragment ions from HCD spectra, >86% of top transitions optimized from direct infusion on QQQ instrument are covered. We also demonstrated that the CE calculated by existing prediction tools was less accurate for +3 precursors, and a significant increase in intensity for transitions could be obtained using a new CE prediction equation constructed from the present experimental data. Overall, our study illustrates the feasibility of expediting the development of larger numbers of high-sensitivity SRM assays through automation of transitions selection and accurate prediction of optimal CE to improve both SRM throughput and measurement quality.« less

  3. Rapid, targeted and culture-free viral infectivity assay in drop-based microfluidics.

    PubMed

    Tao, Ye; Rotem, Assaf; Zhang, Huidan; Chang, Connie B; Basu, Anindita; Kolawole, Abimbola O; Koehler, Stephan A; Ren, Yukun; Lin, Jeffrey S; Pipas, James M; Feldman, Andrew B; Wobus, Christiane E; Weitz, David A

    2015-10-01

    A key viral property is infectivity, and its accurate measurement is crucial for the understanding of viral evolution, disease and treatment. Currently viral infectivity is measured using plaque assays, which involve prolonged culturing of host cells, and whose measurement is unable to differentiate between specific strains and is prone to low number fluctuation. We developed a rapid, targeted and culture-free infectivity assay using high-throughput drop-based microfluidics. Single infectious viruses are incubated in a large number of picoliter drops with host cells for one viral replication cycle followed by in-drop gene-specific amplification to detect infection events. Using murine noroviruses (MNV) as a model system, we measure their infectivity and determine the efficacy of a neutralizing antibody for different variants of MNV. Our results are comparable to traditional plaque-based assays and plaque reduction neutralization tests. However, the fast, low-cost, highly accurate genomic-based assay promises to be a superior method for drug screening and isolation of resistant viral strains. Moreover our technique can be adapted to measuring the infectivity of other pathogens, such as bacteria and fungi. PMID:26304791

  4. ImmunoPET and biodistribution with human epidermal growth factor receptor 3 targeting antibody 89Zr-RG7116

    PubMed Central

    Terwisscha van Scheltinga, Anton GT; Lub-de Hooge, Marjolijn N; Abiraj, Keelara; Schröder, Carolien P; Pot, Linda; Bossenmaier, Birgit; Thomas, Marlene; Hölzlwimmer, Gabriele; Friess, Thomas; Kosterink, Jos GW; de Vries, Elisabeth GE

    2014-01-01

    The humanized monoclonal antibody with high affinity for the human epidermal growth factor receptor (HER) 3, RG7116, is a glycoengineered, IgG1 class antibody. By labeling RG7116 with zirconium-89 (89Zr) we aimed to visualize in vivo HER3 expression and study the biodistribution of this antibody in human tumor-bearing mice. Biodistribution of 89Zr-RG7116 was studied in subcutaneously xenografted FaDu tumor cells (HER3-positive). Dose-dependency of 89Zr-RG7116 organ distribution and specific tumor uptake was assessed by administering doses ranging from 0.05 to 10 mg/kg RG7116 to SCID/Beige mice. Biodistribution was analyzed at 24 and 144 h after injection. MicroPET imaging was performed at 1, 3, and 6 days after injection of 1.0 mg/kg 89Zr-RG7116 in the FaDu, H441, QG-56 and Calu-1 xenografts with varying HER3 expression. The excised tumors were analyzed for HER3 expression. Biodistribution analyses showed a dose- and time-dependent 89Zr-RG7116 tumor uptake in FaDu tumors. The highest tumor uptake of 89Zr-RG7116 was observed in the 0.05 mg/kg dose group with 27.5%ID/g at 144 h after tracer injection. MicroPET imaging revealed specific tumor uptake of 89Zr-RG7116 in FaDu and H441 models with an increase in tumor uptake over time. Biodistribution data was consistent with the microPET findings in FaDu, H441, QG56 and Calu-1 xenografts, which correlated with HER3 expression levels. In conclusion, 89Zr-RG7116 specifically accumulates in HER3 expressing tumors. PET imaging with this tracer provides real-time non-invasive information about RG7116 distribution, tumor targeting and tumor HER3 expression levels. PMID:24870719

  5. Modified procedure for labelling target cells in a europium release assay of natural killer cell activity.

    PubMed

    Pacifici, R; Di Carlo, S; Bacosi, A; Altieri, I; Pichini, S; Zuccaro, P

    1993-05-01

    Lanthanide europium chelated to diethylenetriaminopentaacetate (EuDTPA) can be used to label target cells such as tumor cells and lymphocytes (Blomberg et al., 1986a,b; Granberg et al., 1988). This procedure has permitted the development of new non-radioactive methods for the detection of target cell cytolysis by natural killer (NK) cells (Blomberg et al., 1986a,b), cytotoxic T lymphocytes (CTL) (Granberg et al., 1988) or complement-mediated cytolysis (Cui et al., 1992). However, we had no success with this method because of a lack of comparability between human NK cell activity simultaneously measured by a classical 51Cr release assay (Seaman et al., 1981) and EuDTPA release assay (Blomberg et al., 1986a). Furthermore, cell division and cell viability were significantly impaired by the suggested concentrations of EuCl3. In this paper, we present a modified non-cytotoxic method for target cell labelling with EuDTPA while cells are growing in culture medium. PMID:8486925

  6. Impact of [18F]-fluoro-ethyl-tyrosine PET imaging on target definition for radiation therapy of high-grade glioma

    PubMed Central

    Munck af Rosenschold, Per; Costa, Junia; Engelholm, Svend Aage; Lundemann, Michael J.; Law, Ian; Ohlhues, Lars; Engelholm, Silke

    2015-01-01

    Background We sought to assess the impact of amino-acid 18F-fluoro-ethyl-tyrosine (FET) positron emission tomography (PET) on the volumetric target definition for radiation therapy of high-grade glioma versus the current standard using MRI alone. Specifically, we investigated the influence of tumor grade, MR-defined tumor volume, and the extent of surgical resection on PET positivity. Methods Fifty-four consecutive high-grade glioma patients (World Health Organization grades III–IV) with confirmed histology were scanned using FET-PET/CT and T1 and T2/fluid attenuated inversion recovery MRI. Gross tumor volume and clinical target volumes (CTVs) were defined in a blinded fashion based on MRI and subsequently PET, and volumetric analysis was performed. The extent of the surgical resection was reviewed using postoperative MRI. Results Overall, for ∼90% of the patients, the PET-positive volumes were encompassed by T1 MRI with contrast-defined tumor plus a 20-mm margin. The tumor volume defined by PET was larger for glioma grade IV (P < .001) and smaller for patients with more extensive surgical resection (P = .004). The margin required to be added to the MRI-defined tumor in order to fully encompass the FET-PET positive volume tended to be larger for grade IV tumors (P = .018). Conclusion With an unchanged CTV margin and by including FET-PET for gross tumor volume definition, the CTV will increase moderately for most patients, and quite substantially for a minority of patients. Patients with grade IV glioma were found to be the primary candidates for PET-guided radiation therapy planning. PMID:25537018

  7. Impact of 18FDG-PET/CT on biological target volume (BTV) definition for treatment planning for non-small cell lung cancer patients

    NASA Astrophysics Data System (ADS)

    Devic, Slobodan; Tomic, Nada; Faria, Sergio; Dean, Geoffrey; Lisbona, Robert; Parker, William; Kaufman, Chris; Podgorsak, Ervin B.

    2007-02-01

    This work represents our effort to test feasibility of FDG-based PET/CT on target volume delineation in radiotherapy treatment planning of NSCLC patients. Different methods have been developed to enable more precise target outlining using PET: Qualitative Visual Method, CTV=2.5 SUV units, linear SUV threshold function method, and CTV=40% Iso of Maximum Uptake Value. We are proposing reconstruction of three biological target volumes: necrotic BTV (same as PTV created by radiation oncologist using CT data), proliferating BTV (based on PET signal to background ratio 1:3) and hypoxic BTV (based on PET signal to background ratio of 1:19). Two IMRT plans were created and compared to the conventional treatment plan: "conservative" IMRT plan delivers 52.5 Gy to the necrotic BTV and 65 Gy to the hypoxic BTV; "radical" IMRT plan delivers 30 Gy to necrotic BTV, 52.5 Gy to proliferating BTV and 65 Gy to hypoxic BTV. Use of BTVs in IMRT plans is attractive because it increases dose to targets considered to need higher doses. It reduces considerably dose to heart and spinal cord, organs considered to limit dose escalation approaches in NSCLC treatment. "Conservative" IMRT approach can be understood as a PET/CT-based concomitant boost to the tumor expressing the highest FDG uptake. "Radical" plan implies deviation from the traditional uniform dose target coverage approach, with the intention of achieving better surrounding tissue sparing and ultimately allowing for dose escalation protocols relying on biologically based treatment planning.

  8. SU-C-BRA-02: Gradient Based Method of Target Delineation On PET/MR Image of Head and Neck Cancer Patients

    SciTech Connect

    Dance, M; Chera, B; Falchook, A; Das, S; Lian, J

    2015-06-15

    Purpose: Validate the consistency of a gradient-based segmentation tool to facilitate accurate delineation of PET/CT-based GTVs in head and neck cancers by comparing against hybrid PET/MR-derived GTV contours. Materials and Methods: A total of 18 head and neck target volumes (10 primary and 8 nodal) were retrospectively contoured using a gradient-based segmentation tool by two observers. Each observer independently contoured each target five times. Inter-observer variability was evaluated via absolute percent differences. Intra-observer variability was examined by percentage uncertainty. All target volumes were also contoured using the SUV percent threshold method. The thresholds were explored case by case so its derived volume matched with the gradient-based volume. Dice similarity coefficients (DSC) were calculated to determine overlap of PET/CT GTVs and PET/MR GTVs. Results: The Levene’s test showed there was no statistically significant difference of the variances between the observer’s gradient-derived contours. However, the absolute difference between the observer’s volumes was 10.83%, with a range from 0.39% up to 42.89%. PET-avid regions with qualitatively non-uniform shapes and intensity levels had a higher absolute percent difference near 25%, while regions with uniform shapes and intensity levels had an absolute percent difference of 2% between observers. The average percentage uncertainty between observers was 4.83% and 7%. As the volume of the gradient-derived contours increased, the SUV threshold percent needed to match the volume decreased. Dice coefficients showed good agreement of the PET/CT and PET/MR GTVs with an average DSC value across all volumes at 0.69. Conclusion: Gradient-based segmentation of PET volume showed good consistency in general but can vary considerably for non-uniform target shapes and intensity levels. PET/CT-derived GTV contours stemming from the gradient-based tool show good agreement with the anatomically and

  9. Development of a Novel PET Tracer [18F]AlF-NOTA-C6 Targeting MMP2 for Tumor Imaging

    PubMed Central

    Cheng, Chao; Zhang, Dazhi; Zhang, Anyu; Wang, Lizhen; Jiang, Hongdie; Wang, Tao; Liu, Hongrui; Xu, Yuping; Yang, Runlin; Chen, Fei; Yang, Min; Zuo, Changjing

    2015-01-01

    Background and Objective The overexpression of gelatinases, that is, matrix metalloproteinase MMP2 and MMP9, has been associated with tumor progression, invasion, and metastasis. To image MMP2 in tumors, we developed a novel ligand termed [18F]AlF-NOTA-C6, with consideration that: c(KAHWGFTLD)NH2 (herein, C6) is a selective gelatinase inhibitor; Cy5.5-C6 has been visualized in many in vivo tumor models; positron emission tomography (PET) has a higher detection sensitivity and a wider field of view than optical imaging; fluorine-18 (18F) is the optimal PET radioisotope, and the creation of a [18F]AlF-peptide complex is a simple procedure. Methods C6 was conjugated to the bifunctional chelator NOTA (1, 4, 7-triazacyclononanetriacetic acid) for radiolabeling [18F]AlF conjugation. The MMP2-binding characteristics and tumor-targeting efficacy of [18F]AlF-NOTA-C6 were tested in vitro and in vivo. Results The non-decay corrected yield of [18F]AlF-NOTA-C6 was 46.2–64.2%, and the radiochemical purity exceeded 95%. [18F]AlF-NOTA-C6 was favorably retained in SKOV3 and PC3 cells, determined by cell uptake. Using NOTA-C6 as a competitive ligand, the uptake of [18F]AlF-NOTA-C6 in SKOV3 cells decreased in a dose-dependent manner. In biodistribution and PET imaging studies, higher radioactivity concentrations were observed in tumors. Pre-injection of C6 caused a marked reduction in tumor tissue uptake. Immunohistochemistry showed MMP2 in tumor tissues. Conclusions [18F]AlF-NOTA-C6 was easy to synthesize and has substantial potential as an imaging agent that targets MMP2 in tumors. PMID:26540114

  10. Bevacizumab Targeting Diffuse Intrinsic Pontine Glioma: Results of 89Zr-Bevacizumab PET Imaging in Brain Tumor Models.

    PubMed

    Jansen, Marc H A; Lagerweij, Tonny; Sewing, A Charlotte P; Vugts, Danielle J; van Vuurden, Dannis G; Molthoff, Carla F M; Caretti, Viola; Veringa, Susanna J E; Petersen, Naomi; Carcaboso, Angel M; Noske, David P; Vandertop, W Peter; Wesseling, Pieter; van Dongen, Guus A M S; Kaspers, Gertjan J L; Hulleman, Esther

    2016-09-01

    The role of the VEGF inhibitor bevacizumab in the treatment of diffuse intrinsic pontine glioma (DIPG) is unclear. We aim to study the biodistribution and uptake of zirconium-89 ((89)Zr)-labeled bevacizumab in DIPG mouse models. Human E98-FM, U251-FM glioma cells, and HSJD-DIPG-007-FLUC primary DIPG cells were injected into the subcutis, pons, or striatum of nude mice. Tumor growth was monitored by bioluminescence imaging (BLI) and visualized by MRI. Seventy-two to 96 hours after (89)Zr-bevacizumab injections, mice were imaged by positron emission tomography (PET), and biodistribution was analyzed ex vivo High VEGF expression in human DIPG was confirmed in a publically available mRNA database, but no significant (89)Zr-bevacizumab uptake could be detected in xenografts located in the pons and striatum at an early or late stage of the disease. E98-FM, and to a lesser extent the U251-FM and HSJD-DIPG-007 subcutaneous tumors, showed high accumulation of (89)Zr-bevacizumab. VEGF expression could not be demonstrated in the intracranial tumors by in situ hybridization (ISH) but was clearly present in the perinecrotic regions of subcutaneous E98-FM tumors. The poor uptake of (89)Zr-bevacizumab in xenografts located in the brain suggests that VEGF targeting with bevacizumab has limited efficacy for diffuse infiltrative parts of glial brain tumors in mice. Translating these results to the clinic would imply that treatment with bevacizumab in patients with DIPG is only justified after targeting of VEGF has been demonstrated by (89)Zr-bevacizumab immuno-PET. We aim to confirm this observation in a clinical PET study with patients with DIPG. Mol Cancer Ther; 15(9); 2166-74. ©2016 AACR. PMID:27325687

  11. Target Volume Delineation in Oropharyngeal Cancer: Impact of PET, MRI, and Physical Examination

    SciTech Connect

    Thiagarajan, Anuradha; Caria, Nicola; Schoeder, Heiko; Iyer, N. Gopalakrishna; Wolden, Suzanne; Wong, Richard J.; Sherman, Eric; Fury, Matthew G.; Lee, Nancy

    2012-05-01

    Introduction: Sole utilization of computed tomography (CT) scans in gross tumor volume (GTV) delineation for head-and-neck cancers is subject to inaccuracies. This study aims to evaluate contributions of magnetic resonance imaging (MRI), positron emission tomography (PET), and physical examination (PE) to GTV delineation in oropharyngeal cancer (OPC). Methods: Forty-one patients with OPC were studied. All underwent contrast-enhanced CT simulation scans (CECTs) that were registered with pretreatment PETs and MRIs. For each patient, three sets of primary and nodal GTV were contoured. First, reference GTVs (GTVref) were contoured by the treating radiation oncologist (RO) using CT, MRI, PET, and PE findings. Additional GTVs were created using fused CT/PET scans (GTVctpet) and CT/MRI scans (GTVctmr) by two other ROs blinded to GTVref. To compare GTVs, concordance indices (CI) were calculated by dividing the respective overlap volumes by overall volumes. To evaluate the contribution of PE, composite GTVs derived from CT, MRI, and PET (GTVctpetmr) were compared with GTVref. Results: For primary tumors, GTVref was significantly larger than GTVctpet and GTVctmr (p < 0.001). Although no significant difference in size was noted between GTVctpet and GTVctmr (p = 0.39), there was poor concordance between them (CI = 0.62). In addition, although CI (ctpetmr vs. ref) was low, it was significantly higher than CI (ctpet vs. ref) and CI (ctmr vs. ref) (p < 0.001), suggesting that neither modality should be used alone. Qualitative analyses to explain the low CI (ctpetmr vs. ref) revealed underestimation of mucosal disease when GTV was contoured without knowledge of PE findings. Similar trends were observed for nodal GTVs. However, CI (ctpet vs. ref), CI (ctmr vs. ref), and CI (ctpetmr vs. ref) were high (>0.75), indicating that although the modalities were complementary, the added benefit was small in the context of CECTs. In addition, PE did not aid greatly in nodal GTV delineation

  12. Yeast-based assay identifies novel Shh/Gli target genes in vertebrate development

    PubMed Central

    2012-01-01

    Background The increasing number of developmental events and molecular mechanisms associated with the Hedgehog (Hh) pathway from Drosophila to vertebrates, suggest that gene regulation is crucial for diverse cellular responses, including target genes not yet described. Although several high-throughput, genome-wide approaches have yielded information at the genomic, transcriptional and proteomic levels, the specificity of Gli binding sites related to direct target gene activation still remain elusive. This study aims to identify novel putative targets of Gli transcription factors through a protein-DNA binding assay using yeast, and validating a subset of targets both in-vitro and in-vivo. Testing in different Hh/Gli gain- and loss-of-function scenarios we here identified known (e.g., ptc1) and novel Hh-regulated genes in zebrafish embryos. Results The combined yeast-based screening and MEME/MAST analysis were able to predict Gli transcription factor binding sites, and position mapping of these sequences upstream or in the first intron of promoters served to identify new putative target genes of Gli regulation. These candidates were validated by qPCR in combination with either the pharmacological Hh/Gli antagonist cyc or the agonist pur in Hh-responsive C3H10T1/2 cells. We also used small-hairpin RNAs against Gli proteins to evaluate targets and confirm specific Gli regulation their expression. Taking advantage of mutants that have been identified affecting different components of the Hh/Gli signaling system in the zebrafish model, we further analyzed specific novel candidates. Studying Hh function with pharmacological inhibition or activation complemented these genetic loss-of-function approaches. We provide evidence that in zebrafish embryos, Hh signaling regulates sfrp2, neo1, and c-myc expression in-vivo. Conclusion A recently described yeast-based screening allowed us to identify new Hh/Gli target genes, functionally important in different contexts of vertebrate

  13. Performance of PCR-based assays targeting Bacteroidales genetic markers of human fecal pollution in sewage and fecal samples

    EPA Science Inventory

    There are numerous PCR-based methods available to characterize human fecal pollution in ambient waters. Each assay employs distinct oligonucleotides and many target different genes and microorganisms leading to potential variations in method performance. Laboratory comparisons ...

  14. Genome Editing-Enabled HTS Assays Expand Drug Target Pathways for Charcot–Marie–Tooth Disease

    PubMed Central

    2015-01-01

    Copy number variation resulting in excess PMP22 protein causes the peripheral neuropathy Charcot–Marie–Tooth disease, type 1A. To broadly interrogate chemically sensitive transcriptional pathways controlling PMP22 protein levels, we used the targeting precision of TALEN-mediated genome editing to embed reporters within the genetic locus harboring the Peripheral Myelin Protein 22 (Pmp22) gene. Using a Schwann cell line with constitutively high endogenous levels of Pmp22, we obtained allelic insertion of secreted bioluminescent reporters with sufficient signal to enable a 1536-well assay. Our findings from the quantitative high-throughput screening (qHTS) of several thousand drugs and clinically investigated compounds using this assay design both overlapped and expanded results from a previous assay using a randomly inserted reporter gene controlled by a single regulatory element of the Pmp22 gene. A key difference was the identification of a kinase-controlled inhibitory pathway of Pmp22 transcription revealed by the activity of the Protein kinase C (PKC)-modulator bryostatin. PMID:25188731

  15. Isothermal target and probe amplification assay for the real-time rapid detection of Staphylococcus aureus.

    PubMed

    Shin, Hyewon; Kim, Minhwan; Yoon, Eunju; Kang, Gyoungwon; Kim, Seungyu; Song, Aelee; Kim, Jeongsoon

    2015-04-01

    Staphylococcus aureus, the species most commonly associated with staphylococcal food poisoning, is one of the most prevalent causes of foodborne disease in Korea and other parts of the world, with much damage inflicted to the health of individuals and economic losses estimated at $120 million. To reduce food poisoning outbreaks by implementing prevention methods, rapid detection of S. aureus in foods is essential. Various types of detection methods for S. aureus are available. Although each method has advantages and disadvantages, high levels of sensitivity and specificity are key aspects of a robust detection method. Here, we describe a novel real-time isothermal target and probe amplification (iTPA) method that allows the rapid and simultaneous amplification of target DNA (the S. aureus nuc gene) and a fluorescence resonance energy transfer-based signal probe under isothermal conditions at 61 °C or detection of S. aureus in real time. The assay was able to specifically detect all 91 S. aureus strains tested without nonspecific detection of 51 non-S. aureus strains. The real-time iTPA assay detected S. aureus at an initial level of 10(1) CFU in overnight cultures of preenriched food samples (kiwi dressing, soybean milk, and custard cream). The advantage of this detection system is that it does not require a thermal cycler, reducing the cost of the real-time PCR and its footprint. Combined with a miniaturized fluorescence detector, this system can be developed into a simplified quantitative hand-held real-time device, which is often required. The iTPA assay was highly reliable and therefore may be used as a rapid and sensitive means of identifying S. aureus in foods. PMID:25836397

  16. Development and evaluation of a quantitative PCR assay targeting sandhill crane (Grus canadensis) fecal pollution.

    PubMed

    Ryu, Hodon; Lu, Jingrang; Vogel, Jason; Elk, Michael; Chávez-Ramírez, Felipe; Ashbolt, Nicholas; Santo Domingo, Jorge

    2012-06-01

    While the microbial water quality in the Platte River is seasonally impacted by excreta from migrating cranes, there are no methods available to study crane fecal contamination. Here we characterized microbial populations in crane feces using phylogenetic analysis of 16S rRNA gene fecal clone libraries. Using these sequences, a novel crane quantitative PCR (Crane1) assay was developed, and its applicability as a microbial source tracking (MST) assay was evaluated by determining its host specificity and detection ability in environmental waters. Bacteria from crane excreta were dominated by bacilli and proteobacteria, with a notable paucity of sequences homologous to Bacteroidetes and Clostridia. The Crane1 marker targeted a dominant clade of unclassified Lactobacillales sequences closely related to Catellicoccus marimammalium. The host distribution of the Crane1 marker was relatively high, being positive for 69% (66/96) of the crane excreta samples tested. The assay also showed high host specificity, with 95% of the nontarget fecal samples (i.e., n = 553; 20 different free-range hosts) being negative. Of the presumed crane-impacted water samples (n = 16), 88% were positive for the Crane1 assay, whereas none of the water samples not impacted by cranes were positive (n = 165). Bayesian statistical models of the Crane1 MST marker demonstrated high confidence in detecting true-positive signals and a low probability of false-negative signals from environmental water samples. Altogether, these data suggest that the newly developed marker could be used in environmental monitoring studies to study crane fecal pollution dynamics. PMID:22492437

  17. Mini-PEG spacering of VAP-1-targeting 68Ga-DOTAVAP-P1 peptide improves PET imaging of inflammation

    PubMed Central

    2011-01-01

    Background Vascular adhesion protein-1 (VAP-1) is an adhesion molecule that plays a key role in recruiting leucocytes into sites of inflammation. We have previously shown that 68Gallium-labelled VAP-1-targeting peptide (68Ga-DOTAVAP-P1) is a positron emission tomography (PET) imaging agent, capable of visualising inflammation in rats, but disadvantaged by its short metabolic half-life and rapid clearance. We hypothesised that prolonging the metabolic half-life of 68Ga-DOTAVAP-P1 could further improve its imaging characteristics. In this study, we evaluated a new analogue of 68Ga-DOTAVAP-P1 modified with a mini-polyethylene glycol (PEG) spacer (68Ga-DOTAVAP-PEG-P1) for in vivo imaging of inflammation. Methods Whole-body distribution kinetics and visualisation of inflammation in a rat model by the peptides 68Ga-DOTAVAP-P1 and 68Ga-DOTAVAP-PEG-P1 were evaluated in vivo by dynamic PET imaging and ex vivo by measuring the radioactivity of excised tissues. In addition, plasma samples were analysed by radio-HPLC for the in vivo stability of the peptides. Results The peptide with the mini-PEG spacer showed slower renal excretion but similar liver uptake as the original peptide. At 60 min after injection, the standardised uptake value of the inflammation site was 0.33 ± 0.07 for 68Ga-DOTAVAP-P1 and 0.53 ± 0.01 for 68Ga-DOTAVAP-PEG-P1 by PET. In addition, inflammation-to-muscle ratios were 6.7 ± 1.3 and 7.3 ± 2.1 for 68Ga-DOTAVAP-P1 and 68Ga-DOTAVAP-PEG-P1, respectively. The proportion of unchanged peptide in circulation at 60 min after injection was significantly higher for 68Ga-DOTAVAP-PEG-P1 (76%) than for 68Ga-DOTAVAP-P1 (19%). Conclusion The eight-carbon mini-PEG spacer prolonged the metabolic half-life of the 68Ga-DOTAVAP-P1 peptide, leading to higher target-to-background ratios and improved in vivo PET imaging of inflammation. PMID:22214508

  18. HER1-Targeted 86Y-Panitumumab Possesses Superior Targeting Characteristics than 86Y-Cetuximab for PET Imaging of Human Malignant Mesothelioma Tumors Xenografts

    PubMed Central

    Nayak, Tapan K.; Garmestani, Kayhan; Milenic, Diane E.; Baidoo, Kwamena E.; Brechbiel, Martin W.

    2011-01-01

    Malignant mesothelioma (MM), a rare form of cancer is often associated with previous exposure to fibrous minerals, such as asbestos. Asbestos exposure increases HER1-activity and expression in pre-clinical models. Additionally, HER1 over-expression is observed in the majority of MM cases. In this study, the utility of HER1-targeted chimeric IgG1, cetuximab, and a human IgG2, panitumumab, radiolabeled with 86Y, were evaluated for PET imaging to detect MM non-invasively in vivo, and to select an antibody candidate for radioimmunotherapy (RIT). Methods Radioimmunoconjugates (RICs) of cetuximab and panitumumab were prepared by conjugation with CHX-A’’-DTPA followed by radiolabeling with 86Y. The HER1 expression of NCI-H226, NCI-H2052, NCI-H2452 and MSTO-211H human mesothelioma cells was characterized by flow cytometry. In vivo biodistribution, pharmacokinetic analysis, and PET imaging were performed in tumor bearing athymic mice. Results In vivo studies demonstrated high HER1 tumor uptake of both RICs. Significant reduction in tumor uptake was observed in mice co-injected with excess mAb (0.1 mg), demonstrating that uptake in the tumor was receptor specific. Significant differences were observed in the in vivo characteristics of the RICs. The blood clearance T½α of 86Y-cetuximab (0.9–1.1 h) was faster than 86Y-panitumumab (2.6–3.1 h). Also, the tumor area under the curve (AUC) to liver AUC ratios of 86Y-panitumumab were 1.5 to 2.5 times greater than 86Y-cetuximab as observed by the differences in PET tumor to background ratios, which could be critical when imaging orthotopic tumors and concerns regarding radiation doses to normal organs such as the liver. Conclusion This study demonstrates the more favorable HER1-targeting characteristics of 86Y-panitumumab than 86Y-cetuximab for non-invasive assessment of the HER1 status of MM by PET imaging. Due to lower liver uptake, panitumumab based immunoconjugates may fare better in therapy than corresponding cetuximab

  19. Irradiation of strontium chloride targets at proton energies above 35 MeV to produce PET radioisotope Y-86

    SciTech Connect

    Medvedev D. G.; Mausner, L.F.; Srivastava, S.C.

    2011-12-01

    Proton irradiation of natural and enriched SrCl{sub 2} targets was used to produce PET radioisotope {sup 86}. The proton energy was degraded from the incident 117.8 MeV to induce the {sup 88}Sr(p,3n) reaction. For the irradiation three pellets made of {sup nat}SrCl{sub 2} (6.61 and 74.49 g) and {sup 88}SrCl{sub 2} (5.02 g) were pressed and individually encapsulated in stainless steel target bodies. The two smaller targets were irradiated for 0.5-1 h at the energy - 46 {yields} 37 MeV to take advantage of the peak in the excitation function of the {sup 88}Sr(p,3n) reaction. The larger target was irradiated at 66.4 {yields} 44.6 MeV. The irradiated pellets were chemically processed to selectively separate {sup 86}Y radioisotope using Eichrom DGA (N,N,N{prime},N{prime}-tetra-n-octyldiglycolamide) resin. The production yields of {sup 86}Y were determined to be 10-13 mCi/{mu}A h. Coproduction of {sup 87m}Y in the final product was 34% for {sup nat}SrCl{sub 2} and 54% for {sup 88}SrCl{sub 2} target. The chemical separation yield of yttrium reached 88-92%. The developed chemical procedure allows for the same day processing and shipment of the isotope to users.

  20. Novel Phakopsora pachyrhizi Extracellular Proteins Are Ideal Targets for Immunological Diagnostic Assays

    PubMed Central

    McMahon, Michael B.; Edwards, H. Herb; Boerma, Britney L.; Lewis Ivey, Melanie L.; Miller, Sally A.; Dorrance, Anne E.

    2012-01-01

    Phakopsora pachyrhizi, the causal agent of Asian soybean rust (ASR), continues to spread across the southeast and midsouth regions of the United States, necessitating the use of fungicides by producers. Our objective in this research was to identify ASR proteins expressed early during infection for the development of immunodiagnostic assays. We have identified and partially characterized a small gene family encoding extracellular proteins in the P. pachyrhizi urediniospore wall, termed PHEPs (for Phakopsora extracellular protein). Two highly expressed protein family members, PHEP 107 and PHEP 369, were selected as ideal immunodiagnostic targets for antibody development, after we detected PHEPs in plants as early as 3 days postinfection (dpi). Monoclonal antibodies (MAbs; 2E8E5-1 and 3G6H7-3) generated against recombinant PHEP 369 were tested for sensitivity against the recombinant protein and extracts from ASR-infected plants and for specificity against a set of common soybean pathogens. These antibodies should prove applicable in immunodiagnostic assays to detect infected soybeans and to identify ASR spores from sentinel surveillance plots. PMID:22447596

  1. Human SRMAtlas: A Resource of Targeted Assays to Quantify the Complete Human Proteome.

    PubMed

    Kusebauch, Ulrike; Campbell, David S; Deutsch, Eric W; Chu, Caroline S; Spicer, Douglas A; Brusniak, Mi-Youn; Slagel, Joseph; Sun, Zhi; Stevens, Jeffrey; Grimes, Barbara; Shteynberg, David; Hoopmann, Michael R; Blattmann, Peter; Ratushny, Alexander V; Rinner, Oliver; Picotti, Paola; Carapito, Christine; Huang, Chung-Ying; Kapousouz, Meghan; Lam, Henry; Tran, Tommy; Demir, Emek; Aitchison, John D; Sander, Chris; Hood, Leroy; Aebersold, Ruedi; Moritz, Robert L

    2016-07-28

    The ability to reliably and reproducibly measure any protein of the human proteome in any tissue or cell type would be transformative for understanding systems-level properties as well as specific pathways in physiology and disease. Here, we describe the generation and verification of a compendium of highly specific assays that enable quantification of 99.7% of the 20,277 annotated human proteins by the widely accessible, sensitive, and robust targeted mass spectrometric method selected reaction monitoring, SRM. This human SRMAtlas provides definitive coordinates that conclusively identify the respective peptide in biological samples. We report data on 166,174 proteotypic peptides providing multiple, independent assays to quantify any human protein and numerous spliced variants, non-synonymous mutations, and post-translational modifications. The data are freely accessible as a resource at http://www.srmatlas.org/, and we demonstrate its utility by examining the network response to inhibition of cholesterol synthesis in liver cells and to docetaxel in prostate cancer lines. PMID:27453469

  2. Comparison of two poultry litter qPCR assays targeting the 16S rRNA gene of Brevibacterium sp.

    PubMed

    Ryu, Hodon; Elk, Michael; Khan, Izhar U H; Harwood, Valerie J; Molina, Marirosa; Edge, Thomas A; Domingo, Jorge Santo

    2014-01-01

    Chicken feces commonly contain human pathogens and are also important sources of fecal pollution in environmental waters. Consequently, methods that can detect chicken fecal pollution are needed in public health and environmental monitoring studies. In this study, we compared a previously developed SYBR green qPCR assay (LA35) to a novel TaqMan qPCR assay (CL) for the environmental detection of poultry-associated fecal pollution. We tested both assays against chicken litter (n = 40), chicken fecal samples (n = 186), non-chicken fecal sources (n = 484), and environmental water samples (n = 323). Most chicken litter samples (i.e., ≥ 98%) were positive for both assays with relatively high signal intensities, whereas only 23% and 12% of poultry fecal samples (n = 186) were positive with the LA35 and the CL assays, respectively. Data using fecal samples from non-target animal species showed that the assays are highly host-associated (≥ 95%). Bayesian statistical models showed that the two assays are associated with relatively low probability of false-positive and false-negative signals in water samples. The CL marker had a lower prevalence than the LA35 assay when tested against environmental water samples (i.e., 21% vs. 31% positive signals). However, by combining the results from the two assays the detection levels increased to 41%, suggesting that using multiple assays can improve the detection of chicken-fecal pollution in environmental waters. PMID:24169514

  3. CRISPR Is an Optimal Target for the Design of Specific PCR Assays for Salmonella enterica Serotypes Typhi and Paratyphi A

    PubMed Central

    Fabre, Laetitia; Le Hello, Simon; Roux, Chrystelle; Issenhuth-Jeanjean, Sylvie; Weill, François-Xavier

    2014-01-01

    Background Serotype-specific PCR assays targeting Salmonella enterica serotypes Typhi and Paratyphi A, the causal agents of typhoid and paratyphoid fevers, are required to accelerate formal diagnosis and to overcome the lack of typing sera and, in some situations, the need for culture. However, the sensitivity and specificity of such assays must be demonstrated on large collections of strains representative of the targeted serotypes and all other bacterial populations producing similar clinical symptoms. Methodology Using a new family of repeated DNA sequences, CRISPR (clustered regularly interspaced short palindromic repeats), as a serotype-specific target, we developed a conventional multiplex PCR assay for the detection and differentiation of serotypes Typhi and Paratyphi A from cultured isolates. We also developed EvaGreen-based real-time singleplex PCR assays with the same two sets of primers. Principal findings We achieved 100% sensitivity and specificity for each protocol after validation of the assays on 188 serotype Typhi and 74 serotype Paratyphi A strains from diverse genetic groups, geographic origins and time periods and on 70 strains of bacteria frequently encountered in bloodstream infections, including 29 other Salmonella serotypes and 42 strains from 38 other bacterial species. Conclusions The performance and convenience of our serotype-specific PCR assays should facilitate the rapid and accurate identification of these two major serotypes in a large range of clinical and public health laboratories with access to PCR technology. These assays were developed for use with DNA from cultured isolates, but with modifications to the assay, the CRISPR targets could be used in the development of assays for use with clinical and other samples. PMID:24498453

  4. Identification of Druggable Targets for Radiation Mitigation Using a Small Interfering RNA Screening Assay

    PubMed Central

    Zellefrow, Crystal D.; Sharlow, Elizabeth R.; Epperly, Michael W.; Reese, Celeste E.; Shun, Tongying; Lira, Ana; Greenberger, Joel S.; Lazo, John S.

    2013-01-01

    Currently, there is a serious absence of pharmaceutically attractive small molecules that mitigate the lethal effects of an accidental or intentional public exposure to toxic doses of ionizing radiation. Moreover, cellular systems that emulate the radiobiologically relevant cell populations and that are suitable for high-throughput screening have not been established. Therefore, we examined two human pluripotent embryonal carcinoma cell lines for use in an unbiased phenotypic small interfering RNA (siRNA) assay to identify proteins with the potential of being drug targets for the protection of human cell populations against clinically relevant ionizing radiation doses that cause acute radiation syndrome. Of the two human cell lines tested, NCCIT cells had optimal growth characteristics in a 384 well format, exhibited radiation sensitivity (D0 = 1.3 ± 0.1 Gy and ñ = 2.0 ± 0.6) comparable to the radiosensitivity of stem cell populations associated with human death within 30 days after total-body irradiation. Moreover, they internalized siRNA after 4 Gy irradiation enabling siRNA library screening. Therefore, we used the human NCCIT cell line for the radiation mitigation study with a siRNA library that silenced 5,520 genes known or hypothesized to be potential therapeutic targets. Exploiting computational methodologies, we identified 113 siRNAs with potential radiomitigative properties, which were further refined to 29 siRNAs with phosphoinositide-3-kinase regulatory subunit 1 (p85α) being among the highest confidence candidate gene products. Colony formation assays revealed radiation mitigation when the phosphoinositide-3-kinase inhibitor LY294002 was given after irradiation of 32D cl 3 cells (D0 = 1.3 ± 0.1 Gy and ñ = 2.3 ± 0.3 for the vehicle control treated cells compared to D0 = 1.2 ± 0.1 Gy and ñ = 6.0 ± 0.8 for the LY294002 treated cells, P = 0.0004). LY294002 and two other PI3K inhibitors, PI 828 and GSK 1059615, also mitigated radiation

  5. Identification of druggable targets for radiation mitigation using a small interfering RNA screening assay.

    PubMed

    Zellefrow, Crystal D; Sharlow, Elizabeth R; Epperly, Michael W; Reese, Celeste E; Shun, Tongying; Lira, Ana; Greenberger, Joel S; Lazo, John S

    2012-09-01

    Currently, there is a serious absence of pharmaceutically attractive small molecules that mitigate the lethal effects of an accidental or intentional public exposure to toxic doses of ionizing radiation. Moreover, cellular systems that emulate the radiobiologically relevant cell populations and that are suitable for high-throughput screening have not been established. Therefore, we examined two human pluripotent embryonal carcinoma cell lines for use in an unbiased phenotypic small interfering RNA (siRNA) assay to identify proteins with the potential of being drug targets for the protection of human cell populations against clinically relevant ionizing radiation doses that cause acute radiation syndrome. Of the two human cell lines tested, NCCIT cells had optimal growth characteristics in a 384 well format, exhibited radiation sensitivity (D(0) = 1.3 ± 0.1 Gy and ñ = 2.0 ± 0.6) comparable to the radiosensitivity of stem cell populations associated with human death within 30 days after total-body irradiation. Moreover, they internalized siRNA after 4 Gy irradiation enabling siRNA library screening. Therefore, we used the human NCCIT cell line for the radiation mitigation study with a siRNA library that silenced 5,520 genes known or hypothesized to be potential therapeutic targets. Exploiting computational methodologies, we identified 113 siRNAs with potential radiomitigative properties, which were further refined to 29 siRNAs with phosphoinositide-3-kinase regulatory subunit 1 (p85α) being among the highest confidence candidate gene products. Colony formation assays revealed radiation mitigation when the phosphoinositide-3-kinase inhibitor LY294002 was given after irradiation of 32D cl 3 cells (D(0) = 1.3 ± 0.1 Gy and ñ = 2.3 ± 0.3 for the vehicle control treated cells compared to D(0) = 1.2 ± 0.1 Gy and ñ = 6.0 ± 0.8 for the LY294002 treated cells, P = 0.0004). LY294002 and two other PI3K inhibitors, PI 828 and GSK 1059615, also mitigated radiation

  6. PET CT Thresholds for Radiotherapy Target Definition in Non-Small-Cell Lung Cancer: How Close are we to the Pathologic Findings?

    SciTech Connect

    Wu Kailiang; Ung, Yee C.; Hornby, Jennifer

    2010-07-01

    Purpose: Optimal target delineation threshold values for positron emission tomography (PET) and computed tomography (CT) radiotherapy planning is controversial. In this present study, different PET CT threshold values were used for target delineation and then compared pathologically. Methods and Materials: A total of 31 non-small-cell lung cancer patients underwent PET CT before surgery. The maximal diameter (MD) of the pathologic primary tumor was obtained. The CT-based gross tumor volumes (GTV{sub CT}) were delineated for CT window-level thresholds at 1,600 and -300 Hounsfield units (HU) (GTV{sub CT1}); 1,600 and -400 (GTV{sub CT2}); 1,600 and -450 HU (GTV{sub CT3}); 1,600 and -600 HU (GTV{sub CT4}); 1,200 and -700 HU (GTV{sub CT5}); 900 and -450 HU (GTV{sub CT6}); and 700 and -450 HU (GTV{sub CT7}). The PET-based GTVs (GTV{sub PET}) were autocontoured at 20% (GTV{sub 20}), 30% (GTV{sub 30}), 40% (GTV{sub 40}), 45% (GTV{sub 45}), 50% (GTV{sub 50}), and 55% (GTV{sub 55}) of the maximal intensity level. The MD of each image-based GTV in three-dimensional orientation was determined. The MD of the GTV{sub PET} and GTV{sub CT} were compared with the pathologically determined MD. Results: The median MD of the GTV{sub CT} changed from 2.89 (GTV{sub CT2}) to 4.46 (GTV{sub CT7}) as the CT thresholds were varied. The correlation coefficient of the GTV{sub CT} compared with the pathologically determined MD ranged from 0.76 to 0.87. The correlation coefficient of the GTV{sub CT1} was the best (r = 0.87). The median MD of GTV{sub PET} changed from 5.72cm to 2.67cm as the PET thresholds increased. The correlation coefficient of the GTV{sub PET} compared with the pathologic finding ranged from 0.51 to 0.77. The correlation coefficient of GTV{sub 50} was the best (r = 0.77). Conclusion: Compared with the MD of GTV{sub PET}, the MD of GTV{sub CT} had better correlation with the pathologic MD. The GTV{sub CT1} and GTV{sub 50} had the best correlation with the pathologic results.

  7. Detection of early stage atherosclerotic plaques using PET and CT fusion imaging targeting P-selectin in low density lipoprotein receptor-deficient mice

    SciTech Connect

    Nakamura, Ikuko; Hasegawa, Koki; Wada, Yasuhiro; Hirase, Tetsuaki; Node, Koichi; Watanabe, Yasuyoshi

    2013-03-29

    Highlights: ► P-selectin regulates leukocyte recruitment as an early stage event of atherogenesis. ► We developed an antibody-based molecular imaging probe targeting P-selectin for PET. ► This is the first report on successful PET imaging for delineation of P-selectin. ► P-selectin is a candidate target for atherosclerotic plaque imaging by clinical PET. -- Abstract: Background: Sensitive detection and qualitative analysis of atherosclerotic plaques are in high demand in cardiovascular clinical settings. The leukocyte–endothelial interaction mediated by an adhesion molecule P-selectin participates in arterial wall inflammation and atherosclerosis. Methods and results: A {sup 64}Cu-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid conjugated anti-P-selectin monoclonal antibody ({sup 64}Cu-DOTA-anti-P-selectin mAb) probe was prepared by conjugating an anti-P-selectin monoclonal antibody with DOTA followed by {sup 64}Cu labeling. Thirty-six hours prior to PET and CT fusion imaging, 3 MBq of {sup 64}Cu-DOTA-anti-P-selectin mAb was intravenously injected into low density lipoprotein receptor-deficient Ldlr-/- mice. After a 180 min PET scan, autoradiography and biodistribution of {sup 64}Cu-DOTA-anti-P-selectin monoclonal antibody was examined using excised aortas. In Ldlr-/- mice fed with a high cholesterol diet for promotion of atherosclerotic plaque development, PET and CT fusion imaging revealed selective and prominent accumulation of the probe in the aortic root. Autoradiography of aortas that demonstrated probe uptake into atherosclerotic plaques was confirmed by Oil red O staining for lipid droplets. In Ldlr-/- mice fed with a chow diet to develop mild atherosclerotic plaques, probe accumulation was barely detectable in the aortic root on PET and CT fusion imaging. Probe biodistribution in aortas was 6.6-fold higher in Ldlr-/- mice fed with a high cholesterol diet than in those fed with a normal chow diet. {sup 64}Cu-DOTA-anti-P-selectin m

  8. Cellular thermal shift and clickable chemical probe assays for the determination of drug-target engagement in live cells.

    PubMed

    Xu, Hua; Gopalsamy, Ariamala; Hett, Erik C; Salter, Shores; Aulabaugh, Ann; Kyne, Robert E; Pierce, Betsy; Jones, Lyn H

    2016-07-14

    Proof of drug-target engagement in physiologically-relevant contexts is a key pillar of successful therapeutic target validation. We developed two orthogonal technologies, the cellular thermal shift assay (CETSA) and a covalent chemical probe reporter approach (harnessing sulfonyl fluoride tyrosine labeling and subsequent click chemistry) to measure the occupancy of the mRNA-decapping scavenger enzyme DcpS by a small molecule inhibitor in live cells. Enzyme affinity determined using isothermal dose response fingerprinting (ITDRFCETSA) and the concentration required to occupy 50% of the enzyme (OC50) using the chemical probe reporter assay were very similar. In this case, the chemical probe method worked well due to the long offset kinetics of the reversible inhibitor (determined using a fluorescent dye-tagged probe). This work suggests that CETSA could become the first choice assay to determine in-cell target engagement due to its simplicity. PMID:27216142

  9. Performance of PCR-based assays targeting Bacteroidales genetic markers of human fecal pollution in sewage and fecal samples.

    PubMed

    Shanks, Orin C; White, Karen; Kelty, Catherine A; Sivaganesan, Mano; Blannon, Janet; Meckes, Mark; Varma, Manju; Haugland, Richard A

    2010-08-15

    There are numerous PCR-based assays available to characterize human fecal pollution in ambient waters. Each assay employs distinct oligonucleotides and many target different genes and microorganisms leading to potential variations in assay performance. Performance comparisons utilizing feces and raw sewage samples are needed to determine which assays are best suited for expensive and time-consuming field validation, fate, transport, and epidemiology studies. We report the assessment of five end-point PCR and 10 real-time quantitative PCR (qPCR) assays that target genes from presumptive Bacteroidales microorganisms reported to be associated with human feces. Each assay was tested against a reference collection of 54 primary influent sewage samples collected from different geographical locations across the United States and 174 fecal DNA extracts from 23 different animal sources. Experiments indicate that human-associated genetic markers are distributed across a broad range of human populations but show substantial differences in specificity for human feces suggesting that particular assays may be more suitable than others depending on the abundance of genetic marker required for detection and the animal sources impacting a particular watershed or beach of interest. PMID:20704227

  10. Comparison of Gull Feces-specific Assays Targeting the 16S rRNA Gene of Catellicoccus Marimammalium and Streptococcus spp.

    EPA Science Inventory

    Two novel gull-specific qPCR assays were developed using 16S rRNA gene sequences from gull fecal clone libraries: a SYBR-green-based assay targeting Streptococcus spp. (i.e., gull3) and a TaqMan qPCR assay targeting Catellicoccus marimammalium (i.e., gull4). The main objectives ...

  11. Contribution of {sup 68}Ga-DOTATOC PET/CT to Target Volume Delineation of Skull Base Meningiomas Treated With Stereotactic Radiation Therapy

    SciTech Connect

    Graf, Reinhold; Nyuyki, Fonyuy; Steffen, Ingo G.; Michel, Roger; Fahdt, Daniel; Wust, Peter; Brenner, Winfried; Budach, Volker; Wurm, Reinhard; Plotkin, Michail

    2013-01-01

    Purpose: To investigate the potential impact of {sup 68}Ga-DOTATOC positron emission tomography ({sup 68}Ga-DOTATOC-PET) in addition to magnetic resonance imaging (MRI) and computed tomography (CT) for retrospectively assessing the gross tumor volume (GTV) delineation of meningiomas of the skull base in patients treated with fractionated stereotactic radiation therapy (FSRT). Methods and Materials: The study population consisted of 48 patients with 54 skull base meningiomas, previously treated with FSRT. After scans were coregistered, the GTVs were first delineated with MRI and CT data (GTV{sub MRI/CT}) and then by PET (GTV{sub PET}) data. The overlapping regions of both datasets resulted in the GTV{sub common}, which was enlarged to the GTV{sub final} by adding volumes defined by only one of the complementary modalities (GTV{sub MRI/CT-added} or GTV{sub PET-added}). We then evaluated the contribution of conventional imaging modalities (MRI, CT) and {sup 68}Ga-DOTATOC-PET to the GTV{sub final}, which was used for planning purposes. Results: Forty-eight of the 54 skull base lesions in 45 patients showed increased {sup 68}Ga-DOTATOC uptake and were further analyzed. The mean GTV{sub MRI/CT} and GTV{sub PET} were approximately 21 cm{sup 3} and 25 cm{sup 3}, with a common volume of approximately 15 cm{sup 3}. PET contributed a mean additional GTV of approximately 1.5 cm{sup 3} to the common volume (16% {+-} 34% of the GTV{sub common}). Approximately 4.5 cm{sup 3} of the GTV{sub MRI/CT} was excluded from the contribution to the common volume. The resulting mean GTV{sub final} was significantly smaller than both the GTV{sub MRI/CT} and the GTV{sub PET}. Compared with the initial GTV{sub MRI/CT}, the addition of {sup 68}Ga-DOTATOC-PET resulted in more than 10% modification of the size of the GTV{sub final} in 32 (67%) meningiomas Conclusions: {sup 68}Ga-DOTATOC-PET/CT seems to improve the target volume delineation in skull base meningiomas, often leading to a reduction of

  12. 68Ga-complex lipophilicity and the targeting property of a urea-based PSMA inhibitor for PET imaging.

    PubMed

    Eder, Matthias; Schäfer, Martin; Bauder-Wüst, Ulrike; Hull, William-Edmund; Wängler, Carmen; Mier, Walter; Haberkorn, Uwe; Eisenhut, Michael

    2012-04-18

    Urea-based inhibitors of the prostate-specific membrane antigen (PSMA) represent low-molecular-weight pepidomimetics showing the ability to image PSMA-expressing prostate tumors. The highly efficient, acyclic Ga(III) chelator N,N'-bis [2-hydroxy-5-(carboxyethyl)benzyl] ethylenediamine-N,N'- diacetic acid (HBED-CC) was introduced as a lipophilic side chain into the hydrophilic pharmacophore Glu-NH-CO-NH-Lys which was found favorable to interact with the PSMA "active binding site". This report describes the syntheses, in vitro binding analyses, and biodistribution data of the radiogallium labeled PSMA inhibitor Glu-NH-CO-NH-Lys(Ahx)-HBED-CC in comparison to the corresponding DOTA conjugate. The binding properties were analyzed using competitive cell binding and enzyme-based assays followed by internalization experiments. Compared to the DOTA-conjugate, the HBED-CC derivative showed reduced unspecific binding and considerable higher specific internalization in LNCaP cells. The (68)Ga complex of the HBED-CC ligand exhibited higher specificity for PSMA expressing tumor cells resulting in improved in vivo properties. (68)Ga labeled Glu-NH-CO-NH-Lys(Ahx)-HBED-CC showed fast blood and organ clearances, low liver accumulation, and high specific uptake in PSMA expressing organs and tumor. It could be demonstrated that the PET-imaging property of a urea-based PSMA inhibitor could significantly be improved with HBED-CC. PMID:22369515

  13. Cytotoxicity, tumor targeting and PET imaging of sub-5 nm KGdF4 multifunctional rare earth nanoparticles

    NASA Astrophysics Data System (ADS)

    Cao, Xinmin; Cao, Fengwen; Xiong, Liqin; Yang, Yang; Cao, Tianye; Cai, Xi; Hai, Wangxi; Li, Biao; Guo, Yixiao; Zhang, Yimin; Li, Fuyou

    2015-08-01

    Ultrasmall sub-5 nm KGdF4 rare earth nanoparticles were synthesized as multifunctional probes for fluorescent, magnetic, and radionuclide imaging. The cytotoxicity of these nanoparticles in human glioblastoma U87MG and human non-small cell lung carcinoma H1299 cells was evaluated, and their application for in vitro and in vivo tumor targeted imaging has also been demonstrated.Ultrasmall sub-5 nm KGdF4 rare earth nanoparticles were synthesized as multifunctional probes for fluorescent, magnetic, and radionuclide imaging. The cytotoxicity of these nanoparticles in human glioblastoma U87MG and human non-small cell lung carcinoma H1299 cells was evaluated, and their application for in vitro and in vivo tumor targeted imaging has also been demonstrated. Electronic supplementary information (ESI) available: Details of the experimental section as well as EDXA, XRD, zeta potential, FTIR, TGA, stability, TEM, Z scanning, ICP-MS, and MicroPET/CT images. See DOI: 10.1039/c5nr03374h

  14. Convergent effects of mouse Pet-1 deletion and human PET-1 variation on amygdala fear and threat processing.

    PubMed

    Wellman, Cara L; Camp, Marguerite; Jones, V Morgan; MacPherson, Kathryn P; Ihne, Jessica; Fitzgerald, Paul; Maroun, Mouna; Drabant, Emily; Bogdan, Ryan; Hariri, Ahmad R; Holmes, Andrew

    2013-12-01

    Serotonin is critical for shaping the development of neural circuits regulating emotion. Pet-1 (FEV-1) is an ETS-domain transcription factor essential for differentiation and forebrain targeting of serotonin neurons. Constitutive Pet-1 knockout (KO) causes major loss of serotonin neurons and forebrain serotonin availability, and behavioral abnormalities. We phenotyped Pet-1 KO mice for fear conditioning and extinction, and on a battery of assays for anxiety- and depression-related behaviors. Morphology of Golgi-stained neurons in basolateral amygdala (BLA) and prelimbic cortex was examined. Using human imaging genetics, a common variant (rs860573) in the PET-1 (FEV) gene was tested for effects on threat-related amygdala reactivity and psychopathology in 88 Asian-ancestry subjects. Pet-1 KO mice exhibited increased acquisition and expression of fear, and elevated fear recovery following extinction, relative to wild-type (WT). BLA dendrites of Pet-1 KO mice were significantly longer than in WT. Human PET-1 variation associated with differences in amygdala threat processing and psychopathology. This novel evidence for the role of Pet-1 in fear processing and dendritic organization of amygdala neurons and in human amygdala threat processing extends a growing literature demonstrating the influence of genetic variation in the serotonin system on emotional regulation via effects on structure and function of underlying corticolimbic circuitry. PMID:24100022

  15. Synthesis and Characterization of αvβ3-Targeting Peptidomimetic Chelate Conjugates for PET and SPECT Imaging

    PubMed Central

    Kim, Young-Seung; Nwe, Kido; Milenic, Diane E.; Brechbiel, Martin W.; Satz, Stanley; Baidoo, Kwamena E.

    2012-01-01

    There is growing interest in small peptidomimetic αvβ3 integrin antagonists that are readily synthesized and characterized and can be easily handled using physiological conditions. Peptidomimetic 4-[2-(3,4,5,6-tetrahydropyrimidine-2-ylamino)ethyloxy]benzoyl-2-[N-(3-amino-neopenta-1-carbamyl)]-aminoethylsulfonyl-amino-β-alanine (IAC) was successfully conjugated to 1-(1-carboxy-3-carbo-t-butoxypropyl)-4,7-(carbo-tert-butoxymethyl)-1,4,7-triazacyclononane (NODAGA(tBu)3) and 1-(1-carboxy-3-carbotertbutoxymethyl)-1,4,7,10-tetraazacyclododecane (DOTAGA(tBu)4) and radiolabeled with 111In, 67Ga and 203Pb. Results of a radioimmunoassay demonstrated binding to purified αvβ3 integrin when one to four equivalents of integrin were added to the reaction. Based on this promising result, investigations are moving forward to evaluate the NODA-GA-IAC and DOTA-GA-IAC conjugates for the targeting tumor associated angiogenesis and αvβ3 integrin positive tumors to define their PET and SPECT imaging qualities as well as their potential for delivery of therapeutic radionuclides. PMID:22853992

  16. Internal Ribosome Entry Site-Based Bicistronic In Situ Reporter Assays for Discovery of Transcription-Targeted Lead Compounds.

    PubMed

    Lang, Liwei; Ding, Han-Fei; Chen, Xiaoguang; Sun, Shi-Yong; Liu, Gang; Yan, Chunhong

    2015-07-23

    Although transgene-based reporter gene assays have been used to discover small molecules targeting expression of cancer-driving genes, the success is limited due to the fact that reporter gene expression regulated by incomplete cis-acting elements and foreign epigenetic environments does not faithfully reproduce chemical responses of endogenous genes. Here, we present an internal ribosome entry site-based strategy for bicistronically co-expressing reporter genes with an endogenous gene in the native gene locus, yielding an in situ reporter assay closely mimicking endogenous gene expression without disintegrating its function. This strategy combines the CRISPR-Cas9-mediated genome-editing tool with the recombinase-mediated cassette-exchange technology, and allows for rapid development of orthogonal assays for excluding false hits generated from primary screens. We validated this strategy by developing a screening platform for identifying compounds targeting oncogenic eIF4E, and demonstrated that the novel reporter assays are powerful in searching for transcription-targeted lead compounds with high confidence. PMID:26144883

  17. Synthesis and Evaluation of [(18)F]RAGER: A First Generation Small-Molecule PET Radioligand Targeting the Receptor for Advanced Glycation Endproducts.

    PubMed

    Cary, Brian P; Brooks, Allen F; Fawaz, Maria V; Drake, Lindsey R; Desmond, Timothy J; Sherman, Phillip; Quesada, Carole A; Scott, Peter J H

    2016-03-16

    The receptor for advanced glycation endproducts (RAGE) is a 35 kDa transmembrane receptor that belongs to the immunoglobulin superfamily of cell surface molecules. Its role in Alzheimer's disease (AD) is complex, but it is thought to mediate influx of circulating amyloid-β into the brain as well as amplify Aβ-induced pathogenic responses. RAGE is therefore of considerable interest as both a diagnostic and a therapeutic target in AD. Herein we report the synthesis and preliminary preclinical evaluation of [(18)F]RAGER, the first small molecule PET radiotracer for RAGE (Kd = 15 nM). Docking studies proposed a likely binding interaction between RAGE and RAGER, [(18)F]RAGER autoradiography showed colocalization with RAGE identified by immunohistochemistry in AD brain samples, and [(18)F]RAGER microPET confirmed CNS penetration and increased uptake in areas of the brain known to express RAGE. This first generation radiotracer represents initial proof-of-concept and a promising first step toward quantifying CNS RAGE activity using PET. However, there were high levels of nonspecific [(18)F]RAGER binding in vitro, likely due to its high log P (experimental log P = 3.5), and rapid metabolism of [(18)F]RAGER in rat liver microsome studies. Therefore, development of second generation ligands with improved imaging properties would be advantageous prior to anticipated translation into clinical PET imaging studies. PMID:26771209

  18. Bacterial Zoonoses Transmitted by Household Pets: State-of-the-Art and Future Perspectives for Targeted Research and Policy Actions.

    PubMed

    Damborg, P; Broens, E M; Chomel, B B; Guenther, S; Pasmans, F; Wagenaar, J A; Weese, J S; Wieler, L H; Windahl, U; Vanrompay, D; Guardabassi, L

    2016-07-01

    The close contact between household pets and people offers favourable conditions for bacterial transmission. In this article, the aetiology, prevalence, transmission, impact on human health and preventative measures are summarized for selected bacterial zoonoses transmissible by household pets. Six zoonoses representing distinct transmission routes were selected arbitrarily based on the available information on incidence and severity of pet-associated disease caused by zoonotic bacteria: bite infections and cat scratch disease (physical injuries), psittacosis (inhalation), leptospirosis (contact with urine), and campylobacteriosis and salmonellosis (faecal-oral ingestion). Antimicrobial resistance was also included due to the recent emergence of multidrug-resistant bacteria of zoonotic potential in dogs and cats. There is a general lack of data on pathogen prevalence in the relevant pet population and on the incidence of human infections attributable to pets. In order to address these gaps in knowledge, and to minimize the risk of human infection, actions at several levels are recommended, including: (1) coordinated surveillance of zoonotic pathogens and antimicrobial resistance in household pets, (2) studies to estimate the burden of human disease attributable to pets and to identify risk behaviours facilitating transmission, and (3) education of those in charge of pets, animal caretakers, veterinarians and human medical healthcare practitioners on the potential zoonotic risks associated with exposure to pets. Disease-specific recommendations include incentives to undertake research aimed at the development of new diagnostic tests, veterinary-specific antimicrobial products and vaccines, as well as initiatives to promote best practices in veterinary diagnostic laboratories and prudent antimicrobial usage. PMID:25958184

  19. [Rectal mucosa metastasis in recurrent prostate cancer : (68)Ga-PSMA-PET/CT allows targeted salvage radiotherapy].

    PubMed

    Düwel, C; Blümel, C; Westenfelder, K; Wagner-Thiessen, E; Becker, A; Gschwend, J E; Eiber, M; Maurer, T

    2016-08-01

    This article presents for the first time a case of rectal mucosa metastasis of recurrent prostate cancer that was diagnosed with (68)Ga-PSMA PET/CT. After histological confirmation, the patient was treated with salvage radiotherapy. This case report underlines the specificity and efficacy of PSMA-based PET imaging. In case of biochemical relapse, it can be used even at low PSA levels to detect prostate cancer metastases that might also be in atypical locations. Thus, (68)Ga-PSMA PET/CT may allow new options for salvage therapy. PMID:27385310

  20. Simultaneous assay of DNA and RNA targets in the whole blood using novel isolation procedure and molecular colony amplification.

    PubMed

    Chetverina, Helena V; Falaleeva, Marina V; Chetverin, Alexander B

    2004-11-15

    A universal procedure that permits the whole human blood to be tested for the presence of single molecules of DNA and RNA targets is described. The procedure includes a novel protocol for the isolation of total nucleic acids from the guanidinium thiocyanate lysate of unfractionated blood in which, prior to phenol/chloroform extraction, the sample is deproteinized by precipitation with isopropanol. The procedure results in a nearly 100% yield of DNA and RNA, preserves the integrity of RNA, and removes any polymerase chain reaction (PCR) inhibitors. Following reverse transcription (RT), target molecules are counted after having been amplified as molecular colonies by carrying out PCR in a polyacrylamide gel. The entire procedure was checked by assaying viral DNA and RNA in 100-microl aliquots of the whole blood and was found to be capable of detecting 100% molecules of DNA target and 50% molecules of RNA target. Unexpectedly, nucleic acids at relatively high concentrations (1 ng/microl) were found to selectively inhibit the RT activity of Thermus thermophilus DNA polymerase without affecting its DNA-dependent polymerization activity. It follows that the popular single-enzyme RT-PCR format, in which this DNA polymerase serves for both RT and PCR, is not appropriate for assaying rare RNA targets. PMID:15494145

  1. Brain-inspired cheminformatics of drug-target brain interactome, synthesis, and assay of TVP1022 derivatives.

    PubMed

    Romero-Durán, Francisco J; Alonso, Nerea; Yañez, Matilde; Caamaño, Olga; García-Mera, Xerardo; González-Díaz, Humberto

    2016-04-01

    The use of Cheminformatics tools is gaining importance in the field of translational research from Medicinal Chemistry to Neuropharmacology. In particular, we need it for the analysis of chemical information on large datasets of bioactive compounds. These compounds form large multi-target complex networks (drug-target interactome network) resulting in a very challenging data analysis problem. Artificial Neural Network (ANN) algorithms may help us predict the interactions of drugs and targets in CNS interactome. In this work, we trained different ANN models able to predict a large number of drug-target interactions. These models predict a dataset of thousands of interactions of central nervous system (CNS) drugs characterized by > 30 different experimental measures in >400 different experimental protocols for >150 molecular and cellular targets present in 11 different organisms (including human). The model was able to classify cases of non-interacting vs. interacting drug-target pairs with satisfactory performance. A second aim focus on two main directions: the synthesis and assay of new derivatives of TVP1022 (S-analogues of rasagiline) and the comparison with other rasagiline derivatives recently reported. Finally, we used the best of our models to predict drug-target interactions for the best new synthesized compound against a large number of CNS protein targets. PMID:26721628

  2. A novel assay for screening inhibitors targeting HIV-1 integrase dimerization based on Ni-NTA magnetic agarose beads

    PubMed Central

    Zhang, Dawei; He, Hongqiu; Liu, Mengmeng; Meng, Zhixia; Guo, Shunxing

    2016-01-01

    Human immunodeficiency virus (HIV)-1 integrase (IN), which mediates integration of viral cDNA into the cellular chromosome, is a validated antiviral drug target. Three IN inhibitors, raltegravir, elvitegravir and dolutegravir, have been clinically approved since 2008. However, drug resistance have emerged in infected patients receiving treatment using these drugs which share the same mechanism of action and have a low genetic barrier for resistance. Therefore, there is an urgent need to develop drugs with novel mechanism. IN requires a precise and dynamic equilibrium between several oligomeric species for its activities. The modulation of the process which is termed as IN oligomerization, presents an interesting allosteric target for drug development. In this research, we developed a magnetic beads based approach to assay the IN dimerization. Then, using the assay we screened a library of 1000 Food and Drug Administration (FDA)-approved drugs for IN dimerization inhibitors and identified dexlansoprazole as a potential IN dimerization inhibitor. In conclusion, the assay presented here has been proven to be sensitive and specific for the detection of IN dimerization as well as for the identification of antiviral drugs targeting IN dimerization. Moreover, a FDA-approved proton-pump inhibitors, dexlansoprazole, was identified as a potential inhibitor for IN dimerization. PMID:27137477

  3. Loop-mediated isothermal amplification (LAMP) assays for detection of Theileria parva infections targeting the PIM and p150 genes.

    PubMed

    Thekisoe, Oriel M M; Rambritch, Natasha E; Nakao, Ryo; Bazie, Raoul S; Mbati, Peter; Namangala, Boniface; Malele, Imna; Skilton, Robert A; Jongejan, Frans; Sugimoto, Chihiro; Kawazu, Shin-Ichiro; Inoue, Noboru

    2010-01-01

    We have developed two loop-mediated isothermal amplification (LAMP) assays for the detection of Theileria parva, the causative agent of East Coast fever (ECF), an economically important cattle disease in eastern, central and southern Africa. These assays target the polymorphic immunodominant molecule (PIM) and p150 LAMP genes. The primer set for each gene target consists of six primers, and each set recognises eight distinct regions on the target gene to give highly specific detection of T. parva. The detection limit of each primer set is 1fg, which is equivalent to one copy of the PIM and p150 T. parva genes. These PIM and p150 LAMP primer sets amplify DNA of T. parva isolates from cattle and buffalo from different countries including Kenya, South Africa, Tanzania, Rwanda, Uganda and Burundi, indicating their ability to detect T. parva from different countries. With the advantages of simplicity, rapidity and cost effectiveness, these LAMP assays are good candidates for molecular epidemiology studies and for monitoring control programs in ECF-endemic, resource poor countries. PMID:19654009

  4. A disk-diffusion-based target identification platform for antibacterials (TIPA): an inducible assay for profiling MOAs of antibacterial compounds.

    PubMed

    Silva, Isba; Real, Lilian J; Ward, Matthew S; Xu, H Howard

    2014-06-01

    One of the challenges in antibiotic lead discovery is the difficulty and time-consuming task of determining the mechanism of action (MOA) of antibacterial compounds. In this report, we describe the development and validation of a facile and inexpensive assay system utilizing disk diffusion of inhibitors on solid agar medium embedded with mixed pools of a comprehensive collection of Escherichia coli clones each containing a plasmid-borne inducible essential gene from E. coli. From individual clones, pilot small-scale (48 or 50 clones) assays, to full-scale target identification platform for antibacterials (TIPA) system, involving a variety of assay formats (liquid vs solid media, individual vs mix clones), we demonstrate that elevated resistance phenotypes of relevant cell clones were highly specific. In particular, the TIPA system was able to reveal cellular targets of several known antibacterial inhibitors: cerulenin, diazaborine, indolmycin, phosphomycin, and triclosan. Complementary to several existing MOA profiling schemes, the TIPA system offers a simple and low-cost method for elucidating the target proteins of antibacterial inhibitors, thus will facilitate discovery and development of novel antibacterial compounds to combat multidrug-resistant bacterial pathogens. PMID:24622888

  5. A novel assay for screening inhibitors targeting HIV-1 integrase dimerization based on Ni-NTA magnetic agarose beads.

    PubMed

    Zhang, Dawei; He, Hongqiu; Liu, Mengmeng; Meng, Zhixia; Guo, Shunxing

    2016-01-01

    Human immunodeficiency virus (HIV)-1 integrase (IN), which mediates integration of viral cDNA into the cellular chromosome, is a validated antiviral drug target. Three IN inhibitors, raltegravir, elvitegravir and dolutegravir, have been clinically approved since 2008. However, drug resistance have emerged in infected patients receiving treatment using these drugs which share the same mechanism of action and have a low genetic barrier for resistance. Therefore, there is an urgent need to develop drugs with novel mechanism. IN requires a precise and dynamic equilibrium between several oligomeric species for its activities. The modulation of the process which is termed as IN oligomerization, presents an interesting allosteric target for drug development. In this research, we developed a magnetic beads based approach to assay the IN dimerization. Then, using the assay we screened a library of 1000 Food and Drug Administration (FDA)-approved drugs for IN dimerization inhibitors and identified dexlansoprazole as a potential IN dimerization inhibitor. In conclusion, the assay presented here has been proven to be sensitive and specific for the detection of IN dimerization as well as for the identification of antiviral drugs targeting IN dimerization. Moreover, a FDA-approved proton-pump inhibitors, dexlansoprazole, was identified as a potential inhibitor for IN dimerization. PMID:27137477

  6. Implementation of an HIV-1 Triple-Target NAT Assay in the Routine Screening at Three German Red Cross Blood Centres

    PubMed Central

    Zolt, Silke De; Thermann, Rolf; Bangsow, Thorsten; Pichl, Lutz; Müller, Benjamin; Jork, Christine; Weber-Schehl, Marijke; Hedges, Doris; Schupp, Ingo; Unverzagt, Patrick; de Rue, Katrin; Roth, W. Kurt

    2016-01-01

    Summary Background Blood product safety was significantly improved by the introduction of NAT testing in the late 1990s, resulting in a strong decrease of transfusion-transmitted infections (TTIs). Due to the occurrence of HIV-1 NAT test failures as a consequence of mismatch mutations in the amplicon regions of mono-target NAT assays, the Paul Ehrlich Institute mandated the implementation of multi-target NAT assays for HIV-1 in 2014. Commercial suppliers mostly developed dual-target NAT assays, with only one implementing a triple-target NAT assay. Methods The HIV-1 triple-target NAT assay v3 (GFE Blut) was tested on mutated specimens and synthetic DNA bearing mutations that resulted in sample underquantification or false-negative test results. In addition, data from 2 years routine testing at three German Red Cross Blood centres were analysed. Results The HIV-1 triple-target PCR could compensate for all mutations tested and could compensate the loss of one amplicon without a significant loss of sensitivity. Data from 2 years routine testing showed a solid performance. Conclusion The HIV-1 triple-target v3 assay (GFE Blut) can compensate mutations in target sequences better than a dual-target assay and is applicable to high-throughput screening, thus increasing blood product safety. PMID:27403090

  7. Establishing reliable production of the PET isotope 89Zr for research use: From target fabrication to preclinical imaging

    NASA Astrophysics Data System (ADS)

    Scharli, R. K.; Price, R. I.; Chan, S.; Cryer, D.; Jeffery, C. M.; Asad, A. H.; Morandeau, L.; Eu, P.; Cullinane, C.; Kasbollah, A.; Katsifis, A.

    2012-12-01

    A semi-automated, in-house external beamline, ≤40 μA at 11.7 MeV for 120 min (degraded from 18 MeV to suppress 88Y & 88Zr co-production) produced 89Zr from 89Y(p,n)89Zr. EOB activity (by HPGe γ-spectr.) of 89Zr in target discs, derived from multiple runs, was 1.42 GBq (±0.45 GBq [SD], n=4) which was 67% (±21%, n=4) of the theoretical activity, with a maximum of 1.84 GBq (87% of theory) achieved. Recovery was 88% (±9%, n=4), radionuclidic purity >99% (n=4) and chemical purity 0.2 ppm Zr (±0.3 ppm, n=3, ICP-MS). The Zr:Y ratio improved from 1:10000 in the pre-filtered solution to 1:10 in the product purified by hydroxamate column. Efficiency of radiolabeling to monoclonal antibody (mAb; trastuzumab) was 100% and purified 89Zr did not bind non-specifically to mAb. Chelator:mAb ratio was 1.3:1. No-carrier-added specific activity of purified 89Zr was 408 MBq/μg (±26 MBq/μg, n=2) via the titration-by-chelator method. Minimum ligand concentration for which 100% labeling occurred was 302 nmol/L. Small animal PET imaging (Philips Mosaic; scan acquisition time 10 min; decay & randoms corrected; image reconstructed using a 3-D RAMLA algorithm) demonstrated marked tumor-specific uptake of 89Zr-labeled mAb but nil 'free' 89Zr (as chloride) tumor uptake.

  8. PET Imaging of CRF1 with [11C]R121920 and [11C]DMP696: Is the Target of Sufficient Density?

    PubMed Central

    Sullivan, Gregory M.; Parsey, Ramin V.; Kumar, J. S. Dileep; Arango, Victoria; Kassir, Suham A.; Huang, Yung‐yu; Simpson, Norman R.; Van Heertum, Ronald L.; Mann, J. John

    2007-01-01

    Aim Overstimulation of the CRF type 1 receptor (CRF1) is implicated in anxiety and depressive disorders. The aim of this study was to investigate the in vivo binding characteristics of [11C]R121920 and [11C]DMP696 in the non‐human primate for application in positron emission tomography (PET) studies of CRF1. Methods PET imaging with the two novel CRF1 radioligands was performed in baboon. In vitro binding studies for CRF1 were performed in postmortem brain tissue of baboon and human to assess sufficiency of receptor density for PET. Results Both [11C]R121920 and [11C]DMP696 distributed rapidly and uniformly throughout brain. Washout was comparable across brain regions, without differences in volume of distribution between regions reported to have high and low in vitro CRF1 binding. Membrane‐enriched tissue homogenate assay using [125I]Tyr0‐sauvagine and specific CRF1 antagonists CP154,526 and SN003 in human occipital cortex yielded maximal binding (Bmax) of 63.3 and 147.3 fmol/mg protein, respectively, and in human cerebellar cortex yielded Bmax of 103.6 and 64.6 fmol/mg protein, respectively. Dissociation constants (KD) were subnanomolar. In baboon, specific binding was not detectable in the same regions; therefore Bmax and KD were not measurable. Autoradiographic results were consistent except there was also detectable CRF1‐specific binding in baboon cerebellum. Conclusion Neither [11C]R121920 nor [11C]DMP696 demonstrated quantifiable regional binding in vivo in baboon. In vitro results suggest CRF1 density in baboon may be insufficient for PET. Studies in man may generate more promising results due to the higher CRF1 density compared with baboon in cerebral cortex and cerebellum. PMID:17499724

  9. Coregistration of Prechemotherapy PET-CT for Planning Pediatric Hodgkin's Disease Radiotherapy Significantly Diminishes Interobserver Variability of Clinical Target Volume Definition

    SciTech Connect

    Metwally, Hussein; Courbon, Frederic; David, Isabelle; Filleron, Thomas; Blouet, Aurelien; Rives, Michel; Izar, Francoise; Zerdoud, Slimane; Plat, Genevieve; Vial, Julie; Robert, Alain; Laprie, Anne

    2011-07-01

    Purpose: To assess the interobserver variability in clinical target volume (CTV) definitions when using registered {sup 18}F-labeled deoxyglucose positron emission tomography (FDG-PET-CT) versus side-by-side image sets in pediatric Hodgkin's disease (HD). Methods and Materials: Prechemotherapy FDG-PET-CT scans performed in the treatment position were acquired from 20 children (median age, 14 years old) with HD (stages 2A to 4B) and registered with postchemotherapy planning CT scans. The patients had a median age of 14 years and stages of disease ranging between 2A and 4B. Image sets were coregistered using a semiautomatic coregistration system. The biological target volume was defined on all the coregistered images as a guide to defining the initial site of involvement and to avoid false-positive or negative results. Five radiation oncologists independently defined the CTV for all 20 patients: once using separate FDG-PET-CT images as a guide (not registered) to define CTVa and once using the registered FDG-PET-CT data to define CTVb. The total volumes were compared, as well as their coefficients of variation (COV). To assess the interobserver variability, the percentages of intersection between contours drawn by all observers for each patient were calculated for CTVa and for CTVb. Results: The registration of a prechemotherapy FDG-PET-CT scan caused a change in the CTV for all patients. Comparing CTVa with CTVb showed that the mean CTVb increased in 14 patients (range, 0.61%-101.96%) and decreased in 6 patients (range, 2.97%-37.26%). The COV for CTVb significantly decreased for each patient; the mean COVs for CTVa and CTVb were 45% (21%-65%) and 32% (13%-57%), respectively (p = 0.0004). The percentage of intersection among all CTVbs for the five observers increased significantly by 89.77% (1.99%-256.41%) compared to that of CTVa (p = 0.0001). Conclusions: High observer variability can occur during CT-based definition of CTVs for children diagnosed with HD

  10. A Prospective Evaluation of Staging and Target Volume Definition of Lymph Nodes by {sup 18}FDG PET/CT in Patients With Squamous Cell Carcinoma of Thoracic Esophagus

    SciTech Connect

    Yu Wen; Fu Xiaolong; Zhang Yingjian; Xiang Jiaqing; Shen Lei; Chang, Joe Y.

    2011-12-01

    Purpose: To determine an optimal standardized uptake value (SUV) threshold for detecting lymph node (LN) metastases in esophageal cancer using {sup 18}F-Fluorodeoxyglucose positron emission tomography/computer tomography ({sup 18}FDG PET/CT) and to define the resulting nodal target volume, using histopathology as a 'gold standard.' Methods: Sixteen patients with esophageal squamous cell carcinoma who underwent radical esophagectomy and three-field LN dissection after {sup 18}FDG PET/CT and CT scans were enrolled into this study. Locations of LN groups were recorded according to a uniform LN map. Diagnostic performance of different SUV thresholds was assessed by receiver operating characteristic analysis. The optimal cutoff SUV was determined by plotting the false-negative rate (FNR) and false-positive rate (FPR), the sum of both error rates (FNR+FPR), and accuracy against a hypothetical SUV threshold. For each patient, nodal gross tumor volumes (GTVNs) were generated with CT alone (GTVNCT), PET/CT (GTVNPET), and pathologic data (GTVNpath). GTVNCT or GTVNPET was compared with GTVNpath by means of a conformity index (CI), which is the intersection of the two GTVNs divided by the sum of them minus the intersection, e.g., CI{sub CT} and {sub path} = GTVN{sub CT} and {sub path}/(GTVN{sub CT}+ GTVN{sub path} - GTVN{sub CT} and {sub path}). Results: LN metastases occurred in 21 LN groups among the 144 specimens taken from the 16 patients. The area under the receiver operating characteristic curve was 0.9017 {+-} 0.0410. The plot of error rates showed a minimum of FNR+FPR for an SUV of 2.36, at which the sensitivity, specificity, and accuracy were 76.19%, 95.93%, and 93.06%, respectively, whereas those of CT were 33.33%, 94.31%, and 85.42% (p values: 0.0117, 0.7539, and 0.0266). Mean GTVN{sub CT}, GTVN{sub PET}, and GTVN{sub path} were 1.52 {+-} 2.38, 2.82 {+-} 4.51, and 2.68 {+-} 4.16cm{sup 3}, respectively. Mean CI{sub CT} and {sub path} and CI{sub PET} and {sub path

  11. [Oncology PET imaging].

    PubMed

    Inubushi, Masayuki

    2014-01-01

    At the beginning of this article, likening medical images to "Where is Waldo?" I indicate the concept of diagnostic process of PET/CT imaging, so that medical physics specialists could understand the role of each imaging modality and infer our distress for image diagnosis. Then, I state the present situation of PET imaging and the basics (e.g. health insurance coverage, clinical significance, principle, protocol, and pitfall) of oncology FDG-PET imaging which accounts for more than 99% of all clinical PET examinations in Japan. Finally, I would like to give a wishful prospect of oncology PET that will expand to be more cancer-specific in order to assess therapeutic effects of emerging molecular targeted drugs targeting the "hallmarks of cancer". PMID:25199271

  12. [Rapid detection of Pseudomonas aeruginosa by the fluorescence quantitative PCR assay targeting 16S rDNA].

    PubMed

    Xue, Li-Jun; Wang, Yong-Zhi; Ren, Hao; Tong, Yi-Min; Zhao, Ping; Zhu, Shi-Ying; Qi, Zhong-Tian

    2006-09-01

    The 16S rDNA specific primers were designed for rapid detection of Pseudomonas aeruginosa (PA) by the fluorescence quantitative PCR (FQ-PCR) assay, based upon multiple sequence alignment and phylogenetic tree analysis of the 16S rDNAs of over 20 bacteria. After extraction of PA genomic DNA, the target 16S rDNA fragment was amplified by PCR with specific primers, and used to construct recombinant pMDT-Pfr plasmid, the dilution gradients of which were subjected to the standard quantitation curve in FQ-PCR assay. Different concentrations of PA genomic DNA were detected by FQ-PCR in a 20microL of reaction system with SYBR Green I. At the same time, various genomic DNAs of Staphylococcus aureus, Salmonella typhi, Shigella flexneri, Proteus vulgaris, Staphylococcus epidermidis, Escherichia coli, and Mycobacterium tuberculosis were used as negative controls to confirm specificity of the FQ-PCR detection assay. Results demonstrated that the predicted amplified product of designed primers was of high homology only with PA 16S rDNA, and that sensitivity of the FQ-PCR assay was of 3.6pg/microL of bacterial DNA or (2.1 x 10(3) +/- 3.1 x 10(2)) copies/microL of 16S rDNA, accompanied with high specificity, and that the whole detection process including DNA extraction could be completed in about two hours. In contrast to traditional culture method, the FQ-PCR assay targeting 16S rDNA gene can be used to detect PA rapidly, which exhibits perfect application prospect in future. PMID:17037203

  13. A Novel SERRS Sandwich-Hybridization Assay to Detect Specific DNA Target

    PubMed Central

    Gillet, Benjamin; Montagnac, Gilles; Daniel, Isabelle; Hänni, Catherine

    2011-01-01

    In this study, we have applied Surface Enhanced Resonance Raman Scattering (SERRS) technology to the specific detection of DNA. We present an innovative SERRS sandwich-hybridization assay that allows specific DNA detection without any enzymatic amplification, such as is the case with Polymerase Chain Reaction (PCR). In some substrates, such as ancient or processed remains, enzymatic amplification fails due to DNA alteration (degradation, chemical modification) or to the presence of inhibitors. Consequently, the development of a non-enzymatic method, allowing specific DNA detection, could avoid long, expensive and inconclusive amplification trials. Here, we report the proof of concept of a SERRS sandwich-hybridization assay that leads to the detection of a specific chamois DNA. This SERRS assay reveals its potential as a non-enzymatic alternative technology to DNA amplification methods (particularly the PCR method) with several applications for species detection. As the amount and type of damage highly depend on the preservation conditions, the present SERRS assay would enlarge the range of samples suitable for DNA analysis and ultimately would provide exciting new opportunities for the investigation of ancient DNA in the fields of evolutionary biology and molecular ecology, and of altered DNA in food frauds detection and forensics. PMID:21655320

  14. Assay for peptidoglycan O-acetyltransferase: a potential new antibacterial target.

    PubMed

    Moynihan, Patrick J; Clarke, Anthony J

    2013-08-15

    The O-acetylation of peptidoglycan occurs at the C-6 hydroxyl group of muramoyl residues in many human pathogens, both gram positive and gram negative, such as Staphylococcus aureus and species of Campylobacter, Helicobacter, Neisseria, and Bacillus, including Bacillus anthracis. The process is a maturation event being catalyzed either by integral membrane O-acetylpeptidoglycan transferase (Oat) of gram-positive bacteria or by a two-component peptidoglycan O-acetyltransferase system (PatA/PatB) in gram-negative cells. Here, we describe the development of the first in vitro assay for any peptidoglycan O-acetyltransferase using PatB from Neisseria gonorrhoeae as the model enzyme. This assay is based on the use of chromogenic p-nitrophenyl acetate as the donor substrate and chitooligosaccharides as model acceptor substrates in place of peptidoglycan. The identity of the O-acetylated chitooligosaccharides was confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Rates of transacetylations were determined spectrophotometrically by monitoring p-nitrophenol release after accounting for both spontaneous and enzyme-catalyzed hydrolysis of the acetate donor. Conditions were established for use of the assay in microtiter plate format, and its applicability was demonstrated by determining the first Michaelis-Menten kinetic parameters for PatB. The assay is readily amenable for application in the high-throughput screening for potential inhibitors of peptidoglycan O-acetyltransferases that may prove to be leads for novel classes of antibiotics. PMID:23660013

  15. Identifying New Drug Targets for Potent Phospholipase D Inhibitors: Combining Sequence Alignment, Molecular Docking, and Enzyme Activity/Binding Assays.

    PubMed

    Djakpa, Helene; Kulkarni, Aditya; Barrows-Murphy, Scheneque; Miller, Greg; Zhou, Weihong; Cho, Hyejin; Török, Béla; Stieglitz, Kimberly

    2016-05-01

    Phospholipase D enzymes cleave phospholipid substrates generating choline and phosphatidic acid. Phospholipase D from Streptomyces chromofuscus is a non-HKD (histidine, lysine, and aspartic acid) phospholipase D as the enzyme is more similar to members of the diverse family of metallo-phosphodiesterase/phosphatase enzymes than phospholipase D enzymes with active site HKD repeats. A highly efficient library of phospholipase D inhibitors based on 1,3-disubstituted-4-amino-pyrazolopyrimidine core structure was utilized to evaluate the inhibition of purified S. chromofuscus phospholipase D. The molecules exhibited inhibition of phospholipase D activity (IC50 ) in the nanomolar range with monomeric substrate diC4 PC and micromolar range with phospholipid micelles and vesicles. Binding studies with vesicle substrate and phospholipase D strongly indicate that these inhibitors directly block enzyme vesicle binding. Following these compelling results as a starting point, sequence searches and alignments with S. chromofuscus phospholipase D have identified potential new drug targets. Using AutoDock, inhibitors were docked into the enzymes selected from sequence searches and alignments (when 3D co-ordinates were available) and results analyzed to develop next-generation inhibitors for new targets. In vitro enzyme activity assays with several human phosphatases demonstrated that the predictive protocol was accurate. The strategy of combining sequence comparison, docking, and high-throughput screening assays has helped to identify new drug targets and provided some insight into how to make potential inhibitors more specific to desired targets. PMID:26691755

  16. Detection of miRNA Targets in High-throughput Using the 3′LIFE Assay

    PubMed Central

    Wolter, Justin M.; Kotagama, Kasuen; Babb, Cody S.; Mangone, Marco

    2015-01-01

    Luminescent Identification of Functional Elements in 3′UTRs (3′LIFE) allows the rapid identification of targets of specific miRNAs within an array of hundreds of queried 3′UTRs. Target identification is based on the dual-luciferase assay, which detects binding at the mRNA level by measuring translational output, giving a functional readout of miRNA targeting. 3′LIFE uses non-proprietary buffers and reagents, and publically available reporter libraries, making genome-wide screens feasible and cost-effective. 3′LIFE can be performed either in a standard lab setting or scaled up using liquid handling robots and other high-throughput instrumentation. We illustrate the approach using a dataset of human 3′UTRs cloned in 96-well plates, and two test miRNAs, let-7c and miR-10b. We demonstrate how to perform DNA preparation, transfection, cell culture and luciferase assays in 96-well format, and provide tools for data analysis. In conclusion 3′LIFE is highly reproducible, rapid, systematic, and identifies high confidence targets. PMID:26066857

  17. Quenching methods for background reduction in luminescence-based probe-target binding assays

    DOEpatents

    Cai, Hong; Goodwin, Peter M; Keller, Richard A.; Nolan, Rhiannon L.

    2007-04-10

    Background luminescence is reduced from a solution containing unbound luminescent probes, each having a first molecule that attaches to a target molecule and having an attached luminescent moiety, and luminescent probe/target adducts. Quenching capture reagent molecules are formed that are capable of forming an adduct with the unbound luminescent probes and having an attached quencher material effective to quench luminescence of the luminescent moiety. The quencher material of the capture reagent molecules is added to a solution of the luminescent probe/target adducts and binds in a proximity to the luminescent moiety of the unbound luminescent probes to quench luminescence from the luminescent moiety when the luminescent moiety is exposed to exciting illumination. The quencher capture reagent does not bind to probe molecules that are bound to target molecules and the probe/target adduct emission is not quenched.

  18. Off-Target Effects of Psychoactive Drugs Revealed by Genome-Wide Assays in Yeast

    PubMed Central

    Ericson, Elke; Gebbia, Marinella; Heisler, Lawrence E.; Wildenhain, Jan; Tyers, Mike; Giaever, Guri; Nislow, Corey

    2008-01-01

    To better understand off-target effects of widely prescribed psychoactive drugs, we performed a comprehensive series of chemogenomic screens using the budding yeast Saccharomyces cerevisiae as a model system. Because the known human targets of these drugs do not exist in yeast, we could employ the yeast gene deletion collections and parallel fitness profiling to explore potential off-target effects in a genome-wide manner. Among 214 tested, documented psychoactive drugs, we identified 81 compounds that inhibited wild-type yeast growth and were thus selected for genome-wide fitness profiling. Many of these drugs had a propensity to affect multiple cellular functions. The sensitivity profiles of half of the analyzed drugs were enriched for core cellular processes such as secretion, protein folding, RNA processing, and chromatin structure. Interestingly, fluoxetine (Prozac) interfered with establishment of cell polarity, cyproheptadine (Periactin) targeted essential genes with chromatin-remodeling roles, while paroxetine (Paxil) interfered with essential RNA metabolism genes, suggesting potential secondary drug targets. We also found that the more recently developed atypical antipsychotic clozapine (Clozaril) had no fewer off-target effects in yeast than the typical antipsychotics haloperidol (Haldol) and pimozide (Orap). Our results suggest that model organism pharmacogenetic studies provide a rational foundation for understanding the off-target effects of clinically important psychoactive agents and suggest a rational means both for devising compound derivatives with fewer side effects and for tailoring drug treatment to individual patient genotypes. PMID:18688276

  19. The screening of everyday life chemicals in validated assays targeting the pituitary-gonadal axis.

    PubMed

    Tinwell, H; Colombel, S; Blanck, O; Bars, R

    2013-07-01

    Ten structurally diverse chemicals (vitamins C, B9, B6, B3, sucrose, caffeine, gingerol, xanthan gum, paracetamol, ibuprofen) deemed intrinsic to modern life but not considered as endocrine active, were tested in vitro using the human estrogen receptor transcriptional activation (hERTa) and the H295R steroidogenesis assays. All were inactive in the hERTa assay but paracetamol, gingerol, caffeine and vitamin C affected steroidogenesis in vitro from 250, 25, 500 and 750 μM respectively. One molecule, caffeine, was further tested in rat pubertal assays at the tumorigenic dose-level and at dose-levels relevant for human consumption. In females pubertal parameters (vaginal opening, estrus cycle), ovarian weight and Fsh and prolactin transcript levels were affected. In males, plasma progesterone levels and prostate and seminal vesicle weights were affected. Although the current regulatory focus is synthetic chemicals that can cause adverse effects on the hypothalamus-pituitary-gonadal axis, our data infer that the range of natural chemicals with the potential to affect this axis may be extensive and is probably overlooked. Thus, to avoid regulation of an overwhelming number of chemicals, a weight of evidence approach, combining hazard identification and characterization with exposure considerations, is needed to identify those chemicals of real regulatory concern. PMID:23590819

  20. A short target real-time RT-PCR assay for detection of pestiviruses infecting cattle.

    PubMed

    La Rocca, S A; Sandvik, T

    2009-10-01

    A rapid single step real-time duplex TaqMan RT-PCR was developed for detection of bovine viral diarrhoea virus (BVDV)-1, BVDV-2 and border disease virus (BDV). Based on alignment of available and newly generated partial 5'-UTR nucleotide sequences, one forward and two reverse primers were designed, which amplify a 104bp PCR product. Two TaqMan probes labelled with different fluorochromes were designed to detect BVDV-1/BVDV-2 and BDVs, respectively. The assay was able to detect a selection of strains and isolates that represent the genetic diversity of these three viruses, with an analytical sensitivity that corresponded to 3.6, 48 and 4.8 TCID(50) of BVDV-1, BVDV-2 and BDV, respectively. With an overall cycling time of around 70 min, the assay allows rapid diagnosis and efficient use of modern thermocycling machines. Although developed principally for the diagnosis of BVD, the assay should be equally useful for diagnosis of BD in sheep. PMID:19523981

  1. 3′LIFE: a functional assay to detect miRNA targets in high-throughput

    PubMed Central

    Wolter, Justin M.; Kotagama, Kasuen; Pierre-Bez, Alexandra C.; Firago, Mari; Mangone, Marco

    2014-01-01

    MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene output at the post-transcriptional level by targeting degenerate elements primarily in 3′untranslated regions (3′UTRs) of mRNAs. Individual miRNAs can regulate networks of hundreds of genes, yet for the majority of miRNAs few, if any, targets are known. Misexpression of miRNAs is also a major contributor to cancer progression, thus there is a critical need to validate miRNA targets in high-throughput to understand miRNAs' contribution to tumorigenesis. Here we introduce a novel high-throughput assay to detect miRNA targets in 3′UTRs, called Luminescent Identification of Functional Elements in 3′UTRs (3′LIFE). We demonstrate the feasibility of 3′LIFE using a data set of 275 human 3′UTRs and two cancer-relevant miRNAs, let-7c and miR-10b, and compare our results to alternative methods to detect miRNA targets throughout the genome. We identify a large number of novel gene targets for these miRNAs, with only 32% of hits being bioinformatically predicted and 27% directed by non-canonical interactions. Functional analysis of target genes reveals consistent roles for each miRNA as either a tumor suppressor (let-7c) or oncogenic miRNA (miR-10b), and preferentially target multiple genes within regulatory networks, suggesting 3′LIFE is a rapid and sensitive method to detect miRNA targets in high-throughput. PMID:25074381

  2. Application of vitamin B(12)-targeting site on Lactobacillus helveticus B-1 to vitamin B(12) assay by chemiluminescence method.

    PubMed

    Sato, Kazuyoshi; Muramatsu, Kumi; Amano, Setsumi

    2002-09-01

    Lactobacillus helveticus B-1 is assumed to have a vitamin B(12)-targeting (or B(12)-binding) site on the cells, since the binding reaction of vitamin B(12) with L. helveticus B-1 cells proceeded instantly and quantitatively. This reaction is specific to complete B(12) compounds, cobalamins, and can be used for a vitamin B(12) assay method by chemiluminescence. The calibration graph was linear from 0.1 to 10.0 ng/mL. The B(12) contents in oyster and sardine were 75.9 and 39.4 microg/100g, respectively. These values were very close to those obtained using a chemilumi-ADVIA Centaur immunoassay system with intrinsic factor and to those obtained by microbiological assays. PMID:12234457

  3. Automated Microchromatography Enables Multiplexing of Immunoaffinity Enrichment of Peptides to Greater than 150 for Targeted MS-Based Assays.

    PubMed

    Ippoliti, Paul J; Kuhn, Eric; Mani, D R; Fagbami, Lola; Keshishian, Hasmik; Burgess, Michael W; Jaffe, Jacob D; Carr, Steven A

    2016-08-01

    Immunoaffinity enrichment of peptides coupled with analysis by stable isotope dilution multiple reaction mass spectrometry has been shown to have analytical performance and detection limits suitable for many biomarker verification studies and biological applications. Prior studies have shown that antipeptide antibodies can be multiplexed up to 50 in a single assay without significant loss of performance. Achieving higher multiplex levels is relevant to all studies involving precious biological material as this minimizes the amount of sample that must be consumed to measure a given set of analytes and reduces the assay cost per analyte. Here we developed automated methods employing the Agilent AssayMAP Bravo microchromatography platform and used these methods to characterize the performance of immunoaffinity enrichment of peptides up to multiplex levels of 172. Median capture efficiency for the target peptides remained high (88%) even at levels of 150-plex and declined to 70% at 172-plex compared to antibody performance observed at standard lower multiplex levels (n = 25). Subsequently, we developed and analytically characterized a multiplexed immuno-multiple reaction monitoring-mass spectrometry (immuno-MRM-MS) assay (n = 110) and applied it to measure candidate protein biomarkers of cardiovascular disease in plasma of patients undergoing planned myocardial infarction. The median lower limit of detection of all peptides was 71.5 amol/μL (nM), and the coefficient of variation (CV) was less than 15% at the lower limit of quantification. The results demonstrate that high multiplexed immuno-MRM-MS assays are readily achievable using the optimized sample processing and peptide capture methods described here. PMID:27321643

  4. Integrated-boost IMRT or 3-D-CRT using FET-PET based auto-contoured target volume delineation for glioblastoma multiforme - a dosimetric comparison

    PubMed Central

    2009-01-01

    Background Biological brain tumor imaging using O-(2-[18F]fluoroethyl)-L-tyrosine (FET)-PET combined with inverse treatment planning for locally restricted dose escalation in patients with glioblastoma multiforme seems to be a promising approach. The aim of this study was to compare inverse with forward treatment planning for an integrated boost dose application in patients suffering from a glioblastoma multiforme, while biological target volumes are based on FET-PET and MRI data sets. Methods In 16 glioblastoma patients an intensity-modulated radiotherapy technique comprising an integrated boost (IB-IMRT) and a 3-dimensional conventional radiotherapy (3D-CRT) technique were generated for dosimetric comparison. FET-PET, MRI and treatment planning CT (P-CT) were co-registrated. The integrated boost volume (PTV1) was auto-contoured using a cut-off tumor-to-brain ratio (TBR) of ≥ 1.6 from FET-PET. PTV2 delineation was MRI-based. The total dose was prescribed to 72 and 60 Gy for PTV1 and PTV2, using daily fractions of 2.4 and 2 Gy. Results After auto-contouring of PTV1 a marked target shape complexity had an impact on the dosimetric outcome. Patients with 3-4 PTV1 subvolumes vs. a single volume revealed a significant decrease in mean dose (67.7 vs. 70.6 Gy). From convex to complex shaped PTV1 mean doses decreased from 71.3 Gy to 67.7 Gy. The homogeneity and conformity for PTV1 and PTV2 was significantly improved with IB-IMRT. With the use of IB-IMRT the minimum dose within PTV1 (61.1 vs. 57.4 Gy) and PTV2 (51.4 vs. 40.9 Gy) increased significantly, and the mean EUD for PTV2 was improved (59.9 vs. 55.3 Gy, p < 0.01). The EUD for PTV1 was only slightly improved (68.3 vs. 67.3 Gy). The EUD for the brain was equal with both planning techniques. Conclusion In the presented planning study the integrated boost concept based on inversely planned IB-IMRT is feasible. The FET-PET-based automatically contoured PTV1 can lead to very complex geometric configurations, limiting the

  5. Professor Pet.

    ERIC Educational Resources Information Center

    Pet Information Bureau, New York, NY.

    This manual outlines ways in which observation and care of classroom pet animals may be used to enrich the education of elementary school children. Part one deals with the benefits of having pets in the classroom. Part two illustrates ways in which pets can serve as valuable teaching tools and gives examples of lessons in which the use of pets can…

  6. Food Targeting: A Real-Time PCR Assay Targeting 16S rDNA for Direct Quantification of Alicyclobacillus spp. Spores after Aptamer-Based Enrichment.

    PubMed

    Hünniger, Tim; Felbinger, Christine; Wessels, Hauke; Mast, Sophia; Hoffmann, Antonia; Schefer, Anna; Märtlbauer, Erwin; Paschke-Kratzin, Angelika; Fischer, Markus

    2015-05-01

    Spore-forming Alicyclobacillus spp. are able to form metabolites that induce even in small amounts an antiseptical or medicinal off-flavor in fruit juices. Microbial contaminations could occur by endospores, which overcame the pasteurization process. The current detection method for Alicyclobacillus spp. can take up to 1 week because of microbiological enrichment. In a previous study, DNA aptamers were selected and characterized for an aptamer-driven rapid enrichment of Alicyclobacillus spp. spores from orange juice by magnetic separation. In the present work, a direct quantification assay for Alicyclobacillus spp. spores was developed to complete the two-step approach of enrichment and detection. After mechanical treatment of the spores, the isolated DNA was quantified in a real-time PCR-assay targeting 16S rDNA. The assay was evaluated by the performance requirements of the European Network of Genetically Modified Organisms Laboratories (ENGL). Hence, the presented method is applicable for direct spore detection from orange juice in connection with an enrichment step. PMID:25880790

  7. Evaluation of 68Ga-labeled MG7 antibody: a targeted probe for PET/CT imaging of gastric cancer.

    PubMed

    Xu, Bing; Li, Xiaowei; Yin, Jipeng; Liang, Cong; Liu, Lijuan; Qiu, Zhaoyan; Yao, Liping; Nie, Yongzhan; Wang, Jing; Wu, Kaichun

    2015-01-01

    MG7-Ag, a specific gastric cancer-associated antigen, can be used to non-invasively monitor gastric cancer by molecular imaging with positron emission tomography/computed tomography (PET/CT). In this study, we prepared and evaluated a (68)Ga-labeled MG7 antibody as a molecular probe for nanoPET/CT imaging of gastric cancer in a BGC-823 tumor xenografted mouse model. Macrocyclic chelator 1,4,7-triazacyclononane-N,N0,N00-triacetic acid (NOTA)-conjugated MG7 antibody was synthesized and radiolabeled with (68)Ga (t1/2 = 67.71 min). Then, (68)Ga-NOTA-MG7 was tested using in vitro cytological studies, in vivo nanoPET/CT and Cerenkov imaging studies as well as ex vivo biodistribution and histology studies. The in vitro experiments demonstrated that (68)Ga-NOTA-MG7 has an excellent radiolabeling efficiency of approximately 99% without purification, and it is stable in serum after 120 min of incubation. Cell uptake and retention studies confirmed that (68)Ga-NOTA-MG7 has good binding affinity and tumor cell retention. For the nanoPET imaging study, the predominant uptake of (68)Ga-NOTA-MG7 was visualized in tumor, liver and kidneys. The tumor uptake reached at its peak (2.53 ± 0.28%ID/g) at 60 min pi. Cherenkov imaging also confirmed the specificity of tumor uptake. Moreover, the biodistribution results were consistent with the quantification data of nanoPET/CT imaging. Histologic analysis also demonstrated specific staining of BGC-823 tumor cell lines. PMID:25733152

  8. Evaluation of 68Ga-Labeled MG7 Antibody: A Targeted Probe for PET/CT Imaging of Gastric Cancer

    PubMed Central

    Xu, Bing; Li, Xiaowei; Yin, Jipeng; Liang, Cong; Liu, Lijuan; Qiu, Zhaoyan; Yao, Liping; Nie, Yongzhan; Wang, Jing; Wu, Kaichun

    2015-01-01

    MG7-Ag, a specific gastric cancer-associated antigen, can be used to non-invasively monitor gastric cancer by molecular imaging with positron emission tomography/computed tomography (PET/CT). In this study, we prepared and evaluated a 68Ga-labeled MG7 antibody as a molecular probe for nanoPET/CT imaging of gastric cancer in a BGC-823 tumor xenografted mouse model. Macrocyclic chelator 1,4,7-triazacyclononane-N,N0,N00-triacetic acid (NOTA)-conjugated MG7 antibody was synthesized and radiolabeled with 68Ga (t1/2 = 67.71 min). Then, 68Ga-NOTA-MG7 was tested using in vitro cytological studies, in vivo nanoPET/CT and Cerenkov imaging studies as well as ex vivo biodistribution and histology studies. The in vitro experiments demonstrated that 68Ga-NOTA-MG7 has an excellent radiolabeling efficiency of approximately 99% without purification, and it is stable in serum after 120 min of incubation. Cell uptake and retention studies confirmed that 68Ga-NOTA-MG7 has good binding affinity and tumor cell retention. For the nanoPET imaging study, the predominant uptake of 68Ga-NOTA-MG7 was visualized in tumor, liver and kidneys. The tumor uptake reached at its peak (2.53 ± 0.28%ID/g) at 60 min pi. Cherenkov imaging also confirmed the specificity of tumor uptake. Moreover, the biodistribution results were consistent with the quantification data of nanoPET/CT imaging. Histologic analysis also demonstrated specific staining of BGC-823 tumor cell lines. PMID:25733152

  9. A Novel PCR Assay for Listeria welshimeri Targeting Transcriptional Regulator Gene lwe1801

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transcriptional regulator genes encode a group of specialized molecules that play essential roles in microbial responses to changing external conditions. These genes have been shown to possess species or group specificity and are useful as detection targets for diagnostic application. The present st...

  10. Novel phakopsora pachyrhizi extracellular proteins are ideal targets for immunological diagnostic assays

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Phakopsora pachyrhizi, the causal agent of Asian soybean rust (ASR), continues to expand across the southeast and mid-south regions of the U.S., resulting in increased fungicide applications for producers. Our objectives in this research were to identify ASR protein targets for development of immuno...

  11. Multi-laboratory evaluations of the performance of Catellicoccus marimammalium PCR assays developed to target gull fecal sources

    USGS Publications Warehouse

    Sinigalliano, Christopher D.; Ervin, Jared S.; Van De Werfhorst, Laurie C.; Badgley, Brian D.; Ballestée, Elisenda; Bartkowiaka, Jakob; Boehm, Alexandria B.; Byappanahalli, Muruleedhara N.; Goodwin, Kelly D.; Gourmelon, Michèle; Griffith, John; Holden, Patricia A.; Jay, Jenny; Layton, Blythe; Lee, Cheonghoon; Lee, Jiyoung; Meijer, Wim G.; Noble, Rachel; Raith, Meredith; Ryu, Hodon; Sadowsky, Michael J.; Schriewer, Alexander; Wang, Dan; Wanless, David; Whitman, Richard; Wuertz, Stefan; Santo Domingo, Jorge W.

    2013-01-01

    Here we report results from a multi-laboratory (n = 11) evaluation of four different PCR methods targeting the 16S rRNA gene of Catellicoccus marimammalium originally developed to detect gull fecal contamination in coastal environments. The methods included a conventional end-point PCR method, a SYBR® Green qPCR method, and two TaqMan® qPCR methods. Different techniques for data normalization and analysis were tested. Data analysis methods had a pronounced impact on assay sensitivity and specificity calculations. Across-laboratory standardization of metrics including the lower limit of quantification (LLOQ), target detected but not quantifiable (DNQ), and target not detected (ND) significantly improved results compared to results submitted by individual laboratories prior to definition standardization. The unit of measure used for data normalization also had a pronounced effect on measured assay performance. Data normalization to DNA mass improved quantitative method performance as compared to enterococcus normalization. The MST methods tested here were originally designed for gulls but were found in this study to also detect feces from other birds, particularly feces composited from pigeons. Sequencing efforts showed that some pigeon feces from California contained sequences similar to C. marimammalium found in gull feces. These data suggest that the prevalence, geographic scope, and ecology of C. marimammalium in host birds other than gulls require further investigation. This study represents an important first step in the multi-laboratory assessment of these methods and highlights the need to broaden and standardize additional evaluations, including environmentally relevant target concentrations in ambient waters from diverse geographic regions.

  12. Multi-laboratory evaluations of the performance of Catellicoccus marimammalium PCR assays developed to target gull fecal sources.

    PubMed

    Sinigalliano, Christopher D; Ervin, Jared S; Van De Werfhorst, Laurie C; Badgley, Brian D; Ballesté, Elisenda; Bartkowiak, Jakob; Boehm, Alexandria B; Byappanahalli, Muruleedhara; Goodwin, Kelly D; Gourmelon, Michèle; Griffith, John; Holden, Patricia A; Jay, Jenny; Layton, Blythe; Lee, Cheonghoon; Lee, Jiyoung; Meijer, Wim G; Noble, Rachel; Raith, Meredith; Ryu, Hodon; Sadowsky, Michael J; Schriewer, Alexander; Wang, Dan; Wanless, David; Whitman, Richard; Wuertz, Stefan; Santo Domingo, Jorge W

    2013-11-15

    Here we report results from a multi-laboratory (n = 11) evaluation of four different PCR methods targeting the 16S rRNA gene of Catellicoccus marimammalium originally developed to detect gull fecal contamination in coastal environments. The methods included a conventional end-point PCR method, a SYBR(®) Green qPCR method, and two TaqMan(®) qPCR methods. Different techniques for data normalization and analysis were tested. Data analysis methods had a pronounced impact on assay sensitivity and specificity calculations. Across-laboratory standardization of metrics including the lower limit of quantification (LLOQ), target detected but not quantifiable (DNQ), and target not detected (ND) significantly improved results compared to results submitted by individual laboratories prior to definition standardization. The unit of measure used for data normalization also had a pronounced effect on measured assay performance. Data normalization to DNA mass improved quantitative method performance as compared to enterococcus normalization. The MST methods tested here were originally designed for gulls but were found in this study to also detect feces from other birds, particularly feces composited from pigeons. Sequencing efforts showed that some pigeon feces from California contained sequences similar to C. marimammalium found in gull feces. These data suggest that the prevalence, geographic scope, and ecology of C. marimammalium in host birds other than gulls require further investigation. This study represents an important first step in the multi-laboratory assessment of these methods and highlights the need to broaden and standardize additional evaluations, including environmentally relevant target concentrations in ambient waters from diverse geographic regions. PMID:23916157

  13. Planar optical waveguide based sandwich assay sensors and processes for the detection of biological targets including early detection of cancers

    DOEpatents

    Martinez, Jennifer S.; Swanson, Basil I.; Shively, John E.; Li, Lin

    2009-06-02

    An assay element is described including recognition ligands adapted for binding to carcinoembryonic antigen (CEA) bound to a film on a single mode planar optical waveguide, the film from the group of a membrane, a polymerized bilayer membrane, and a self-assembled monolayer containing polyethylene glycol or polypropylene glycol groups therein and an assay process for detecting the presence of CEA is described including injecting a possible CEA-containing sample into a sensor cell including the assay element, maintaining the sample within the sensor cell for time sufficient for binding to occur between CEA present within the sample and the recognition ligands, injecting a solution including a reporter ligand into the sensor cell; and, interrogating the sample within the sensor cell with excitation light from the waveguide, the excitation light provided by an evanescent field of the single mode penetrating into the biological target-containing sample to a distance of less than about 200 nanometers from the waveguide thereby exciting any bound reporter ligand within a distance of less than about 200 nanometers from the waveguide and resulting in a detectable signal.

  14. Biosensor assay of neuropathy target esterase in whole blood as a new approach to OPIDN risk assessment: review of progress.

    PubMed

    Makhaeva, Galina F; Malygin, Vladimir V; Strakhova, Nadezhda N; Sigolaeva, Larisa V; Sokolovskaya, Lidia G; Eremenko, Arkady V; Kurochkin, Ilya N; Richardson, Rudy J

    2007-04-01

    Organophosphates (OPs) that inhibit neuropathy target esterase (NTE) with subsequent ageing can produce OP-induced delayed neuropathy (OPIDN). NTE inhibition in lymphocytes can be used as a biomarker of exposure to neuropathic OPs. An electrochemical method was developed to assay NTE in whole blood. The high sensitivity of the tyrosinase carbon-paste biosensors for the phenol produced by hydrolysis of the substrate, phenyl valerate, allowed NTE activity to be measured in diluted samples of whole blood, which cannot be done using the standard colorimetric assay. The biosensor was used to establish correlations of NTE inhibitions in blood with that in lymphocytes and brain after dosing hens with a neuropathic OP. The results of further studies demonstrated that whole blood NTE is a reliable biomarker of neuropathic OPs for up to 96 hours after exposure. These validation results suggest that the biosensor NTE assay for whole blood could be developed to measure human exposure to neuropathic OPs as a predictor of OPIDN. The small blood volume required (100 microL), simplicity of sample preparation and rapid analysis times indicate that the biosensor should be useful in biomonitoring and epidemiological studies. The present paper is an overview of our previous and ongoing work in this area. PMID:17615108

  15. Luciferase-based protein-denaturation assay for quantification of radiofrequency field-induced targeted hyperthermia: developing an intracellular thermometer

    PubMed Central

    Raoof, Mustafa; Zhu, Cihui; Kaluarachchi, Warna D.; Curley, Steven A.

    2013-01-01

    Background Several studies have reported targeted hyperthermia at the cellular level using remote activation of nanoparticles by radiofrequency waves. To date, methods to quantify intracellular thermal dose have not been reported. In this report we study the relationship between radio wave exposure and luciferase denaturation with and without intracellular nanoparticles. The findings are used to devise a strategy to quantify targeted thermal dose in a primary human liver cancer cell line. Methods Water-bath or non-invasive external RF generator (600W, 13.56 MHz) was used for hyperthermia exposures. Luciferase activity was measured using a bioluminescence assay and viability was assessed using Annexin V-FITC and Propidium iodide staining. Heat shock proteins were analyzed using western-blot analysis Results Duration-dependent luciferase denaturation was observed in SNU449 cells exposed to RF field that preceded measurable loss in viability. Loss of luciferase activity was higher in cetuximab-conjugated gold nanoparticle (C225-AuNP) treated cells. Using a standard curve from water-bath experiments, the intracellular thermal dose was calculated. Cells treated with C225-AuNP accumulated 6.07 times higher intracellular thermal dose than the untreated controls over initial 4 minutes of RF exposure. Conclusions Cancer cells when exposed to an external RF field exhibit dose-dependent protein denaturation. Luciferase denaturation assay can be used to quantify thermal dose delivered after RF exposures to cancer cells with and without nanoparticles. PMID:22515341

  16. Novel Antibacterial Targets and Compounds Revealed by a High-Throughput Cell Wall Reporter Assay

    PubMed Central

    Nayar, Asha S.; Dougherty, Thomas J.; Ferguson, Keith E.; Granger, Brett A.; McWilliams, Lisa; Stacey, Clare; Leach, Lindsey J.; Narita, Shin-ichiro; Tokuda, Hajime; Miller, Alita A.; Brown, Dean G.

    2015-01-01

    ABSTRACT A high-throughput phenotypic screen based on a Citrobacter freundii AmpC reporter expressed in Escherichia coli was executed to discover novel inhibitors of bacterial cell wall synthesis, an attractive, well-validated target for antibiotic intervention. Here we describe the discovery and characterization of sulfonyl piperazine and pyrazole compounds, each with novel mechanisms of action. E. coli mutants resistant to these compounds display no cross-resistance to antibiotics of other classes. Resistance to the sulfonyl piperazine maps to LpxH, which catalyzes the fourth step in the synthesis of lipid A, the outer membrane anchor of lipopolysaccharide (LPS). To our knowledge, this compound is the first reported inhibitor of LpxH. Resistance to the pyrazole compound mapped to mutations in either LolC or LolE, components of the essential LolCDE transporter complex, which is required for trafficking of lipoproteins to the outer membrane. Biochemical experiments with E. coli spheroplasts showed that the pyrazole compound is capable of inhibiting the release of lipoproteins from the inner membrane. Both of these compounds have significant promise as chemical probes to further interrogate the potential of these novel cell wall components for antimicrobial therapy. IMPORTANCE The prevalence of antibacterial resistance, particularly among Gram-negative organisms, signals a need for novel antibacterial agents. A phenotypic screen using AmpC as a sensor for compounds that inhibit processes involved in Gram-negative envelope biogenesis led to the identification of two novel inhibitors with unique mechanisms of action targeting Escherichia coli outer membrane biogenesis. One compound inhibits the transport system for lipoprotein transport to the outer membrane, while the other compound inhibits synthesis of lipopolysaccharide. These results indicate that it is still possible to uncover new compounds with intrinsic antibacterial activity that inhibit novel targets

  17. Defining balanced conditions for inhibitor screening assays that target bisubstrate enzymes.

    PubMed

    Yang, Jingsong; Copeland, Robert A; Lai, Zhihong

    2009-02-01

    High-throughput screening (HTS) is a common mechanism for identifying lead compounds for drug discovery efforts. Small molecules can inhibit enzymes by a variety of mechanisms, such as competitive, noncompetitive, and uncompetitive with respect to the substrate(s) of the catalytic reaction. To optimize the chances of finding the broadest diversity of inhibitor modalities during screening, one must run assays under ;;balanced'' conditions where the potency of inhibitors with various modes of action falls within a similar range. When an enzyme reaction involves more than one substrate, the definition and assessment of the apparent potency of inhibitors (IC(50)), in relation to their true potency (K(i)), can be nontrivial. This article provides a theoretical analysis, on the basis of the Cheng-Prusoff derivation, of the IC(50)/K( i) relationship of bisubstrate enzyme reactions following various sequential kinetic mechanisms, as well as the application and limitations of this information for defining optimal screening conditions for such enzymes. PMID:19196704

  18. {sup 18}F-Fluorodeoxyglucose Positron Emission Tomography/Computed Tomography-Based Radiotherapy Target Volume Definition in Non-Small-Cell Lung Cancer: Delineation by Radiation Oncologists vs. Joint Outlining With a PET Radiologist?

    SciTech Connect

    Hanna, Gerard G.; Carson, Kathryn J.; Lynch, Tom; McAleese, Jonathan; Cosgrove, Vivian P.; Eakin, Ruth L.; Stewart, David P.; Zatari, Ashraf; O'Sullivan, Joe M.; Hounsell, Alan R.

    2010-11-15

    Purpose: {sup 18}F-Fluorodeoxyglucose positron emission tomography/computed tomography (PET/CT) has benefits in target volume (TV) definition in radiotherapy treatment planning (RTP) for non-small-cell lung cancer (NSCLC); however, an optimal protocol for TV delineation has not been determined. We investigate volumetric and positional variation in gross tumor volume (GTV) delineation using a planning PET/CT among three radiation oncologists and a PET radiologist. Methods and Materials: RTP PET/CT scans were performed on 28 NSCLC patients (Stage IA-IIIB) of which 14 patients received prior induction chemotherapy. Three radiation oncologists and one PET radiologist working with a fourth radiation oncologist independently delineated the GTV on CT alone (GTV{sub CT}) and on fused PET/CT images (GTV{sub PETCT}). The mean percentage volume change (PVC) between GTV{sub CT} and GTV{sub PETCT} for the radiation oncologists and the PVC between GTV{sub CT} and GTV{sub PETCT} for the PET radiologist were compared using the Wilcoxon signed-rank test. Concordance index (CI) was used to assess both positional and volume change between GTV{sub CT} and GTV{sub PETCT} in a single measurement. Results: For all patients, a significant difference in PVC from GTV{sub CT} to GTV{sub PETCT} exists between the radiation oncologist (median, 5.9%), and the PET radiologist (median, -0.4%, p = 0.001). However, no significant difference in median concordance index (comparing GTV{sub CT} and GTV{sub FUSED} for individual cases) was observed (PET radiologist = 0.73; radiation oncologists = 0.66; p = 0.088). Conclusions: Percentage volume changes from GTV{sub CT} to GTV{sub PETCT} were lower for the PET radiologist than for the radiation oncologists, suggesting a lower impact of PET/CT in TV delineation for the PET radiologist than for the oncologists. Guidelines are needed to standardize the use of PET/CT for TV delineation in RTP.

  19. A targeted next-generation sequencing assay for the molecular diagnosis of genetic disorders with orodental involvement

    PubMed Central

    Prasad, Megana K; Geoffroy, Véronique; Vicaire, Serge; Jost, Bernard; Dumas, Michael; Le Gras, Stéphanie; Switala, Marzena; Gasse, Barbara; Laugel-Haushalter, Virginie; Paschaki, Marie; Leheup, Bruno; Droz, Dominique; Dalstein, Amelie; Loing, Adeline; Grollemund, Bruno; Muller-Bolla, Michèle; Lopez-Cazaux, Séréna; Minoux, Maryline; Jung, Sophie; Obry, Frédéric; Vogt, Vincent; Davideau, Jean-Luc; Davit-Beal, Tiphaine; Kaiser, Anne-Sophie; Moog, Ute; Richard, Béatrice; Morrier, Jean-Jacques; Duprez, Jean-Pierre; Odent, Sylvie; Bailleul-Forestier, Isabelle; Rousset, Monique Marie; Merametdijan, Laure; Toutain, Annick; Joseph, Clara; Giuliano, Fabienne; Dahlet, Jean-Christophe; Courval, Aymeric; El Alloussi, Mustapha; Laouina, Samir; Soskin, Sylvie; Guffon, Nathalie; Dieux, Anne; Doray, Bérénice; Feierabend, Stephanie; Ginglinger, Emmanuelle; Fournier, Benjamin; de la Dure Molla, Muriel; Alembik, Yves; Tardieu, Corinne; Clauss, François; Berdal, Ariane; Stoetzel, Corinne; Manière, Marie Cécile; Dollfus, Hélène; Bloch-Zupan, Agnès

    2016-01-01

    Background Orodental diseases include several clinically and genetically heterogeneous disorders that can present in isolation or as part of a genetic syndrome. Due to the vast number of genes implicated in these disorders, establishing a molecular diagnosis can be challenging. We aimed to develop a targeted next-generation sequencing (NGS) assay to diagnose mutations and potentially identify novel genes mutated in this group of disorders. Methods We designed an NGS gene panel that targets 585 known and candidate genes in orodental disease. We screened a cohort of 101 unrelated patients without a molecular diagnosis referred to the Reference Centre for Oro-Dental Manifestations of Rare Diseases, Strasbourg, France, for a variety of orodental disorders including isolated and syndromic amelogenesis imperfecta (AI), isolated and syndromic selective tooth agenesis (STHAG), isolated and syndromic dentinogenesis imperfecta, isolated dentin dysplasia, otodental dysplasia and primary failure of tooth eruption. Results We discovered 21 novel pathogenic variants and identified the causative mutation in 39 unrelated patients in known genes (overall diagnostic rate: 39%). Among the largest subcohorts of patients with isolated AI (50 unrelated patients) and isolated STHAG (21 unrelated patients), we had a definitive diagnosis in 14 (27%) and 15 cases (71%), respectively. Surprisingly, COL17A1 mutations accounted for the majority of autosomal-dominant AI cases. Conclusions We have developed a novel targeted NGS assay for the efficient molecular diagnosis of a wide variety of orodental diseases. Furthermore, our panel will contribute to better understanding the contribution of these genes to orodental disease. Trial registration numbers NCT01746121 and NCT02397824. PMID:26502894

  20. Validation, optimisation, and application data in support of the development of a targeted selected ion monitoring assay for degraded cardiac troponin T

    PubMed Central

    Streng, Alexander S.; de Boer, Douwe; Bouwman, Freek G.; Mariman, Edwin C.M.; Scholten, Arjen; van Dieijen-Visser, Marja P.; Wodzig, Will K.W.H.

    2016-01-01

    Cardiac troponin T (cTnT) fragmentation in human serum was investigated using a newly developed targeted selected ion monitoring assay, as described in the accompanying article: “Development of a targeted selected ion monitoring assay for the elucidation of protease induced structural changes in cardiac troponin T” [1]. This article presents data describing aspects of the validation and optimisation of this assay. The data consists of several figures, an excel file containing the results of a sequence identity search, and a description of the raw mass spectrometry (MS) data files, deposited in the ProteomeXchange repository with id PRIDE: PXD003187. PMID:26977445

  1. Validation, optimisation, and application data in support of the development of a targeted selected ion monitoring assay for degraded cardiac troponin T.

    PubMed

    Streng, Alexander S; de Boer, Douwe; Bouwman, Freek G; Mariman, Edwin C M; Scholten, Arjen; van Dieijen-Visser, Marja P; Wodzig, Will K W H

    2016-06-01

    Cardiac troponin T (cTnT) fragmentation in human serum was investigated using a newly developed targeted selected ion monitoring assay, as described in the accompanying article: "Development of a targeted selected ion monitoring assay for the elucidation of protease induced structural changes in cardiac troponin T" [1]. This article presents data describing aspects of the validation and optimisation of this assay. The data consists of several figures, an excel file containing the results of a sequence identity search, and a description of the raw mass spectrometry (MS) data files, deposited in the ProteomeXchange repository with id PRIDE: PXD003187. PMID:26977445

  2. In Vivo Tumor Vasculature Targeted PET/NIRF Imaging with TRC105(Fab)-Conjugated, Dual-Labeled Mesoporous Silica Nanoparticles

    PubMed Central

    2015-01-01

    Multifunctional mesoporous silica nanoparticles (MSN) with well-integrated multimodality imaging properties have generated increasing research interest in the past decade. However, limited progress has been made in developing MSN-based multimodality imaging agents to image tumors. We describe the successful conjugation of, copper-64 (64Cu, t1/2 = 12.7 h), 800CW (a near-infrared fluorescence [NIRF] dye), and TRC105 (a human/murine chimeric IgG1 monoclonal antibody) to the surface of MSN via well-developed surface engineering procedures, resulting in a dual-labeled MSN for in vivo targeted positron emission tomography (PET) imaging/NIRF imaging of the tumor vasculature. Pharmacokinetics and tumor targeting efficacy/specificity in 4T1 murine breast tumor-bearing mice were thoroughly investigated through various in vitro, in vivo, and ex vivo experiments. Dual-labeled MSN is an attractive candidate for future cancer theranostics. PMID:24937108

  3. In vivo tumor vasculature targeted PET/NIRF imaging with TRC105(Fab)-conjugated, dual-labeled mesoporous silica nanoparticles.

    PubMed

    Chen, Feng; Nayak, Tapas R; Goel, Shreya; Valdovinos, Hector F; Hong, Hao; Theuer, Charles P; Barnhart, Todd E; Cai, Weibo

    2014-11-01

    Multifunctional mesoporous silica nanoparticles (MSN) with well-integrated multimodality imaging properties have generated increasing research interest in the past decade. However, limited progress has been made in developing MSN-based multimodality imaging agents to image tumors. We describe the successful conjugation of, copper-64 ((64)Cu, t1/2 = 12.7 h), 800CW (a near-infrared fluorescence [NIRF] dye), and TRC105 (a human/murine chimeric IgG1 monoclonal antibody) to the surface of MSN via well-developed surface engineering procedures, resulting in a dual-labeled MSN for in vivo targeted positron emission tomography (PET) imaging/NIRF imaging of the tumor vasculature. Pharmacokinetics and tumor targeting efficacy/specificity in 4T1 murine breast tumor-bearing mice were thoroughly investigated through various in vitro, in vivo, and ex vivo experiments. Dual-labeled MSN is an attractive candidate for future cancer theranostics. PMID:24937108

  4. Sandwich assay for mixed-sequence recognition of double-stranded DNA: Invader-based detection of targets specific to food pathogens†

    PubMed Central

    Denn, Benjamin; Karmakar, Saswata; Guenther, Dale C.; Hrdlicka, Patrick J.

    2014-01-01

    A 96-well plate sandwich assay based on Invader capture/signalling probes is used to recognize 28-mer mixed-sequence dsDNA targets specific to Salmonella, Campylobacter and Escherichia coli. Targets are detected at 20-55 pM concentration with excellent binding specificity. PMID:24036937

  5. Comparison of Gull Feces-Specific Assays Targeting the 16S rRNA Genes of Catellicoccus marimammalium and Streptococcus spp.

    PubMed Central

    Ryu, Hodon; Griffith, John F.; Khan, Izhar U. H.; Hill, Stephen; Edge, Thomas A.; Toledo-Hernandez, Carlos; Gonzalez-Nieves, Joel

    2012-01-01

    Two novel gull-specific quantitative PCR (qPCR) assays were developed using 16S rRNA gene sequences from gull fecal clone libraries: a SYBR green assay targeting Streptococcus spp. (gull3) and a hydrolysis TaqMan assay targeting Catellicoccus marimammalium (gull4). The objectives of this study were to compare the host specificity of a previous C. marimammalium qPCR assay (gull2) with that of the new markers and to examine the presence of the three gull markers in environmental water samples from different geographic locations. Most of the gull fecal samples tested (n = 255) generated positive signals with the gull2 and gull4 assays (i.e., >86%), whereas only 28% were positive with gull3. Low prevalence and abundance of tested gull markers (0.6 to 15%) were observed in fecal samples from six nonavian species (n = 180 fecal samples), whereas the assays cross-reacted to some extent (13 to 31%) with other (nongull) avian fecal samples. The gull3 assay was positive against fecal samples from 11 of 15 avian species, including gull. Of the presumed gull-impacted water samples (n = 349), 86%, 59%, and 91% were positive with the gull2, the gull3, and the gull4 assays, respectively. Approximately 5% of 239 non-gull-impacted water samples were positive with the gull2 and the gull4 assays, whereas 21% were positive witg the gull3 assay. While the relatively high occurrence of gull2 and gull4 markers in waters impacted by gull feces suggests that these assays could be used in environmental monitoring studies, the data also suggest that multiple avian-specific assays will be needed to accurately assess the contribution of different avian sources in recreational waters. PMID:22226950

  6. Comparison of gull feces-specific assays targeting the 16S rRNA genes of Catellicoccus marimammalium and Streptococcus spp.

    PubMed

    Ryu, Hodon; Griffith, John F; Khan, Izhar U H; Hill, Stephen; Edge, Thomas A; Toledo-Hernandez, Carlos; Gonzalez-Nieves, Joel; Santo Domingo, Jorge

    2012-03-01

    Two novel gull-specific quantitative PCR (qPCR) assays were developed using 16S rRNA gene sequences from gull fecal clone libraries: a SYBR green assay targeting Streptococcus spp. (gull3) and a hydrolysis TaqMan assay targeting Catellicoccus marimammalium (gull4). The objectives of this study were to compare the host specificity of a previous C. marimammalium qPCR assay (gull2) with that of the new markers and to examine the presence of the three gull markers in environmental water samples from different geographic locations. Most of the gull fecal samples tested (n = 255) generated positive signals with the gull2 and gull4 assays (i.e., >86%), whereas only 28% were positive with gull3. Low prevalence and abundance of tested gull markers (0.6 to 15%) were observed in fecal samples from six nonavian species (n = 180 fecal samples), whereas the assays cross-reacted to some extent (13 to 31%) with other (nongull) avian fecal samples. The gull3 assay was positive against fecal samples from 11 of 15 avian species, including gull. Of the presumed gull-impacted water samples (n = 349), 86%, 59%, and 91% were positive with the gull2, the gull3, and the gull4 assays, respectively. Approximately 5% of 239 non-gull-impacted water samples were positive with the gull2 and the gull4 assays, whereas 21% were positive witg the gull3 assay. While the relatively high occurrence of gull2 and gull4 markers in waters impacted by gull feces suggests that these assays could be used in environmental monitoring studies, the data also suggest that multiple avian-specific assays will be needed to accurately assess the contribution of different avian sources in recreational waters. PMID:22226950

  7. Rapid and reliable identification of Staphylococcus equorum by a species-specific PCR assay targeting the sodA gene.

    PubMed

    Blaiotta, Giuseppe; Ercolini, Danilo; Mauriello, Gianluigi; Salzano, Giovanni; Villani, Francesco

    2004-11-01

    Rapid and reliable identification of Staphylococcus (S.) equorum was achieved by species-specific PCR assays. A set of primers targeting the manganese-dependent superoxide dismutase (sodA) gene of S. equorum was designed. Species-specificity of the primer set was evaluated by using a total of 112 strains (including 27 reference strains of the DSM collection), representing 26 different species of the genus Staphylococcus, 3 species of the genus Kocuria, and different strains of Macrococcus caseolyticus. By using primers SdAEqF and SdAEqR the expected PCR fragment was obtained only when DNA from S. equorum strains was used as template. The rapidity (about 4 h from DNA isolation to results) and reliability of the PCR procedures established suggests that the method may be profitably applied for specific detection and identification of S. equorum strains. PMID:15612627

  8. Effect of peptide assay library size and composition in targeted data-independent acquisition-MS analyses.

    PubMed

    Parker, Sarah J; Venkatraman, Vidya; Van Eyk, Jennifer E

    2016-08-01

    The quantification of peptides using targeted analysis of data-independent acquisition MS (DIA-MS) is dependent on the size and characteristics of the assay library. We addressed several important questions on how library composition influences: (1) the number of peptides extracted from DIA-MS datasets, (2) the quality of these peptides and proteins, and (3) the biological conclusions inferred. To answer these questions we constructed five libraries from mouse vascular smooth muscle cell (VSMC) lysate, each unique in depth, input sample complexity, data acquisition mode (DDA-MS or DIA-MS), and precursor fragmentation mode (TOF-CID or Orbitrap HCD) and extracted them against the same eight DIA-MS files of VSMCs treated with vehicle or transforming growth factor β-1 (TGF-β1). We found that along with differences in peptide and protein composition, the fragments representing a given peptide differed between the libraries. Collectively these differences impacted both peak group score profile and protein abundance estimates. Surprisingly, there was little overlap in the TGF-β1 response proteome between libraries. We conclude that additional work is needed to optimize peptide assay library building for DIA-MS applications, particularly in terms of selecting optimal peptides and their respective fragments for protein quantification. PMID:27432805

  9. Pet Health

    MedlinePlus

    ... Before getting a pet, think carefully about which animal is best for your family. What is each ... Does anyone have pet allergies? What type of animal suits your lifestyle and budget? Once you own ...

  10. Identification of Hepatitis C Virus Inhibitors Targeting Different Aspects of Infection Using a Cell-Based Assay

    PubMed Central

    Yu, Xuemei; Sainz, Bruno; Petukhov, Pavel A.

    2012-01-01

    With 2 to 3% of the worldwide population chronically infected, hepatitis C virus (HCV) infection continues to be a major health care burden. Unfortunately, current interferon-based treatment options are not effective in all patients and are associated with significant side effects. Consequently, there is an ongoing need to identify and develop new anti-HCV therapies. Toward this goal, we previously developed a cell-based HCV infection assay for antiviral compound screening based on a low-multiplicity-of-infection approach that uniquely allows for the identification of antiviral compounds that target cell culture-derived HCV (HCVcc) at any step of the viral infection cycle. Using this assay, here we report the screening of the NCI Diversity Set II library, containing 1,974 synthesized chemical compounds, and the identification of compounds with specific anti-HCV activity. In combination with toxicity counterscreening, we identified 30 hits from the compound library, 13 of which showed reproducible and dose-dependent inhibition of HCV with mean therapeutic indices (50% cytotoxic concentration [CC50]/50% effective concentration [EC50]) of greater than 6. Using HCV pseudotype and replicon systems of multiple HCV genotypes, as well as infectious HCVcc-based assembly and secretion analysis, we determined that different compounds within this group of candidate inhibitors target different steps of viral infection. The compounds identified not only will serve as biological probes to study and further dissect the biology of viral infection but also should facilitate the development of new anti-HCV therapeutic treatments. PMID:22948883

  11. Identification of hepatitis C virus inhibitors targeting different aspects of infection using a cell-based assay.

    PubMed

    Yu, Xuemei; Sainz, Bruno; Petukhov, Pavel A; Uprichard, Susan L

    2012-12-01

    With 2 to 3% of the worldwide population chronically infected, hepatitis C virus (HCV) infection continues to be a major health care burden. Unfortunately, current interferon-based treatment options are not effective in all patients and are associated with significant side effects. Consequently, there is an ongoing need to identify and develop new anti-HCV therapies. Toward this goal, we previously developed a cell-based HCV infection assay for antiviral compound screening based on a low-multiplicity-of-infection approach that uniquely allows for the identification of antiviral compounds that target cell culture-derived HCV (HCVcc) at any step of the viral infection cycle. Using this assay, here we report the screening of the NCI Diversity Set II library, containing 1,974 synthesized chemical compounds, and the identification of compounds with specific anti-HCV activity. In combination with toxicity counterscreening, we identified 30 hits from the compound library, 13 of which showed reproducible and dose-dependent inhibition of HCV with mean therapeutic indices (50% cytotoxic concentration [CC(50)]/50% effective concentration [EC(50)]) of greater than 6. Using HCV pseudotype and replicon systems of multiple HCV genotypes, as well as infectious HCVcc-based assembly and secretion analysis, we determined that different compounds within this group of candidate inhibitors target different steps of viral infection. The compounds identified not only will serve as biological probes to study and further dissect the biology of viral infection but also should facilitate the development of new anti-HCV therapeutic treatments. PMID:22948883

  12. Initial Evaluation of [18F]DCFPyL for Prostate-Specific Membrane Antigen (PSMA)-Targeted PET Imaging of Prostate Cancer

    PubMed Central

    Szabo, Zsolt; Mena, Esther; Rowe, Steven P.; Plyku, Donika; Nidal, Rosa; Eisenberger, Mario A.; Antonarakis, Emmanuel S.; Fan, Hong; Dannals, Robert F.; Chen, Ying; Mease, Ronnie C.; Vranesic, Melin; Bhatnagar, Akrita; Sgouros, George; Cho, Steve Y.; Pomper, Martin G.

    2015-01-01

    Purpose Prostate-specific membrane antigen (PSMA) is a recognized target for imaging prostate cancer. Here we present initial safety, biodistribution, and radiation dosimetry results with [18F]DCFPyL, a second-generation fluorine-18-labeled small-molecule PSMA inhibitor, in patients with prostate cancer. Procedures Biodistribution was evaluated using sequential positron-emission tomography (PET) scans in nine patients with prostate cancer. Time-activity curves from the most avid tumor foci were determined. The radiation dose to selected organs was estimated using OLINDA/EXM. Results No major radiotracer-specific adverse events were observed. Physiologic accumulation was observed in known sites of PSMA expression. Accumulation in putative sites of prostate cancer was observed (SUVmax up to >100, and tumor-to-blood ratios up to >50). The effective radiation dose from [18F]DCFPyL was 0.0139 mGy/MBq or 5 mGy (0.5 rem) from an injected dose of 370 MBq (10 mCi). Conclusions [18F]DCFPyL is safe with biodistribution as expected, and its accumulation is high in presumed primary and metastatic foci. The radiation dose from [18F]DCFPyL is similar to that from other PET radiotracers. PMID:25896814

  13. Targeted Peptide Measurements in Biology and Medicine: Best Practices for Mass Spectrometry-based Assay Development Using a Fit-for-Purpose Approach

    SciTech Connect

    Carr, Steven A.; Abbateillo, Susan E.; Ackermann, Bradley L.; Borchers, Christoph H.; Domon, Bruno; Deutsch, Eric W.; Grant, Russel; Hoofnagle, Andrew N.; Huttenhain, Ruth; Koomen, John M.; Liebler, Daniel; Liu, Tao; MacLean, Brendan; Mani, DR; Mansfield, Elizabeth; Neubert, Hendrik; Paulovich, Amanda G.; Reiter, Lukas; Vitek, Olga; Aebersold, Ruedi; Anderson, Leigh N.; Bethem, Robert; Blonder, Josip; Boja, Emily; Botelho, Julianne; Boyne, Michael; Bradshaw, Ralph A.; Burlingame, Alma S.; Chan, Daniel W.; Keshishian, Hasmik; Kuhn, Eric; Kingsinger, Christopher R.; Lee, Jerry S.; Lee, Sang-Won; Moritz, Robert L.; Oses-Prieto, Juan; Rifai, Nader; Ritchie, James E.; Rodriguez, Henry; Srinivas, Pothur R.; Townsend, Reid; Van Eyk , Jennifer; Whiteley, Gordon; Wiita, Arun; Weintraub, Susan

    2014-01-14

    Adoption of targeted mass spectrometry (MS) approaches such as multiple reaction monitoring (MRM) to study biological and biomedical questions is well underway in the proteomics community. Successful application depends on the ability to generate reliable assays that uniquely and confidently identify target peptides in a sample. Unfortunately, there is a wide range of criteria being applied to say that an assay has been successfully developed. There is no consensus on what criteria are acceptable and little understanding of the impact of variable criteria on the quality of the results generated. Publications describing targeted MS assays for peptides frequently do not contain sufficient information for readers to establish confidence that the tests work as intended or to be able to apply the tests described in their own labs. Guidance must be developed so that targeted MS assays with established performance can be made widely distributed and applied by many labs worldwide. To begin to address the problems and their solutions, a workshop was held at the National Institutes of Health with representatives from the multiple communities developing and employing targeted MS assays. Participants discussed the analytical goals of their experiments and the experimental evidence needed to establish that the assays they develop work as intended and are achieving the required levels of performance. Using this “fit-for-purpose” approach, the group defined three tiers of assays distinguished by their performance and extent of analytical characterization. Computational and statistical tools useful for the analysis of targeted MS results were described. Participants also detailed the information that authors need to provide in their manuscripts to enable reviewers and readers to clearly understand what procedures were performed and to evaluate the reliability of the peptide or protein quantification measurements reported. This paper presents a summary of the meeting and

  14. Targeted Peptide Measurements in Biology and Medicine: Best Practices for Mass Spectrometry-based Assay Development Using a Fit-for-Purpose Approach*

    PubMed Central

    Carr, Steven A.; Abbatiello, Susan E.; Ackermann, Bradley L.; Borchers, Christoph; Domon, Bruno; Deutsch, Eric W.; Grant, Russell P.; Hoofnagle, Andrew N.; Hüttenhain, Ruth; Koomen, John M.; Liebler, Daniel C.; Liu, Tao; MacLean, Brendan; Mani, DR; Mansfield, Elizabeth; Neubert, Hendrik; Paulovich, Amanda G.; Reiter, Lukas; Vitek, Olga; Aebersold, Ruedi; Anderson, Leigh; Bethem, Robert; Blonder, Josip; Boja, Emily; Botelho, Julianne; Boyne, Michael; Bradshaw, Ralph A.; Burlingame, Alma L.; Chan, Daniel; Keshishian, Hasmik; Kuhn, Eric; Kinsinger, Christopher; Lee, Jerry S.H.; Lee, Sang-Won; Moritz, Robert; Oses-Prieto, Juan; Rifai, Nader; Ritchie, James; Rodriguez, Henry; Srinivas, Pothur R.; Townsend, R. Reid; Van Eyk, Jennifer; Whiteley, Gordon; Wiita, Arun; Weintraub, Susan

    2014-01-01

    Adoption of targeted mass spectrometry (MS) approaches such as multiple reaction monitoring (MRM) to study biological and biomedical questions is well underway in the proteomics community. Successful application depends on the ability to generate reliable assays that uniquely and confidently identify target peptides in a sample. Unfortunately, there is a wide range of criteria being applied to say that an assay has been successfully developed. There is no consensus on what criteria are acceptable and little understanding of the impact of variable criteria on the quality of the results generated. Publications describing targeted MS assays for peptides frequently do not contain sufficient information for readers to establish confidence that the tests work as intended or to be able to apply the tests described in their own labs. Guidance must be developed so that targeted MS assays with established performance can be made widely distributed and applied by many labs worldwide. To begin to address the problems and their solutions, a workshop was held at the National Institutes of Health with representatives from the multiple communities developing and employing targeted MS assays. Participants discussed the analytical goals of their experiments and the experimental evidence needed to establish that the assays they develop work as intended and are achieving the required levels of performance. Using this “fit-for-purpose” approach, the group defined three tiers of assays distinguished by their performance and extent of analytical characterization. Computational and statistical tools useful for the analysis of targeted MS results were described. Participants also detailed the information that authors need to provide in their manuscripts to enable reviewers and readers to clearly understand what procedures were performed and to evaluate the reliability of the peptide or protein quantification measurements reported. This paper presents a summary of the meeting and

  15. PET/CT in radiation oncology

    SciTech Connect

    Pan, Tinsu; Mawlawi, Osama

    2008-11-15

    PET/CT is an effective tool for the diagnosis, staging and restaging of cancer patients. It combines the complementary information of functional PET images and anatomical CT images in one imaging session. Conventional stand-alone PET has been replaced by PET/CT for improved patient comfort, patient throughput, and most importantly the proven clinical outcome of PET/CT over that of PET and that of separate PET and CT. There are over two thousand PET/CT scanners installed worldwide since 2001. Oncology is the main application for PET/CT. Fluorine-18 deoxyglucose is the choice of radiopharmaceutical in PET for imaging the glucose uptake in tissues, correlated with an increased rate of glycolysis in many tumor cells. New molecular targeted agents are being developed to improve the accuracy of targeting different disease states and assessing therapeutic response. Over 50% of cancer patients receive radiation therapy (RT) in the course of their disease treatment. Clinical data have demonstrated that the information provided by PET/CT often changes patient management of the patient and/or modifies the RT plan from conventional CT simulation. The application of PET/CT in RT is growing and will become increasingly important. Continuing improvement of PET/CT instrumentation will also make it easier for radiation oncologists to integrate PET/CT in RT. The purpose of this article is to provide a review of the current PET/CT technology, to project the future development of PET and CT for PET/CT, and to discuss some issues in adopting PET/CT in RT and potential improvements in PET/CT simulation of the thorax in radiation therapy.

  16. Evaluation of Multiple Immunoassay Technology Platforms to Select the Anti-Drug Antibody Assay Exhibiting the Most Appropriate Drug and Target Tolerance.

    PubMed

    Collet-Brose, Justine; Couble, Pierre-Jean; Deehan, Maureen R; Nelson, Robert J; Ferlin, Walter G; Lory, Sabrina

    2016-01-01

    The aim of this study was, at the assay development stage and thus with an appropriate degree of rigor, to select the most appropriate technology platform and sample pretreatment procedure for a clinical ADA assay. Thus, ELISA, MSD, Gyrolab, and AlphaLISA immunoassay platforms were evaluated in association with target depletion and acid dissociation sample pretreatment steps. An acid dissociation step successfully improved the drug tolerance for all 4 technology platforms and the required drug tolerance was achieved with the Gyrolab and MSD platforms. The target tolerance was shown to be better for the ELISA format, where an acid dissociation treatment step alone was sufficient to achieve the desired target tolerance. However, inclusion of a target depletion step in conjunction with the acid treatment raised the target tolerance to the desired level for all of the technologies. A higher sensitivity was observed for the MSD and Gyrolab assays and the ELISA, MSD, and Gyrolab all displayed acceptable interdonor variability. This study highlights the usefulness of evaluating the performance of different assay platforms at an early stage in the assay development process to aid in the selection of the best fit-for-purpose technology platform and sample pretreatment steps. PMID:27243038

  17. Evaluation of Multiple Immunoassay Technology Platforms to Select the Anti-Drug Antibody Assay Exhibiting the Most Appropriate Drug and Target Tolerance

    PubMed Central

    Collet-Brose, Justine

    2016-01-01

    The aim of this study was, at the assay development stage and thus with an appropriate degree of rigor, to select the most appropriate technology platform and sample pretreatment procedure for a clinical ADA assay. Thus, ELISA, MSD, Gyrolab, and AlphaLISA immunoassay platforms were evaluated in association with target depletion and acid dissociation sample pretreatment steps. An acid dissociation step successfully improved the drug tolerance for all 4 technology platforms and the required drug tolerance was achieved with the Gyrolab and MSD platforms. The target tolerance was shown to be better for the ELISA format, where an acid dissociation treatment step alone was sufficient to achieve the desired target tolerance. However, inclusion of a target depletion step in conjunction with the acid treatment raised the target tolerance to the desired level for all of the technologies. A higher sensitivity was observed for the MSD and Gyrolab assays and the ELISA, MSD, and Gyrolab all displayed acceptable interdonor variability. This study highlights the usefulness of evaluating the performance of different assay platforms at an early stage in the assay development process to aid in the selection of the best fit-for-purpose technology platform and sample pretreatment steps. PMID:27243038

  18. Rational design of a redox-labeled chiral target for an enantioselective aptamer-based electrochemical binding assay.

    PubMed

    Moreau, Julie; Challier, Lylian; Lalaoui, Noémie; Mavré, François; Noël, Vincent; Limoges, Benoît; Schöllhorn, Bernd; Fave, Claire

    2014-03-01

    A series of redox-labeled L-tyrosinamide (L-Tym) derivatives was prepared and the nature of the functional group and the chain length of the spacer were systematically varied in a step-by-step affinity optimization process of the tracer for the L-Tym aptamer. The choice of the labeling position on L-Tym proved to be crucial for the molecular recognition event, which could be monitored by cyclic voltammetry and is based on the different diffusion rates of free and bound targets in solution. From this screening approach an efficient electroactive tracer emerged. Comparable dissociation constants Kd were obtained for the unlabeled and labeled targets in direct or competitive binding assays. The enantiomeric tracer was prepared and its enantioselective recognition by the corresponding anti-D-Tym aptamer was demonstrated. The access to both enantiomeric tracer molecules opens the door for the development of one-pot determination of the enantiomeric excess when using different labels with well-separated redox potentials for each enantiomer. PMID:24519626

  19. Targeting CD146 with a 64Cu-labeled antibody enables in vivo immunoPET imaging of high-grade gliomas

    PubMed Central

    Yang, Yunan; Hernandez, Reinier; Rao, Jun; Yin, Li; Qu, Yazhuo; Wu, Jinrong; England, Christopher G.; Graves, Stephen A.; Lewis, Christina M.; Wang, Pu; Meyerand, Mary E.; Nickles, Robert J.; Bian, Xiu-wu; Cai, Weibo

    2015-01-01

    Given the highly heterogeneous character of brain malignancies and the associated implication for its proper diagnosis and treatment, finding biomarkers that better characterize this disease from a molecular standpoint is imperative. In this study, we evaluated CD146 as a potential molecular target for diagnosis and targeted therapy of glioblastoma multiforme (GBM), the most common and lethal brain malignancy. YY146, an anti-CD146 monoclonal antibody, was generated and radiolabeled for noninvasive positron-emission tomography (PET) imaging of orthotopic GBM models. 64Cu-labeled YY146 preferentially accumulated in the tumors of mice bearing U87MG xenografts, which allowed the acquisition of high-contrast PET images of small tumor nodules (∼2 mm). Additionally, we found that tumor uptake correlated with the levels of CD146 expression in a highly specific manner. We also explored the potential therapeutic effects of YY146 on the cancer stem cell (CSC) and epithelial-to-mesenchymal (EMT) properties of U87MG cells, demonstrating that YY146 can mitigate those aggressive phenotypes. Using YY146 as the primary antibody, we performed histological studies of World Health Organization (WHO) grades I through IV primary gliomas. The positive correlation found between CD146-positive staining and high tumor grade (χ2 = 9.028; P = 0.029) concurred with the GBM data available in The Cancer Genome Atlas (TCGA) and validated the clinical value of YY146. In addition, we demonstrate that YY146 can be used to detect CD146 in various cancer cell lines and human resected tumor tissues of multiple other tumor types (gastric, ovarian, liver, and lung), indicating a broad applicability of YY146 in solid tumors. PMID:26553993

  20. Targeting CD146 with a 64Cu-labeled antibody enables in vivo immunoPET imaging of high-grade gliomas.

    PubMed

    Yang, Yunan; Hernandez, Reinier; Rao, Jun; Yin, Li; Qu, Yazhuo; Wu, Jinrong; England, Christopher G; Graves, Stephen A; Lewis, Christina M; Wang, Pu; Meyerand, Mary E; Nickles, Robert J; Bian, Xiu-Wu; Cai, Weibo

    2015-11-24

    Given the highly heterogeneous character of brain malignancies and the associated implication for its proper diagnosis and treatment, finding biomarkers that better characterize this disease from a molecular standpoint is imperative. In this study, we evaluated CD146 as a potential molecular target for diagnosis and targeted therapy of glioblastoma multiforme (GBM), the most common and lethal brain malignancy. YY146, an anti-CD146 monoclonal antibody, was generated and radiolabeled for noninvasive positron-emission tomography (PET) imaging of orthotopic GBM models. (64)Cu-labeled YY146 preferentially accumulated in the tumors of mice bearing U87MG xenografts, which allowed the acquisition of high-contrast PET images of small tumor nodules (∼ 2 mm). Additionally, we found that tumor uptake correlated with the levels of CD146 expression in a highly specific manner. We also explored the potential therapeutic effects of YY146 on the cancer stem cell (CSC) and epithelial-to-mesenchymal (EMT) properties of U87MG cells, demonstrating that YY146 can mitigate those aggressive phenotypes. Using YY146 as the primary antibody, we performed histological studies of World Health Organization (WHO) grades I through IV primary gliomas. The positive correlation found between CD146-positive staining and high tumor grade (χ(2) = 9.028; P = 0.029) concurred with the GBM data available in The Cancer Genome Atlas (TCGA) and validated the clinical value of YY146. In addition, we demonstrate that YY146 can be used to detect CD146 in various cancer cell lines and human resected tumor tissues of multiple other tumor types (gastric, ovarian, liver, and lung), indicating a broad applicability of YY146 in solid tumors. PMID:26553993

  1. Large scale integration of drug-target information reveals poly-pharmacological drug action mechanisms in tumor cell line growth inhibition assays

    PubMed Central

    Knight, Richard A.; Gostev, Mikhail; Ilisavskii, Sergei; Willis, Anne E.; Melino, Gerry; Antonov, Alexey V.

    2014-01-01

    Understanding therapeutic mechanisms of drug anticancer cytotoxicity represents a key challenge in preclinical testing. Here we have performed a meta-analysis of publicly available tumor cell line growth inhibition assays (~ 70 assays from 6 independent experimental groups covering ~ 500 000 molecules) with the primary goal of understanding molecular therapeutic mechanisms of cancer cytotoxicity. To implement this we have collected currently available information on protein targets for molecules that were tested in the assays. We used a statistical methodology to identify protein targets overrepresented among molecules exhibiting cancer cytotoxicity with the particular focus of identifying overrepresented patterns consisting of several proteins (i.e. proteins “A” and “B” and “C”). Our analysis demonstrates that targeting individual proteins can result in a significant increase (up to 50-fold) of the observed odds for a molecule to be an efficient inhibitor of tumour cell line growth. However, further insight into potential molecular mechanisms reveals a multi-target mode of action: targeting a pattern of several proteins drastically increases the observed odds (up to 500-fold) for a molecule to be tumour cytotoxic. In contrast, molecules targeting only one protein but not targeting an additional set of proteins tend to be nontoxic. Our findings support a poly-pharmacology drug discovery paradigm, demonstrating that anticancer cytotoxicity is a product, in most cases, of multi-target mode of drug action PMID:24553133

  2. Development of PCR Assays Targeting Genes in O-Antigen Gene Clusters for Detection and Identification of Escherichia coli O45 and O55 Serogroups

    PubMed Central

    DebRoy, Chitrita; Fratamico, Pina M.; Roberts, Elisabeth; Davis, Michael A.; Liu, Yanhong

    2005-01-01

    The Escherichia coli O45 O-antigen gene cluster of strain O45:H2 96-3285 was sequenced, and conventional (singleplex), multiplex, and real-time PCR assays were designed to amplify regions in the wzx (O-antigen flippase) and wzy (O-antigen polymerase) genes. In addition, PCR assays targeting the E. coli O55 wzx and wzy genes were designed based on previously published sequences. PCR assays targeting E. coli O45 showed 100% specificity for this serogroup, whereas by PCR assays specific for E. coli O55, 97/102 strains serotyped as E. coli O55 were positive for wzx and 98/102 for wzy. Multiplex PCR assays targeting the E. coli O45 and the E. coli O55 wzx and wzy genes were used to detect the organisms in fecal samples spiked at levels of 106 and 108 CFU/0.2 g feces. Thus, the PCR assays can be used to detect and identify E. coli serogroups O45 and O55. PMID:16085897

  3. Topoisomerase Assays

    PubMed Central

    Nitiss, John L.; Soans, Eroica; Rogojina, Anna; Seth, Aman; Mishina, Margarita

    2012-01-01

    Topoisomerases are nuclear enzymes that play essential roles in DNA replication, transcription, chromosome segregation, and recombination. All cells have two major forms of topoisomerases: type I, which makes single-stranded cuts in DNA, and type II enzymes, which cut and pass double-stranded DNA. DNA topoisomerases are important targets of approved and experimental anti-cancer agents. The protocols described in this unit are of assays used to assess new chemical entities for their ability to inhibit both forms of DNA topoisomerase. Included are an in vitro assay for topoisomerase I activity based on relaxation of supercoiled DNA and an assay for topoisomerase II based on the decatenation of double-stranded DNA. The preparation of mammalian cell extracts for assaying topoisomerase activity is described, along with a protocol for an ICE assay for examining topoisomerase covalent complexes in vivo and an assay for measuring DNA cleavage in vitro. PMID:22684721

  4. Comparison of gull-specific assays targeting 16S rRNA gene of Catellicoccus marimammalium and Streptococcus spp.

    EPA Science Inventory

    Gulls have been implicated as a source of fecal contamination in inland and coastal waters. Only one gull-specific assay is currently available (i.e., gull2 qPCR assay). This assay is based on the 16S rRNA gene of Catellicocclls marimammalium and has showed a high level of host-s...

  5. Pitfalls of the MTT assay: Direct and off-target effects of inhibitors can result in over/underestimation of cell viability.

    PubMed

    Stepanenko, A A; Dmitrenko, V V

    2015-12-15

    The MTT assay (to a less degree MTS, XTT or WST) is a widely exploited approach for measuring cell viability/drug cytotoxicity. MTT reduction occurs throughout a cell and can be significantly affected by a number of factors, including metabolic and energy perturbations, changes in the activity of oxidoreductases, endo-/exocytosis and intracellular trafficking. Over/underestimation of cell viability by the MTT assay may be due to both adaptive metabolic and mitochondrial reprogramming of cells subjected to drug treatment-mediated stress and inhibitor off-target effects. Previously, imatinib, rottlerin, ursolic acid, verapamil, resveratrol, genistein nanoparticles and some polypeptides were shown to interfere with MTT reduction rate resulting in inconsistent results between the MTT assay and alternative assays. Here, to test the under/overestimation of viability by the MTT assay, we compared results derived from the MTT assay with the trypan blue exclusion assay after treatment of glioblastoma U251, T98G and C6 cells with three widely used inhibitors with the known direct and side effects on energy and metabolic homeostasis - temozolomide (TMZ), a DNA-methylating agent, temsirolimus (TEM), an inhibitor of mTOR kinase, and U0126, an inhibitor of MEK1/2 kinases. Inhibitors were applied shortly as in IC50 evaluating studies or long as in studies focusing on drug resistance acquisition. We showed that over/underestimation of cell viability by the MTT assay and its significance depends on a cell line, a time point of viability measurement and other experimental parameters. Furthermore, we provided a comprehensive survey of factors that should be accounted in the MTT assay. To avoid result misinterpretation, supplementation of the tetrazolium salt-based assays with other non-metabolic assays is recommended. PMID:26260013

  6. Assessment of a targeted resequencing assay as a support tool in the diagnosis of lysosomal storage disorders

    PubMed Central

    2014-01-01

    Background With over 50 different disorders and a combined incidence of up to 1/3000 births, lysosomal storage diseases (LSDs) constitute a major public health problem and place an enormous burden on affected individuals and their families. Many factors make LSD diagnosis difficult, including phenotype and penetrance variability, shared signs and symptoms, and problems inherent to biochemical diagnosis. Developing a powerful diagnostic tool could mitigate the protracted diagnostic process for these families, lead to better outcomes for current and proposed therapies, and provide the basis for more appropriate genetic counseling. Methods We have designed a targeted resequencing assay for the simultaneous testing of 57 lysosomal genes, using in-solution capture as the enrichment method and two different sequencing platforms. A total of 84 patients with high to moderate-or low suspicion index for LSD were enrolled in different centers in Spain and Portugal, including 18 positive controls. Results We correctly diagnosed 18 positive blinded controls, provided genetic diagnosis to 25 potential LSD patients, and ended with 18 diagnostic odysseys. Conclusion We report the assessment of a next–generation-sequencing-based approach as an accessory tool in the diagnosis of LSDs, a group of disorders which have overlapping clinical profiles and genetic heterogeneity. We have also identified and quantified the strengths and limitations of next generation sequencing (NGS) technology applied to diagnosis. PMID:24767253

  7. Development and Validation of an Immuno-PET Tracer as a Companion Diagnostic Agent for Antibody-Drug Conjugate Therapy to Target the CA6 Epitope

    PubMed Central

    Ilovich, Ohad; Natarajan, Arutselvan; Hori, Sharon; Sathirachinda, Ataya; Kimura, Richard; Srinivasan, Ananth; Gebauer, Mathias; Kruip, Jochen; Focken, Ingo; Lange, Christian; Carrez, Chantal; Sassoon, Ingrid; Blanc, Veronique; Sarkar, Susanta K.

    2015-01-01

    Purpose To develop and compare three copper 64 (64Cu)–labeled antibody fragments derived from a CA6-targeting antibody (huDS6) as immuno-positron emission tomography (immuno-PET)–based companion diagnostic agents for an antibody-drug conjugate by using huDS6. Materials and Methods Three antibody fragments derived from huDS6 were produced, purified, conjugated to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), and evaluated in the following ways: (a) the affinity of the fragments and the DOTA conjugates was measured via flow cytometry, (b) the stability of the labeled fragments was determined ex vivo in human serum over 24 hours, and (c) comparison of the in vivo imaging potential of the fragments was evaluated in mice bearing subcutaneous CA6-positive and CA6-negative xenografts by using serial PET imaging and biodistribution. Isotype controls with antilysozyme and anti-DM4 B-Fabs and blocking experiments with an excess of either B-Fab or huDS6 were used to determine the extent of the antibody fragment 64Cu-DOTA-B-Fab binding specificity. Immunoreactivity and tracer kinetics were evaluated by using cellular uptake and 48-hour imaging experiments, respectively. Statistical analyses were performed by using t tests, one-way analysis of variance, and Wilcoxon and Mann-Whitney tests. Results The antibody fragment 64Cu-DOTA-B-Fab was more than 95% stable after 24 hours in human serum, had an immunoreactivity of more than 70%, and allowed differentiation between CA6-positive and CA6-negative tumors in vivo as early as 6 hours after injection, with a 1.7-fold uptake ratio between tumors. Isotype and blocking studies experiments showed tracer-specific uptake in antigen-positive tumors, despite some nonspecific uptake in both tumor models. Conclusion Three antibody fragments were produced and examined as potential companion diagnostic agents. 64Cu-DOTA-B-Fab is a stable and effective immuno-PET tracer for CA6 imaging in vivo. © RSNA, 2015 Online

  8. Lutetium oxyorthosilicate (LSO) intrinsic activity correction and minimal detectable target activity study for SPECT imaging with a LSO-based animal PET scanner.

    PubMed

    Yao, Rutao; Ma, Tianyu; Shao, Yiping

    2008-08-21

    This work is part of a feasibility study to develop SPECT imaging capability on a lutetium oxyorthosilicate (LSO) based animal PET system. The SPECT acquisition was enabled by inserting a collimator assembly inside the detector ring and acquiring data in singles mode. The same LSO detectors were used for both PET and SPECT imaging. The intrinsic radioactivity of (176)Lu in the LSO crystals, however, contaminates the SPECT data, and can generate image artifacts and introduce quantification error. The objectives of this study were to evaluate the effectiveness of a LSO background subtraction method, and to estimate the minimal detectable target activity (MDTA) of image object for SPECT imaging. For LSO background correction, the LSO contribution in an image study was estimated based on a pre-measured long LSO background scan and subtracted prior to the image reconstruction. The MDTA was estimated in two ways. The empirical MDTA (eMDTA) was estimated from screening the tomographic images at different activity levels. The calculated MDTA (cMDTA) was estimated from using a formula based on applying a modified Currie equation on an average projection dataset. Two simulated and two experimental phantoms with different object activity distributions and levels were used in this study. The results showed that LSO background adds concentric ring artifacts to the reconstructed image, and the simple subtraction method can effectively remove these artifacts-the effect of the correction was more visible when the object activity level was near or above the eMDTA. For the four phantoms studied, the cMDTA was consistently about five times of the corresponding eMDTA. In summary, we implemented a simple LSO background subtraction method and demonstrated its effectiveness. The projection-based calculation formula yielded MDTA results that closely correlate with that obtained empirically and may have predicative value for imaging applications. PMID:18670052

  9. Lutetium oxyorthosilicate (LSO) intrinsic activity correction and minimal detectable target activity study for SPECT imaging with a LSO-based animal PET scanner

    NASA Astrophysics Data System (ADS)

    Yao, Rutao; Ma, Tianyu; Shao, Yiping

    2008-08-01

    This work is part of a feasibility study to develop SPECT imaging capability on a lutetium oxyorthosilicate (LSO) based animal PET system. The SPECT acquisition was enabled by inserting a collimator assembly inside the detector ring and acquiring data in singles mode. The same LSO detectors were used for both PET and SPECT imaging. The intrinsic radioactivity of 176Lu in the LSO crystals, however, contaminates the SPECT data, and can generate image artifacts and introduce quantification error. The objectives of this study were to evaluate the effectiveness of a LSO background subtraction method, and to estimate the minimal detectable target activity (MDTA) of image object for SPECT imaging. For LSO background correction, the LSO contribution in an image study was estimated based on a pre-measured long LSO background scan and subtracted prior to the image reconstruction. The MDTA was estimated in two ways. The empirical MDTA (eMDTA) was estimated from screening the tomographic images at different activity levels. The calculated MDTA (cMDTA) was estimated from using a formula based on applying a modified Currie equation on an average projection dataset. Two simulated and two experimental phantoms with different object activity distributions and levels were used in this study. The results showed that LSO background adds concentric ring artifacts to the reconstructed image, and the simple subtraction method can effectively remove these artifacts—the effect of the correction was more visible when the object activity level was near or above the eMDTA. For the four phantoms studied, the cMDTA was consistently about five times of the corresponding eMDTA. In summary, we implemented a simple LSO background subtraction method and demonstrated its effectiveness. The projection-based calculation formula yielded MDTA results that closely correlate with that obtained empirically and may have predicative value for imaging applications.

  10. Structure-Activity Relationship of (18)F-Labeled Phosphoramidate Peptidomimetic Prostate-Specific Membrane Antigen (PSMA)-Targeted Inhibitor Analogues for PET Imaging of Prostate Cancer.

    PubMed

    Dannoon, Shorouk; Ganguly, Tanushree; Cahaya, Hendry; Geruntho, Jonathan J; Galliher, Matthew S; Beyer, Sophia K; Choy, Cindy J; Hopkins, Mark R; Regan, Melanie; Blecha, Joseph E; Skultetyova, Lubica; Drake, Christopher R; Jivan, Salma; Barinka, Cyril; Jones, Ella F; Berkman, Clifford E; VanBrocklin, Henry F

    2016-06-23

    A series of phosphoramidate-based prostate specific membrane antigen (PSMA) inhibitors of increasing lipophilicity were synthesized (4, 5, and 6), and their fluorine-18 analogs were evaluated for use as positron emission tomography (PET) imaging agents for prostate cancer. To gain insight into their modes of binding, they were also cocrystallized with the extracellular domain of PSMA. All analogs exhibited irreversible binding to PSMA with IC50 values ranging from 0.4 to 1.3 nM. In vitro assays showed binding and rapid internalization (80-95%, 2 h) of the radiolabeled ligands in PSMA(+) cells. In vivo distribution demonstrated significant uptake in CWR22Rv1 (PSMA(+)) tumor, with tumor to blood ratios of 25.6:1, 63.6:1, and 69.6:1 for [(18)F]4, [(18)F]5, and [(18)F]6, respectively, at 2 h postinjection. Installation of aminohexanoic acid (AH) linkers in the phosphoramidate scaffold improved their PSMA binding and inhibition and was critical for achieving suitable in vivo imaging properties, positioning [(18)F]5 and [(18)F]6 as favorable candidates for future prostate cancer imaging clinical trials. PMID:27228467

  11. A convenient cellular assay for the identification of the molecular target of ergosterol biosynthesis inhibitors and quantification of their effects on total ergosterol biosynthesis.

    PubMed

    Müller, Christoph; Staudacher, Verena; Krauss, Jürgen; Giera, Martin; Bracher, Franz

    2013-05-01

    Increasing resistance of clinically relevant fungi is causing major problems in anti-mycotic therapy. Particularly for immunosuppressed patients fungal infections are of concern and increasing resistance against clinically used antimycotic drugs is hampering successful treatment. In the search for new antifungals ergosterol biosynthesis still is the most prominent target. However, several pitfalls in the bioactivity testing of such substances remain. Two of the major drawbacks certainly are the membrane association of most enzymes participating in ergosterol biosynthesis, and the difficulty to selectively associate growth inhibitory effects with the target pathway (ergosterol biosynthesis). Here we describe a GC-MS based cellular assay for target identification and selective potency determination of test components. In the qualitative part of the assay GC-MS analysis of cell lysates allows target identification by analysis of the changes in the sterol pattern. The quantitative part of the assay makes use of 13C-acetate feeding combined with GC-MS analysis allowing the selective quantification of a compound's effect on total ergosterol biosynthesis. The described cellular assay was analytically and biologically validated and used to characterize the novel ergosterol biosynthesis inhibitor JK-250. PMID:23454215

  12. Combining multiple FDG-PET radiotherapy target segmentation methods to reduce the effect of variable performance of individual segmentation methods

    SciTech Connect

    McGurk, Ross J.; Bowsher, James; Das, Shiva K.; Lee, John A

    2013-04-15

    Purpose: Many approaches have been proposed to segment high uptake objects in 18F-fluoro-deoxy-glucose positron emission tomography images but none provides consistent performance across the large variety of imaging situations. This study investigates the use of two methods of combining individual segmentation methods to reduce the impact of inconsistent performance of the individual methods: simple majority voting and probabilistic estimation. Methods: The National Electrical Manufacturers Association image quality phantom containing five glass spheres with diameters 13-37 mm and two irregularly shaped volumes (16 and 32 cc) formed by deforming high-density polyethylene bottles in a hot water bath were filled with 18-fluoro-deoxyglucose and iodine contrast agent. Repeated 5-min positron emission tomography (PET) images were acquired at 4:1 and 8:1 object-to-background contrasts for spherical objects and 4.5:1 and 9:1 for irregular objects. Five individual methods were used to segment each object: 40% thresholding, adaptive thresholding, k-means clustering, seeded region-growing, and a gradient based method. Volumes were combined using a majority vote (MJV) or Simultaneous Truth And Performance Level Estimate (STAPLE) method. Accuracy of segmentations relative to CT ground truth volumes were assessed using the Dice similarity coefficient (DSC) and the symmetric mean absolute surface distances (SMASDs). Results: MJV had median DSC values of 0.886 and 0.875; and SMASD of 0.52 and 0.71 mm for spheres and irregular shapes, respectively. STAPLE provided similar results with median DSC of 0.886 and 0.871; and median SMASD of 0.50 and 0.72 mm for spheres and irregular shapes, respectively. STAPLE had significantly higher DSC and lower SMASD values than MJV for spheres (DSC, p < 0.0001; SMASD, p= 0.0101) but MJV had significantly higher DSC and lower SMASD values compared to STAPLE for irregular shapes (DSC, p < 0.0001; SMASD, p= 0.0027). DSC was not significantly

  13. Assay of the Rab-binding specificity of rabphilin and Noc2: target molecules for Rab27.

    PubMed

    Fukuda, Mitsunori; Yamamoto, Akitsugu

    2005-01-01

    Rabphilin and Noc2 were originally described as Rab3A effector proteins involved in the regulation of secretory vesicle exocytosis in neurons and certain endocrine cells. Both proteins share the conserved N-terminal Rab-binding domain (RBD) that consists of two alpha-helical regions separated by two zinc finger motifs. However, the RBD of rabphilin and Noc2 has been shown to bind Rab27A (the closest homologue of Rab3 isoforms) in preference to Rab3A, both in vitro and in vivo. Rabphilin and Noc2 are recruited to dense-core vesicles (DCVs) in neuroendocrine PC12 cells and regulate their exocytosis through interaction with Rab27A rather than with Rab3A. Rab3A-binding-defective mutants of rabphilin(E50A) and Noc2(E51A) retain the ability to target DCVs in PC12 cells, the same as the wild-type proteins, whereas Rab27A-binding-defective mutants of rabphilin(E50A/I54A) and Noc2(E51A/I55A) do not (i.e., they are present throughout the cytoplasm). Expression of the wild-type or the E50A mutant of rabphilin-RBD, but not the E50A/I54A mutant of rabphilin-RBD, in PC12 cells significantly attenuated DCV exocytosis monitored by high-KCl-stimulated neuropeptide Y secretion. In this chapter we describe various assay methods that have been used to characterize the RBD of rabphilin and Noc2 as "RBD27 (Rab-binding domain for Rab27)." PMID:16473612

  14. Planar optical waveguide based sandwich assay sensors and processes for the detection of biological targets including protein markers, pathogens and cellular debris

    DOEpatents

    Martinez, Jennifer S.; Swanson, Basil I.; Grace, Karen M.; Grace, Wynne K.; Shreve, Andrew P.

    2009-06-02

    An assay element is described including recognition ligands bound to a film on a single mode planar optical waveguide, the film from the group of a membrane, a polymerized bilayer membrane, and a self-assembled monolayer containing polyethylene glycol or polypropylene glycol groups therein and an assay process for detecting the presence of a biological target is described including injecting a biological target-containing sample into a sensor cell including the assay element, with the recognition ligands adapted for binding to selected biological targets, maintaining the sample within the sensor cell for time sufficient for binding to occur between selected biological targets within the sample and the recognition ligands, injecting a solution including a reporter ligand into the sensor cell; and, interrogating the sample within the sensor cell with excitation light from the waveguide, the excitation light provided by an evanescent field of the single mode penetrating into the biological target-containing sample to a distance of less than about 200 nanometers from the waveguide thereby exciting the fluorescent-label in any bound reporter ligand within a distance of less than about 200 nanometers from the waveguide and resulting in a detectable signal.

  15. PET scan

    MedlinePlus

    You may feel a sharp sting when the needle with the tracer is placed into your vein. A PET scan causes no pain. The table may be ... The amount of radiation used in a PET scan is about the same amount as used in most CT scans. These scans use ...

  16. Synthesis, radiolabeling and preliminary in vivo evaluation of multimodal radiotracers for PET imaging and targeted radionuclide therapy of pigmented melanoma.

    PubMed

    Billaud, Emilie M F; Maisonial-Besset, Aurélie; Rbah-Vidal, Latifa; Vidal, Aurélien; Besse, Sophie; Béquignat, Jean-Baptiste; Decombat, Caroline; Degoul, Françoise; Audin, Laurent; Deloye, Jean-Bernard; Dollé, Frédéric; Kuhnast, Bertrand; Madelmont, Jean-Claude; Tarrit, Sébastien; Galmier, Marie-Josèphe; Borel, Michèle; Auzeloux, Philippe; Miot-Noirault, Elisabeth; Chezal, Jean-Michel

    2015-03-01

    Melanin pigment represents an attractive target to address specific treatment to melanoma cells, such as cytotoxic radionuclides. However, less than half of the patients have pigmented metastases. Hence, specific marker is required to stratify this patient population before proceeding with melanin-targeted radionuclide therapy. In such a context, we developed fluorinated analogues of a previously studied melanin-targeting ligand, N-(2-diethylaminoethyl)-6-iodoquinoxaline-2-carboxamide (ICF01012). These latter can be labeled either with (18)F or (131)I/(125)I for positron emission tomography imaging (melanin-positive patient selection) and targeted radionuclide therapy purposes. Here we describe the syntheses, radiosyntheses and preclinical evaluations on melanoma-bearing mice model of several iodo- and fluoro(hetero)aromatic derivatives of the ICF01012 scaffold. After preliminary planar gamma scintigraphic and positron emission tomography imaging evaluations, [(125)I]- and [(18)F]-N-[2-(diethylamino)ethyl]-4-fluoro-3-iodobenzamides ([(125)I]4, [(18)F]4) were found to be chemically and biologically stable with quite similar tumor uptakes at 1 h p.i. (9.7 ± 2.6% ID/g and 6.8 ± 1.9% ID/g, respectively). PMID:25637883

  17. Prediction of Multi-Target Networks of Neuroprotective Compounds with Entropy Indices and Synthesis, Assay, and Theoretical Study of New Asymmetric 1,2-Rasagiline Carbamates

    PubMed Central

    Romero Durán, Francisco J.; Alonso, Nerea; Caamaño, Olga; García-Mera, Xerardo; Yañez, Matilde; Prado-Prado, Francisco J.; González-Díaz, Humberto

    2014-01-01

    In a multi-target complex network, the links (Lij) represent the interactions between the drug (di) and the target (tj), characterized by different experimental measures (Ki, Km, IC50, etc.) obtained in pharmacological assays under diverse boundary conditions (cj). In this work, we handle Shannon entropy measures for developing a model encompassing a multi-target network of neuroprotective/neurotoxic compounds reported in the CHEMBL database. The model predicts correctly >8300 experimental outcomes with Accuracy, Specificity, and Sensitivity above 80%–90% on training and external validation series. Indeed, the model can calculate different outcomes for >30 experimental measures in >400 different experimental protocolsin relation with >150 molecular and cellular targets on 11 different organisms (including human). Hereafter, we reported by the first time the synthesis, characterization, and experimental assays of a new series of chiral 1,2-rasagiline carbamate derivatives not reported in previous works. The experimental tests included: (1) assay in absence of neurotoxic agents; (2) in the presence of glutamate; and (3) in the presence of H2O2. Lastly, we used the new Assessing Links with Moving Averages (ALMA)-entropy model to predict possible outcomes for the new compounds in a high number of pharmacological tests not carried out experimentally. PMID:25255029

  18. Integration of Affinity Selection-Mass Spectrometry and Functional Cell-Based Assays to Rapidly Triage Druggable Target Space within the NF-κB Pathway.

    PubMed

    Kutilek, Victoria D; Andrews, Christine L; Richards, Matthew P; Xu, Zangwei; Sun, Tianxiao; Chen, Yiping; Hashke, Andrew; Smotrov, Nadya; Fernandez, Rafael; Nickbarg, Elliott B; Chamberlin, Chad; Sauvagnat, Berengere; Curran, Patrick J; Boinay, Ryan; Saradjian, Peter; Allen, Samantha J; Byrne, Noel; Elsen, Nathaniel L; Ford, Rachael E; Hall, Dawn L; Kornienko, Maria; Rickert, Keith W; Sharma, Sujata; Shipman, Jennifer M; Lumb, Kevin J; Coleman, Kevin; Dandliker, Peter J; Kariv, Ilona; Beutel, Bruce

    2016-07-01

    The primary objective of early drug discovery is to associate druggable target space with a desired phenotype. The inability to efficiently associate these often leads to failure early in the drug discovery process. In this proof-of-concept study, the most tractable starting points for drug discovery within the NF-κB pathway model system were identified by integrating affinity selection-mass spectrometry (AS-MS) with functional cellular assays. The AS-MS platform Automated Ligand Identification System (ALIS) was used to rapidly screen 15 NF-κB proteins in parallel against large-compound libraries. ALIS identified 382 target-selective compounds binding to 14 of the 15 proteins. Without any chemical optimization, 22 of the 382 target-selective compounds exhibited a cellular phenotype consistent with the respective target associated in ALIS. Further studies on structurally related compounds distinguished two chemical series that exhibited a preliminary structure-activity relationship and confirmed target-driven cellular activity to NF-κB1/p105 and TRAF5, respectively. These two series represent new drug discovery opportunities for chemical optimization. The results described herein demonstrate the power of combining ALIS with cell functional assays in a high-throughput, target-based approach to determine the most tractable drug discovery opportunities within a pathway. PMID:26969322

  19. Lung PET scan

    MedlinePlus

    ... emission tomography; PET - chest; PET - lung; PET - tumor imaging ... Grainger & Allison's Diagnostic Radiology: A Textbook of Medical Imaging . 6th ed. Philadelphia, PA: Elsevier Churchill Livingstone; 2015: ...

  20. Access to a polymerase chain reaction assay method targeting 13 respiratory viruses can reduce antibiotics: a randomised, controlled trial

    PubMed Central

    2011-01-01

    Background Viral respiratory infections are common worldwide and range from completely benign disease to life-threatening illness. Symptoms can be unspecific, and an etiologic diagnosis is rarely established because of a lack of suitable diagnostic tools. Improper use of antibiotics is common in this setting, which is detrimental in light of the development of bacterial resistance. It has been suggested that the use of diagnostic tests could reduce antibiotic prescription rates. The objective of this study was to evaluate whether access to a multiplex polymerase chain reaction (PCR) assay panel for etiologic diagnosis of acute respiratory tract infections (ARTIs) would have an impact on antibiotic prescription rate in primary care clinical settings. Methods Adult patients with symptoms of ARTI were prospectively included. Nasopharyngeal and throat swabs were analysed by using a multiplex real-time PCR method targeting thirteen viruses and two bacteria. Patients were recruited at 12 outpatient units from October 2006 through April 2009, and samples were collected on the day of inclusion (initial visit) and after 10 days (follow-up visit). Patients were randomised in an open-label treatment protocol to receive a rapid or delayed result (on the following day or after eight to twelve days). The primary outcome measure was the antibiotic prescription rate at the initial visit, and the secondary outcome was the total antibiotic prescription rate during the study period. Results A total sample of 447 patients was randomised. Forty-one were excluded, leaving 406 patients for analysis. In the group of patients randomised for a rapid result, 4.5% (9 of 202) of patients received antibiotics at the initial visit, compared to 12.3% (25 of 204) (P = 0.005) of patients in the delayed result group. At follow-up, there was no significant difference between the groups: 13.9% (28 of 202) in the rapid result group and 17.2% (35 of 204) in the delayed result group (P = 0

  1. A new fluorescence/PET probe for targeting intracellular human telomerase reverse transcriptase (hTERT) using Tat peptide-conjugated IgM.

    PubMed

    Jung, Kyung Oh; Youn, Hyewon; Kim, Seung Hoo; Kim, Young-Hwa; Kang, Keon Wook; Chung, June-Key

    2016-08-26

    Despite an increasing need for methods to visualize intracellular proteins in vivo, the majority of antibody-based imaging methods available can only detect membrane proteins. The human telomerase reverse transcriptase (hTERT) is an intracellular target of great interest because of its high expression in several types of cancer. In this study, we developed a new probe for hTERT using the Tat peptide. An hTERT antibody (IgG or IgM) was conjugated with the Tat peptide, a fluorescence dye and (64)Cu. HT29 (hTERT+) and U2OS (hTERT-) were used to visualize the intracellular hTERT. The hTERT was detected by RT-PCR and western blot. Fluorescence signals for hTERT were obtained by confocal microscopy, live cell imaging, and analyzed by Tissue-FAXS. In nude mice, tumors were visualized using the fluorescence imaging devices Maestro™ and PETBOX. In RT-PCR and western blot, the expression of hTERT was detected in HT29 cells, but not in U2OS cells. Fluorescence signals were clearly observed in HT29 cells and in U2OS cells after 1 h of treatment, but signals were only detected in HT29 cells after 24 h. Confocal microscopy showed that 9.65% of U2OS and 78.54% of HT29 cells had positive hTERT signals. 3D animation images showed that the probe could target intranuclear hTERT in the nucleus. In mice models, fluorescence and PET imaging showed that hTERT in HT29 tumors could be efficiently visualized. In summary, we developed a new method to visualize intracellular and intranuclear proteins both in vitro and in vivo. PMID:27317485

  2. Interactions between the budding yeast IQGAP homologue Iqg1p and its targets revealed by a split-EGFP bimolecular fluorescence complementation assay.

    PubMed

    Pathmanathan, Sevvel; Barnard, Emma; Timson, David J

    2008-10-01

    A split-EGFP based bimolecular fluorescence complementation (BiFC) assay has been used to detect interactions between the Saccharomyces cerevisiae cytoskeletal scaffolding protein Iqg1p and three targets: myosin essential light chain (Mlc1p), calmodulin (Cmd1p) and the small GTPase Cdc42p. The format of the BiFC assay used ensures that the proteins are expressed at wild type levels thereby avoiding artefacts due to overexpression. This is the first direct in vivo detection of these interactions; in each case, the complex is localised to discrete regions of the yeast cytoplasm. The labelling with EGFP fragments results in changes in growth kinetics, cell size and budding frequency. This is partly due to the reassembled EGFP locking the complexes into essentially permanent interactions. The consequences of this for Iqg1p interactions and BiFC assays in general are discussed. PMID:18675924

  3. Quantitative assays for the measurement of HER1-HER2 heterodimerization and phosphorylation in cell lines and breast tumors: applications for diagnostics and targeted drug mechanism of action

    PubMed Central

    2011-01-01

    Introduction Ligand-bound and phosphorylated ErbB/HER heterodimers are potent signaling forms of this receptor family, and quantitative measurements of these active receptors may be predictive of patient response to targeted therapies. Using VeraTag™ technology, we developed and characterized quantitative assays measuring epidermal growth factor (EGF)-dependent increases in activated HER receptors in tumor cell line lysates and formalin-fixed, paraffin-embedded (FFPE) tumor sections. We demonstrated the ability of the assays to quantitatively measure changes in activated HER1 and HER2 receptor levels in cell lines following treatment with 2C4, erlotinib, and lapatinib. We utilized these assays to determine the prevalence and distribution of activated HER1, HER2, and HER1-HER2 heterodimers in 43 HER2-positive breast tumors. Methods Assays for activated HER1 and HER2 receptors in FFPE and cell lysate formats were developed using VeraTag™ technology, which requires the proximity of an antibody pair for light-dependent release of a fluorescently labeled tag, followed by capillary electrophoresis-based quantitation. Results Ligand-dependent and independent HER1-HER2 heterodimer levels measured by lysate and FFPE VeraTag™ assays trended with HER1 and HER2 expression levels in tumor cell lines, which was confirmed by co-immunoprecipitation. The formation of EGF-dependent HER1-HER2 heterodimers were inhibited by the HER2-targeted monoclonal antibody 2C4 and stabilized by the HER1 tyrosine kinase inhibitor (TKI) erlotinib. EGF-dependent HER1 and HER2 phosphorylation was inhibited by lapatinib and erlotinib. Further, we observed that dominant receptor signaling patterns may switch between HER1-HER1 and HER1-HER2, depending on drug mechanism of action and relative levels of HER receptors. In FFPE breast tumors that expressed both HER1 and HER2, HER1-HER2 heterodimers were detected in 25 to 50% of tumors, depending on detection method. The levels of activated phospho

  4. Double Gene Targeting Multiplex Polymerase Chain Reaction-Restriction Fragment Length Polymorphism Assay Discriminates Beef, Buffalo, and Pork Substitution in Frankfurter Products.

    PubMed

    Hossain, M A Motalib; Ali, Md Eaqub; Abd Hamid, Sharifah Bee; Asing; Mustafa, Shuhaimi; Mohd Desa, Mohd Nasir; Zaidul, I S M

    2016-08-17

    Beef, buffalo, and pork adulteration in the food chain is an emerging and sensitive issue. Current molecular techniques to authenticate these species depend on polymerase chain reaction (PCR) assays involving long and single targets which break down under natural decomposition and/or processing treatments. This novel multiplex polymerase chain reaction-restriction fragment length polymorphism assay targeted two different gene sites for each of the bovine, buffalo, and porcine materials. This authentication ensured better security, first through a complementation approach because it is highly unlikely that both sites will be missing under compromised states, and second through molecular fingerprints. Mitochondrial cytochrome b and ND5 genes were targeted, and all targets (73, 90, 106, 120, 138, and 146 bp) were stable under extreme boiling and autoclaving treatments. Target specificity and authenticity were ensured through cross-amplification reaction and restriction digestion of PCR products with AluI, EciI, FatI, and CviKI-1 enzymes. A survey of Malaysian frankfurter products revealed rampant substitution of beef with buffalo but purity in porcine materials. PMID:27501408

  5. Preclinical TSPO Ligand PET to Visualize Human Glioma Xenotransplants: A Preliminary Study

    PubMed Central

    Buck, Jason R.; McKinley, Eliot T.; Fu, Allie; Abel, Ty W.; Thompson, Reid C.; Chambless, Lola; Watchmaker, Jennifer M.; Harty, James P.; Cooper, Michael K.; Manning, H. Charles

    2015-01-01

    Current positron emission tomography (PET) imaging biomarkers for detection of infiltrating gliomas are limited. Translocator protein (TSPO) is a novel and promising biomarker for glioma PET imaging. To validate TSPO as a potential target for molecular imaging of glioma, TSPO expression was assayed in a tumor microarray containing 37 high-grade (III, IV) gliomas. TSPO staining was detected in all tumor specimens. Subsequently, PET imaging was performed with an aryloxyanilide-based TSPO ligand, [18F]PBR06, in primary orthotopic xenograft models of WHO grade III and IV gliomas. Selective uptake of [18F]PBR06 in engrafted tumor was measured. Furthermore, PET imaging with [18F]PBR06 demonstrated infiltrative glioma growth that was undetectable by traditional magnetic resonance imaging (MRI). Preliminary PET with [18F]PBR06 demonstrated a preferential tumor-to-normal background ratio in comparison to 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG). These results suggest that TSPO PET imaging with such high-affinity radiotracers may represent a novel strategy to characterize distinct molecular features of glioma growth, as well as better define the extent of glioma infiltration for therapeutic purposes. PMID:26517124

  6. Preclinical TSPO Ligand PET to Visualize Human Glioma Xenotransplants: A Preliminary Study.

    PubMed

    Buck, Jason R; McKinley, Eliot T; Fu, Allie; Abel, Ty W; Thompson, Reid C; Chambless, Lola; Watchmaker, Jennifer M; Harty, James P; Cooper, Michael K; Manning, H Charles

    2015-01-01

    Current positron emission tomography (PET) imaging biomarkers for detection of infiltrating gliomas are limited. Translocator protein (TSPO) is a novel and promising biomarker for glioma PET imaging. To validate TSPO as a potential target for molecular imaging of glioma, TSPO expression was assayed in a tumor microarray containing 37 high-grade (III, IV) gliomas. TSPO staining was detected in all tumor specimens. Subsequently, PET imaging was performed with an aryloxyanilide-based TSPO ligand, [18F]PBR06, in primary orthotopic xenograft models of WHO grade III and IV gliomas. Selective uptake of [18F]PBR06 in engrafted tumor was measured. Furthermore, PET imaging with [18F]PBR06 demonstrated infiltrative glioma growth that was undetectable by traditional magnetic resonance imaging (MRI). Preliminary PET with [18F]PBR06 demonstrated a preferential tumor-to-normal background ratio in comparison to 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG). These results suggest that TSPO PET imaging with such high-affinity radiotracers may represent a novel strategy to characterize distinct molecular features of glioma growth, as well as better define the extent of glioma infiltration for therapeutic purposes. PMID:26517124

  7. Positron Emission Tomography (PET)

    SciTech Connect

    Welch, M.J.

    1990-01-01

    Positron emission tomography (PET) assesses biochemical processes in the living subject, producing images of function rather than form. Using PET, physicians are able to obtain not the anatomical information provided by other medical imaging techniques, but pictures of physiological activity. In metaphoric terms, traditional imaging methods supply a map of the body's roadways, its, anatomy; PET shows the traffic along those paths, its biochemistry. This document discusses the principles of PET, the radiopharmaceuticals in PET, PET research, clinical applications of PET, the cost of PET, training of individuals for PET, the role of the United States Department of Energy in PET, and the futures of PET. 22 figs.

  8. Positron Emission Tomography (PET)

    DOE R&D Accomplishments Database

    Welch, M. J.

    1990-01-01

    Positron emission tomography (PET) assesses biochemical processes in the living subject, producing images of function rather than form. Using PET, physicians are able to obtain not the anatomical information provided by other medical imaging techniques, but pictures of physiological activity. In metaphoric terms, traditional imaging methods supply a map of the body's roadways, its, anatomy; PET shows the traffic along those paths, its biochemistry. This document discusses the principles of PET, the radiopharmaceuticals in PET, PET research, clinical applications of PET, the cost of PET, training of individuals for PET, the role of the United States Department of Energy in PET, and the futures of PET.

  9. Validation of Bacteroidales quantitative PCR assays targeting human and animal fecal contamination in the public and domestic domains in India.

    PubMed

    Odagiri, Mitsunori; Schriewer, Alexander; Hanley, Kaitlyn; Wuertz, Stefan; Misra, Pravas R; Panigrahi, Pinaki; Jenkins, Marion W

    2015-01-01

    We compared host-associated Bacteroidales qPCR assays developed in the continental United States and Europe for the purpose of measuring the effect of improved sanitation on human fecal exposure in rural Indian communities where both human and animal fecal loading are high. Ten candidate Bacteroidales qPCR assays were tested against fecal samples (human, sewage, cow, buffalo, goat, sheep, dog and chicken) from a test set of 30 individual human, 5 sewage, and 60 pooled animal samples collected in coastal Odisha, India. The two universal/general Bacteroidales assays tested (BacUni, GenBac3) performed equally well, achieving 100% sensitivity on the test set. Across the five human-associated assays tested (HF183 Taqman, BacHum, HumM2, BacH, HF183 SYBR), we found low sensitivity (17 to 49%) except for HF183 SYBR (89%), and moderate to high cross-reactivity with dog (20 to 80%) and chicken fecal samples (60 to 100%). BacHum had the highest accuracy (67%), amplified all sewage samples within the range of quantification (ROQ), and did not cross-react with any fecal samples from cows, the most populous livestock animal in India. Of the ruminant- and cattle-associated assays tested (BacCow, CowM2), BacCow was more sensitive in detecting the full range of common Indian livestock animal fecal sources, while CowM2 only detected cow sources with 50% sensitivity. Neither assay cross-reacted with human sources. BacCan, the dog-associated assay tested, showed no cross-reactivity with human sources, and high sensitivity (90%) for dog fecal samples. Overall, our results indicate BacUni, BacHum, HumM2, BacCan and BacCow would be the most suitable MST assays to distinguish and quantify relative amounts of human-associated and livestock/domestic animal-associated contributions to fecal contamination in Odisha, India. PMID:25285421

  10. TH-E-BRF-11: Dynamic Treatment of Clinical Margins Beyond the PET-Avid Target in Emission Guided Radiation Therapy: A Retrospective Patient Study

    SciTech Connect

    Nanduri, A; Mazin, S; Fan, Q; Yang, J; Graves, E; Loo, B; Yamamoto, T

    2014-06-15

    Purpose: Emission guided radiation therapy (EGRT) is a new modality that uses PET emissions for direct real-time tumor tracking. Radiation beamlets are delivered along PET lines of response (LOR's) by a fast rotating PET-Linac closed ring gantry. In this work, we develop a scheme to treat clinical margins defined proximal to the moving PET-avid tumor, while maintaining EGRT's inherent real-time tracking ability. Methods: The principle of EGRT is to deliver radiation along PET emission paths to concentrate dose in the PET-avid gross tumor volume (GTV). To account for adjacent non- PET avid regions in the clinical volume (CTV) a method was developed that expands the set of radiation beamlet responses to include the effective margin extension from the GTV to the CTV. An LOR detection may now Result in multiple beamlet responses: one along the original LOR, and others that are adjacent to it in the direction of margin extension. Evaluation studies were performed on a 4D digital patient as well as a clinical breast cancer patient with moving lung tumors. Emission data were obtained using GATE and a commercial PET scanner. Dose delivery was simulated using VMC++. For the patient study, Philips Pinnacle was used for planning and Mirada RTx was used for deformable dose registration across multiple breathing phases. Results: Compared with IMRT, the EGRT margin extension method achieved a 25.3% and 9.0% relative increase in dose to 95% of the CTV for the digital and clinical patients, respectively. The corresponding CTV dose increases without margin extension were 9.7% and 1.4%. The organs at risk doses were kept similar or lower for EGRT in both cases, with tumor tracking preserved. Conclusions: With the capability of accurate treatment of the moving CTV, EGRT has the potential to enable a practical and effective implementation of 4D biologically guided radiation therapy. Authors SRM and AN are stockholders of RefleXion Medical.

  11. ImmunoPET In Cancer Models

    PubMed Central

    Reddy, Smitha; Robinson, Matthew

    2010-01-01

    Positron Emission Tomography (PET) is playing an increasingly important role in the diagnosis, staging, and monitoring response to treatment in a variety of cancers. Recent efforts have focused on ImmunoPET, which employs antibody-based radiotracers, to image tumors based on expression of tumor-associated antigens. It is postulated that the specificity afforded by antibody targeting should both improve tumor detection and provide phenotypic information related to primary and metastatic lesions that will guide therapy decisions. Advances in antibody-engineering are providing the tools to develop antibody-based molecules with pharmacokinetic properties optimized for use as immunoPET radiotracers. Coupled with technical advances in the design of PET scanners, immunoPET holds promise to improve diagnostic imaging and to guide the use of targeted therapies. An overview of the preclinical immunoPET studies in cancer models is reviewed here. PMID:20350627

  12. t-Bu2SiF-derivatized D2-receptor ligands: the first SiFA-containing small molecule radiotracers for target-specific PET-imaging.

    PubMed

    Iovkova-Berends, Ljuba; Wängler, Carmen; Zöller, Thomas; Höfner, Georg; Wanner, Klaus Theodor; Rensch, Christian; Bartenstein, Peter; Kostikov, Alexey; Schirrmacher, Ralf; Jurkschat, Klaus; Wängler, Björn

    2011-01-01

    The synthesis, radiolabeling and in vitro evaluation of new silicon-fluoride acceptor (SiFA) derivatized D(2)-receptor ligands is reported. The SiFA-technology simplifies the introduction of fluorine-18 into target specific biomolecules for Positron-Emission-Tomography (PET). However, one of the remaining challenges, especially for small molecules such as receptor-ligands, is the bulkiness of the SiFA-moiety. We therefore synthesized four Fallypride SiFA-conjugates derivatized either directly at the benzoic acid ring system (SiFA-DMFP, SiFA-FP, SiFA-DDMFP) or at the butyl-side chain (SiFA-M-FP) and tested their receptor affinities. We found D(2)-receptor affinities for all compounds in the nanomolar range (K(i(SiFA-DMFP)) = 13.6 nM, K(i(SiFA-FP)) = 33.0 nM, K(i(SiFA-DDMFP)) = 62.7 nM and K(i(SiFA-M-FP)) = 4.21 nM). The radiofluorination showed highest yields when 10 nmol of the precursors were reacted with [(18)F]fluoride/TBAHCO(3) in acetonitrile. After a reversed phased cartridge purification the desired products could be isolated as an injectable solution after only 10 min synthesis time with radiochemical yields (RCY) of more than 40% in the case of SiFA-DMFP resulting in specific activities >41 GBq/µmol (>1,100 Ci/mmol). Furthermore, the radiolabeled products were shown to be stable in the injectable solutions, as well as in human plasma, for at least 90 min. PMID:21892125

  13. Characterization of Optically Resolved 9-fluoropropyl-dihydrotetrabenazine as a Potential PET Imaging Agent Targeting Vesicular Monoamine Transporters

    PubMed Central

    Kung, Mei-Ping; Hou, Catherine; Goswami, Rajesh; E.Ponde, Datta; Kilbourn, Michael R.; Kung, Hank F.

    2007-01-01

    Labeling derivatives of dihydrotetrabenazine (DTBZ) with F-18 (T1/2 = 110 min) instead of C-11 (T1/2 = 20 min), would improve their utility and availability for imaging vesicular monoamine transporters (VMAT2) in clinical settings. The successful synthesis, reported previously, of two novel 9-fluoroalkyl(±)-DTBZ ligands prompted us to study the optically resolved active ligand 9-fluoropropyl-(+)-DTBZ (FP-(+)-DTBZ), which may have more promising characteristics. The inhibition constant (Ki) estimated for FP-(+)-DTBZ (using [3H](±)-DTBZ as the labeled ligand in rat striatal homogenates) showed a lower value as compared to the racemic FP-(±)-DTBZ (0.10 ± 0.01 vs 0.19 ± 0.04 nM). The inactive isomer, FP-(−)-DTBZ, displayed a much lower binding affinity with a Ki value >3000 nM. Biodistribution studies in mice after an iv injection of [18F]FP-(+)-DTBZ exhibited a ratio of striatum (ST, target) to cerebellum (CB, background) of 4.51 at 30 minutes post-injection, which is a higher value than previously obtained with the racemic ligand [18F]FP-(±)-DTBZ (ST/CB = 2.95). Brain extraction at 30 minutes after the tracer injection in mice showed that >95% of the radioactivity corresponded to the parent, non-metabolized, compound remaining in the striatum, suggesting that the tracer has an excellent in vivo stability. Furthermore, localization of the tracer in the brain examined with ex vivo autoradiography displayed a typical distribution pattern consistent with VMAT2 sites. The highest labeling was observed in monoaminergic neuron regions (caudate putamen, olfactory tubercle, nucleus accumbens, substania nigra, dorsal raphe and locus coerules). We also tested the selective labeling of this tracer at the dopamine neurons in unilateral-lesioned mice (treated with 6-hydroxydopamine). When [18F]FP-(+)-DTBZ and [125I]IPT ((N-(3'-iodopropen-2'-yl)-2-beta-carbomethoxy-3-beta-(4-chlorophenyl)tropane, a selective marker for dopamine transporters in dopaminergic neurons) were

  14. ASSESSING POSSIBLE ECOLOGICAL RISKS OF GENETICALLY MODIFIED CROPS: GENE EXPRESSION ASSAYS AND GENETIC MONITORING OF NON-TARGET ORGANISMS

    EPA Science Inventory

    Widespread planting of genetically modified crops with the Bt transgene pesticide has led to concern over non-target effects of Bt compounds in agroecosystems. While some research suggests that non-target organisms exposed to Bt toxin exhibit reduced fecundity and increased morta...

  15. A sandwich-hybridization assay for simultaneous determination of HIV and tuberculosis DNA targets based on signal amplification by quantum dots-PowerVision™ polymer coding nanotracers.

    PubMed

    Yan, Zhongdan; Gan, Ning; Zhang, Huairong; Wang, De; Qiao, Li; Cao, Yuting; Li, Tianhua; Hu, Futao

    2015-09-15

    A novel sandwich-hybridization assay for simultaneous electrochemical detection of multiple DNA targets related to human immune deficiency virus (HIV) and tuberculosis (TB) was developed based on the different quantum dots-PowerVision(TM) polymer nanotracers. The polymer nanotracers were respectively fabricated by immobilizing SH-labeled oligonucleotides (s-HIV or s-TB), which can partially hybrid with virus DNA (HIV or TB), on gold nanoparticles (Au NPs) and then modified with PowerVision(TM) (PV) polymer-encapsulated quantum dots (CdS or PbS) as signal tags. PV is a dendrimer enzyme linked polymer, which can immobilize abundant QDs to amplify the stripping voltammetry signals from the metal ions (Pb or Cd). The capture probes were prepared through the immobilization of SH-labeled oligonucleotides, which can complementary with HIV and TB DNA, on the magnetic Fe3O4@Au (GMPs) beads. After sandwich-hybridization, the polymer nanotracers together with HIV and TB DNA targets were simultaneously introduced onto the surface of GMPs. Then the two encoding metal ions (Cd(2+) and Pb(2+)) were used to differentiate two viruses DNA due to the different subsequent anodic stripping voltammetric peaks at -0.84 V (Cd) and -0.61 V (Pb). Because of the excellent signal amplification of the polymer nanotracers and the great specificity of DNA targets, this assay could detect targets DNA as low as 0.2 femtomolar and exhibited excellent selectivity with the dynamitic range from 0.5 fM to 500 pM. Those results demonstrated that this electrochemical coding assay has great potential in applications for screening more viruses DNA while changing the probes. PMID:25911447

  16. Target Product Profile for a Diagnostic Assay to Differentiate between Bacterial and Non-Bacterial Infections and Reduce Antimicrobial Overuse in Resource-Limited Settings: An Expert Consensus.

    PubMed

    Dittrich, Sabine; Tadesse, Birkneh Tilahun; Moussy, Francis; Chua, Arlene; Zorzet, Anna; Tängdén, Thomas; Dolinger, David L; Page, Anne-Laure; Crump, John A; D'Acremont, Valerie; Bassat, Quique; Lubell, Yoel; Newton, Paul N; Heinrich, Norbert; Rodwell, Timothy J; González, Iveth J

    2016-01-01

    Acute fever is one of the most common presenting symptoms globally. In order to reduce the empiric use of antimicrobial drugs and improve outcomes, it is essential to improve diagnostic capabilities. In the absence of microbiology facilities in low-income settings, an assay to distinguish bacterial from non-bacterial causes would be a critical first step. To ensure that patient and market needs are met, the requirements of such a test should be specified in a target product profile (TPP). To identify minimal/optimal characteristics for a bacterial vs. non-bacterial fever test, experts from academia and international organizations with expertise in infectious diseases, diagnostic test development, laboratory medicine, global health, and health economics were convened. Proposed TPPs were reviewed by this working group, and consensus characteristics were defined. The working group defined non-severely ill, non-malaria infected children as the target population for the desired assay. To provide access to the most patients, the test should be deployable to community health centers and informal health settings, and staff should require <2 days of training to perform the assay. Further, given that the aim is to reduce inappropriate antimicrobial use as well as to deliver appropriate treatment for patients with bacterial infections, the group agreed on minimal diagnostic performance requirements of >90% and >80% for sensitivity and specificity, respectively. Other key characteristics, to account for the challenging environment at which the test is targeted, included: i) time-to-result <10 min (but maximally <2 hrs); ii) storage conditions at 0-40°C, ≤90% non-condensing humidity with a minimal shelf life of 12 months; iii) operational conditions of 5-40°C, ≤90% non-condensing humidity; and iv) minimal sample collection needs (50-100μL, capillary blood). This expert approach to define assay requirements for a bacterial vs. non-bacterial assay should guide product

  17. Target Product Profile for a Diagnostic Assay to Differentiate between Bacterial and Non-Bacterial Infections and Reduce Antimicrobial Overuse in Resource-Limited Settings: An Expert Consensus

    PubMed Central

    Dittrich, Sabine; Tadesse, Birkneh Tilahun; Moussy, Francis; Chua, Arlene; Zorzet, Anna; Tängdén, Thomas; Dolinger, David L.; Page, Anne-Laure; Crump, John A.; D’Acremont, Valerie; Bassat, Quique; Lubell, Yoel; Newton, Paul N.; Heinrich, Norbert; Rodwell, Timothy J.; González, Iveth J.

    2016-01-01

    Acute fever is one of the most common presenting symptoms globally. In order to reduce the empiric use of antimicrobial drugs and improve outcomes, it is essential to improve diagnostic capabilities. In the absence of microbiology facilities in low-income settings, an assay to distinguish bacterial from non-bacterial causes would be a critical first step. To ensure that patient and market needs are met, the requirements of such a test should be specified in a target product profile (TPP). To identify minimal/optimal characteristics for a bacterial vs. non-bacterial fever test, experts from academia and international organizations with expertise in infectious diseases, diagnostic test development, laboratory medicine, global health, and health economics were convened. Proposed TPPs were reviewed by this working group, and consensus characteristics were defined. The working group defined non-severely ill, non-malaria infected children as the target population for the desired assay. To provide access to the most patients, the test should be deployable to community health centers and informal health settings, and staff should require <2 days of training to perform the assay. Further, given that the aim is to reduce inappropriate antimicrobial use as well as to deliver appropriate treatment for patients with bacterial infections, the group agreed on minimal diagnostic performance requirements of >90% and >80% for sensitivity and specificity, respectively. Other key characteristics, to account for the challenging environment at which the test is targeted, included: i) time-to-result <10 min (but maximally <2 hrs); ii) storage conditions at 0–40°C, ≤90% non-condensing humidity with a minimal shelf life of 12 months; iii) operational conditions of 5–40°C, ≤90% non-condensing humidity; and iv) minimal sample collection needs (50–100μL, capillary blood). This expert approach to define assay requirements for a bacterial vs. non-bacterial assay should guide

  18. Inhibitory effect of target binding on hairpin aptamer sticky-end pairing-induced gold nanoparticle assembly for light-up colorimetric protein assay.

    PubMed

    Wu, Zai-Sheng; Lu, Haixia; Liu, Xueping; Hu, Rong; Zhou, Hui; Shen, Guoli; Yu, Ru-Qin

    2010-05-01

    Gold nanoparticles (GNPs) possessing strong distance-dependent optical properties and high extinction coefficients have emerged as important colorimetric materials. Almost all colorimetric studies are based on two working mechanisms: sandwich cross-linking and non-cross-linking systems. In the present study, a new working mechanism, hairpin sticky-end pairing-induced GNP assembly, is introduced based on the discovery of unique aggregation behavior of aptamer-functionalized GNPs. The salt-induced aggregation of oligonucleotide probe-modified GNPs can readily occur due to the sticky-end pairing effect while addition of target molecules favors the formation of the hairpin structure of probe sequences and substantially inhibits the nanoparticle assembly. Along this line, we developed a proof-of-concept colorimetric homogeneous assay using immunoglobulin E (IgE) as an analyte model via transforming a commonly designed "light-down" colorimetric biosensor into a "light-up" one. From the point of view of both conformational transition of aptamer and steric bulk, oligonucleotide-GNPs display an additional stability upon binding to target molecules. The assay showed an extremely high sensitivity from both naked eye observations and absorbance measurements. Compared with almost all existing IgE sensing strategies, the proposed colorimetric system possesses a substantially improved analytical performance. Investigating the assembly behavior of hairpin aptamer-modified GNPs could offer new insight into the dependence of the GNP properties on the structure switching and open a new way to design signaling probes and develop colorimetric assay schemes. PMID:20394414

  19. An Automated High-Throughput Cell-Based Multiplexed Flow Cytometry Assay to Identify Novel Compounds to Target Candida albicans Virulence-Related Proteins

    PubMed Central

    Bernardo, Stella M.; Allen, Christopher P.; Waller, Anna; Young, Susan M.; Oprea, Tudor; Sklar, Larry A.; Lee, Samuel A.

    2014-01-01

    Although three major classes of systemic antifungal agents are clinically available, each is characterized by important limitations. Thus, there has been considerable ongoing effort to develop novel and repurposed agents for the therapy of invasive fungal infections. In an effort to address these needs, we developed a novel high-throughput, multiplexed screening method that utilizes small molecules to probe candidate drug targets in the opportunistic fungal pathogen Candida albicans. This method is amenable to high-throughput automated screening and is based upon detection of changes in GFP levels of individually tagged target proteins. We first selected four GFP-tagged membrane-bound proteins associated with virulence or antifungal drug resistance in C. albicans. We demonstrated proof-of-principle that modulation of fluorescence intensity can be used to assay the expression of specific GFP-tagged target proteins to inhibitors (and inducers), and this change is measurable within the HyperCyt automated flow cytometry sampling system. Next, we generated a multiplex of differentially color-coded C. albicans strains bearing C-terminal GFP-tags of each gene encoding candidate drug targets incubated in the presence of small molecules from the Prestwick Chemical Library in 384-well microtiter plate format. Following incubation, cells were sampled through the HyperCyt system and modulation of protein levels, as indicated by changes in GFP-levels of each strain, was used to identify compounds of interest. The hit rate for both inducers and inhibitors identified in the primary screen did not exceed 1% of the total number of compounds in the small-molecule library that was probed, as would be expected from a robust target-specific, high-throughput screening campaign. Secondary assays for virulence characteristics based on null mutant strains were then used to further validate specificity. In all, this study presents a method for the identification and verification of new

  20. Detection and quantification of schistosome DNA in freshwater snails using either fluorescent probes in real-time PCR or oligochromatographic dipstick assays targeting the ribosomal intergenic spacer.

    PubMed

    Kane, Richard A; Stothard, J Russell; Rollinson, David; Leclipteux, Thierry; Evraerts, Jonathan; Standley, Claire J; Allan, Fiona; Betson, Martha; Kaba, Rehana; Mertens, Pascal; Laurent, Thierry

    2013-11-01

    Several DNA probes were designed for use in real-time polymerase chain reaction (PCR) assays to target sequence variation within the ribosomal intergenic spacer (IGS) of schistosomes. A sub-section of the IGS (∼300bp) was amplified, with cross-specific primers, after which group-specific fluorescent, locked nucleic acid probes were assessed for their ability to differentiate and quantify DNA from Schistosoma haematobium and Schistosoma mansoni group parasites. A number of fluorescent probe candidates were screened and validated against genomic DNA from adult schistosome worms and laboratory infected freshwater snails. Two fluorescent, locked nucleic acid probes ShaemLNA5 and SmanLNA2, of 20-26bp in length, were identified and found to be effective in providing evidence of infection in field-collected snails. To adapt these real-time PCR assays for more resource-poor laboratory settings, a PCR-restriction fragment length polymorphism (RFLP) assay was developed and primer/probe combinations were modified for use in oligochromatography, a DNA 'dipstick' technology. An appropriate dipstick was developed, inclusive of internal amplification and amplicon migration controls that could be of particular importance for assessing schistosome transmission dynamics. These assays and tools also have future potential for use in detection of schistosome infections in humans and livestock. PMID:22100540

  1. Quantitative detection of pork in commercial meat products by TaqMan® real-time PCR assay targeting the mitochondrial D-loop region.

    PubMed

    Kim, Miju; Yoo, Insuk; Lee, Shin-Young; Hong, Yeun; Kim, Hae-Yeong

    2016-11-01

    The TaqMan® real-time PCR assay using the mitochondrial D-loop region was developed for the quantitative detection of pork in processed meat products. The newly designed primers and probe specifically amplified pork without any cross-reactivity with non-target animal species. The limit of detection of the real-time PCR assay was 0.1pg of heat-treated pork meat and 0.1% (w/w) pork meat in beef and chicken meat mixtures. The quantitative real-time PCR assay was applied to analyze the pork meat content in 22 commercial processed meat products including jerkies, press hams, sausages, hamburger patties and steaks, grilled short rib patties, and nuggets. The developed real-time PCR method was able to detect pork meat in various types of processed meat products that declared the use of pork meat on their label. All processed meat products that declared no use of pork meat showed a negative result in the assay. The method developed in this study showed sensitivity and specificity in the quantification of pork meat in commercial processed meat products. PMID:27211626

  2. Loop-mediated isothermal amplification assay for detection of Histomonas meleagridis infection in chickens targeting the 18S rRNA sequences.

    PubMed

    Xu, Jinjun; Qu, Chanbao; Tao, Jianping

    2014-01-01

    Histomonas meleagridis is the causative agent of histomonosis, a disease of gallinaceous fowl characterized by necrotic typhlitis, hepatitis, and high mortality. To develop a rapid and sensitive method for specific detection of H. meleagridis, an assay based on loop-mediated isothermal amplification (LAMP) targeting the 18S rRNA gene was established. The detection limit of the LAMP assay was 10 copies for standard plasmids containing an 18S rRNA gene fragment, which was superior to that of a classical PCR method. Specificity tests revealed that there was no cross-reaction with other protozoa such as Trichomonas gallinae, Blastocytis sp, Tetratrichomonas gallinarum, Plasmodium gallinaceum, Toxoplasma gondii, Eimeria tenella, Leucocytozoon caulleryi and Leucocytozoon sabrazesi. The assay was evaluated for its diagnostic utility using liver and caeca samples collected from suspected field cases, the detection rate was 100 and 97.92%, respectively. These results indicate that the LAMP assay may be a useful tool for rapid detection and identification of H. meleagridis in poultry. PMID:24320623

  3. Chemo-Predictive Assay for Targeting Cancer Stem-Like Cells in Patients Affected by Brain Tumors

    PubMed Central

    Nande, Rounak; Neto, Walter; Lawrence, Logan; McCallister, Danielle R.; Denvir, James; Kimmey, Gerrit A.; Mogul, Mark; Oakley, Gerard; Denning, Krista L.; Dougherty, Thomas; Valluri, Jagan V.; Claudio, Pier Paolo

    2014-01-01

    Administration of ineffective anticancer therapy is associated with unnecessary toxicity and development of resistant clones. Cancer stem-like cells (CSLCs) resist chemotherapy, thereby causing relapse of the disease. Thus, development of a test that identifies the most effective chemotherapy management offers great promise for individualized anticancer treatments. We have developed an ex vivo chemotherapy sensitivity assay (ChemoID), which measures the sensitivity of CSLCs as well as the bulk of tumor cells to a variety of chemotherapy agents. Two patients, a 21-year old male (patient 1) and a 5-month female (patient 2), affected by anaplastic WHO grade-III ependymoma were screened using the ChemoID assay. Patient 1 was found sensitive to the combination of irinotecan and bevacizumab, which resulted in a prolonged disease progression free period of 18 months. Following recurrence, the combination of various chemotherapy drugs was tested again with the ChemoID assay. We found that benzyl isothiocyanate (BITC) greatly increased the chemosensitivity of the ependymoma cells to the combination of irinotecan and bevacizumab. After patient 1 was treated for two months with irinotecan, bevacizumab and supplements of cruciferous vegetable extracts containing BITC, we observed over 50% tumoral regression in comparison with pre-ChemoID scan as evidenced by MRI. Patient 2 was found resistant to all treatments tested and following 6 cycles of vincristine, carboplatin, cyclophosphamide, etoposide, and cisplatin in various combinations, the tumor of this patient rapidly progressed and proton beam therapy was recommended. As expected animal studies conducted with patient derived xenografts treated with ChemoID screened drugs recapitulated the clinical observation. This assay demonstrates that patients with the same histological stage and grade of cancer may vary considerably in their clinical response, suggesting that ChemoID testing which measures the sensitivity of CSLCs as

  4. Chemo-predictive assay for targeting cancer stem-like cells in patients affected by brain tumors.

    PubMed

    Mathis, Sarah E; Alberico, Anthony; Nande, Rounak; Neto, Walter; Lawrence, Logan; McCallister, Danielle R; Denvir, James; Kimmey, Gerrit A; Mogul, Mark; Oakley, Gerard; Denning, Krista L; Dougherty, Thomas; Valluri, Jagan V; Claudio, Pier Paolo

    2014-01-01

    Administration of ineffective anticancer therapy is associated with unnecessary toxicity and development of resistant clones. Cancer stem-like cells (CSLCs) resist chemotherapy, thereby causing relapse of the disease. Thus, development of a test that identifies the most effective chemotherapy management offers great promise for individualized anticancer treatments. We have developed an ex vivo chemotherapy sensitivity assay (ChemoID), which measures the sensitivity of CSLCs as well as the bulk of tumor cells to a variety of chemotherapy agents. Two patients, a 21-year old male (patient 1) and a 5-month female (patient 2), affected by anaplastic WHO grade-III ependymoma were screened using the ChemoID assay. Patient 1 was found sensitive to the combination of irinotecan and bevacizumab, which resulted in a prolonged disease progression free period of 18 months. Following recurrence, the combination of various chemotherapy drugs was tested again with the ChemoID assay. We found that benzyl isothiocyanate (BITC) greatly increased the chemosensitivity of the ependymoma cells to the combination of irinotecan and bevacizumab. After patient 1 was treated for two months with irinotecan, bevacizumab and supplements of cruciferous vegetable extracts containing BITC, we observed over 50% tumoral regression in comparison with pre-ChemoID scan as evidenced by MRI. Patient 2 was found resistant to all treatments tested and following 6 cycles of vincristine, carboplatin, cyclophosphamide, etoposide, and cisplatin in various combinations, the tumor of this patient rapidly progressed and proton beam therapy was recommended. As expected animal studies conducted with patient derived xenografts treated with ChemoID screened drugs recapitulated the clinical observation. This assay demonstrates that patients with the same histological stage and grade of cancer may vary considerably in their clinical response, suggesting that ChemoID testing which measures the sensitivity of CSLCs as

  5. Development of a Highly Automated and Multiplexed Targeted Proteome Pipeline and Assay for 112 Rat Brain Synaptic Proteins

    PubMed Central

    Colangelo, Christopher M.; Ivosev, Gordana; Chung, Lisa; Abbott, Thomas; Shifman, Mark; Sakaue, Fumika; Cox, David; Kitchen, Rob R.; Burton, Lyle; Tate, Stephen A; Gulcicek, Erol; Bonner, Ron; Rinehart, Jesse; Nairn, Angus C.; Williams, Kenneth R.

    2015-01-01

    We present a comprehensive workflow for large scale (>1000 transitions/run) label-free LC-MRM proteome assays. Innovations include automated MRM transition selection, intelligent retention time scheduling (xMRM) that improves Signal/Noise by >2-fold, and automatic peak modeling. Improvements to data analysis include a novel Q/C metric, Normalized Group Area Ratio (NGAR), MLR normalization, weighted regression analysis, and data dissemination through the Yale Protein Expression Database. As a proof of principle we developed a robust 90 minute LC-MRM assay for Mouse/Rat Post-Synaptic Density (PSD) fractions which resulted in the routine quantification of 337 peptides from 112 proteins based on 15 observations per protein. Parallel analyses with stable isotope dilution peptide standards (SIS), demonstrate very high correlation in retention time (1.0) and protein fold change (0.94) between the label-free and SIS analyses. Overall, our first method achieved a technical CV of 11.4% with >97.5% of the 1697 transitions being quantified without user intervention, resulting in a highly efficient, robust, and single injection LC-MRM assay. PMID:25476245

  6. Development and Implementation of a High-Throughput Compound Screening Assay for Targeting Disrupted ER Calcium Homeostasis in Alzheimer's Disease

    PubMed Central

    Honarnejad, Kamran; Daschner, Alexander; Giese, Armin; Zall, Andrea; Schmidt, Boris; Szybinska, Aleksandra; Kuznicki, Jacek; Herms, Jochen

    2013-01-01

    Disrupted intracellular calcium homeostasis is believed to occur early in the cascade of events leading to Alzheimer's disease (AD) pathology. Particularly familial AD mutations linked to Presenilins result in exaggerated agonist-evoked calcium release from endoplasmic reticulum (ER). Here we report the development of a fully automated high-throughput calcium imaging assay utilizing a genetically-encoded FRET-based calcium indicator at single cell resolution for compound screening. The established high-throughput screening assay offers several advantages over conventional high-throughput calcium imaging technologies. We employed this assay for drug discovery in AD by screening compound libraries consisting of over 20,000 small molecules followed by structure-activity-relationship analysis. This led to the identification of Bepridil, a calcium channel antagonist drug in addition to four further lead structures capable of normalizing the potentiated FAD-PS1-induced calcium release from ER. Interestingly, it has recently been reported that Bepridil can reduce Aβ production by lowering BACE1 activity. Indeed, we also detected lowered Aβ, increased sAPPα and decreased sAPPβ fragment levels upon Bepridil treatment. The latter findings suggest that Bepridil may provide a multifactorial therapeutic modality for AD by simultaneously addressing multiple aspects of the disease. PMID:24260442

  7. PET Imaging in Huntington's Disease.

    PubMed

    Roussakis, Andreas-Antonios; Piccini, Paola

    2015-01-01

    To date, little is known about how neurodegeneration and neuroinflammation propagate in Huntington's disease (HD). Unfortunately, no treatment is available to cure or reverse the progressive decline of function caused by the disease, thus considering HD a fatal disease. Mutation gene carriers typically remain asymptomatic for many years although alterations in the basal ganglia and cortex occur early on in mutant HD gene-carriers. Positron Emission Tomography (PET) is a functional imaging technique of nuclear medicine which enables in vivo visualization of numerous biological molecules expressed in several human tissues. Brain PET is most powerful to study in vivo neuronal and glial cells function as well as cerebral blood flow in a plethora of neurodegenerative disorders including Parkinson's disease, Alzheimer's and HD. In absence of HD-specific biomarkers for monitoring disease progression, previous PET studies in HD were merely focused on the study of dopaminergic terminals, cerebral blood flow and glucose metabolism in manifest and premanifest HD-gene carriers. More recently, research interest has been exploring novel PET targets in HD including the state of phosphodiesterse expression and the role of activated microglia. Hence, a better understanding of the HD pathogenesis mechanisms may lead to the development of targeted therapies. PET imaging follow-up studies with novel selective PET radiotracers such as 11C-IMA-107 and 11C-PBR28 may provide insight on disease progression and identify prognostic biomarkers, elucidate the underlying HD pathology and assess novel pharmaceutical agents and over time. PMID:26683130

  8. Polymerase chain reaction assay for verifying the labeling of meat and commercial meat products from game birds targeting specific sequences from the mitochondrial D-loop region.

    PubMed

    Rojas, M; González, I; Pavón, M A; Pegels, N; Hernández, P E; García, T; Martín, R

    2010-05-01

    A PCR assay was developed for the identification of meats and commercial meat products from quail (Coturnix coturnix), pheasant (Phasianus colchicus), partridge (Alectoris spp.), guinea fowl (Numida meleagris), pigeon (Columba spp.), Eurasian woodcock (Scolopax rusticola), and song thrush (Turdus philomelos) based on oligonucleotide primers targeting specific sequences from the mitochondrial D-loop region. The primers designed generated specific fragments of 96, 100, 104, 106, 147, 127, and 154 bp in length for quail, pheasant, partridge, guinea fowl, pigeon, Eurasian woodcock, and song thrush tissues, respectively. The specificity of each primer pair was tested against DNA from various game and domestic species. In this work, satisfactory amplification was accomplished in the analysis of experimentally pasteurized (72 degrees C for 30 min) and sterilized (121 degrees C for 20 min) meats, as well as in commercial meat products from the target species. The technique was also applied to raw and sterilized muscular binary mixtures, with a detection limit of 0.1% (wt/wt) for each of the targeted species. The proposed PCR assay represents a rapid and straightforward method for the detection of possible mislabeling in game bird meat products. PMID:20371856

  9. Current Status of Hybrid PET/MRI in Oncologic Imaging

    PubMed Central

    Rosenkrantz, Andrew B.; Friedman, Kent; Chandarana, Hersh; Melsaether, Amy; Moy, Linda; Ding, Yu-Shin; Jhaveri, Komal; Beltran, Luis; Jain, Rajan

    2016-01-01

    OBJECTIVE This review article explores recent advancements in PET/MRI for clinical oncologic imaging. CONCLUSION Radiologists should understand the technical considerations that have made PET/MRI feasible within clinical workflows, the role of PET tracers for imaging various molecular targets in oncology, and advantages of hybrid PET/MRI compared with PET/CT. To facilitate this understanding, we discuss clinical examples (including gliomas, breast cancer, bone metastases, prostate cancer, bladder cancer, gynecologic malignancy, and lymphoma) as well as future directions, challenges, and areas for continued technical optimization for PET/MRI. PMID:26491894

  10. Development of a Targeted Multi-Disorder High-Throughput Sequencing Assay for the Effective Identification of Disease-Causing Variants

    PubMed Central

    Delio, Maria; Patel, Kunjan; Maslov, Alex; Marion, Robert W.; McDonald, Thomas V.; Cadoff, Evan M.; Golden, Aaron; Greally, John M.; Vijg, Jan; Morrow, Bernice; Montagna, Cristina

    2015-01-01

    Background While next generation sequencing (NGS) is a useful tool for the identification of genetic variants to aid diagnosis and support therapy decision, high sequencing costs have limited its application within routine clinical care, especially in economically depressed areas. To investigate the utility of a multi-disease NGS based genetic test, we designed a custom sequencing assay targeting over thirty disease-associated areas including cardiac disorders, intellectual disabilities, hearing loss, collagenopathies, muscular dystrophy, Ashkenazi Jewish genetic disorders, and complex Mendelian disorders. We focused on these specific areas based on the interest of our collaborative clinical team, suggesting these diseases being the ones in need for the development of a sequencing-screening assay. Results We targeted all coding, untranslated regions (UTR) and flanking intronic regions of 650 known disease-associated genes using the Roche-NimbleGen EZ SeqCapV3 capture system and sequenced on the Illumina HiSeq 2500 Rapid Run platform. Eight controls with known variants and one HapMap sample were first sequenced to assess the performance of the panel. Subsequently, as a proof of principle and to explore the possible utility of our test, we analyzed test disease subjects (n = 16). Eight had known Mendelian disorders and eight had complex pediatric diseases. In addition to assess whether copy number variation may be of utility as a companion assay relative to these specific disease areas, we used the Affymetrix Genome-Wide SNP Array 6.0 to analyze the same samples. Conclusion We identified potentially disease-associated variants: 22 missense, 4 nonsense, 1 frameshift, and 1 splice variants (16 previously identified, 12 novel among dbSNP and 15 novel among NHLBI Exome Variant Server). We found multi-disease targeted high-throughput sequencing to be a cost efficient approach in detecting disease-associated variants to aid diagnosis. PMID:26214305

  11. Pet Health

    MedlinePlus

    ... Know the signs of medical problems. Take your pet to the veterinarian if you notice: Loss of appetite Drinking a lot of water Gaining or losing a lot of weight quickly Strange behavior Being sluggish and tired Trouble getting up or down Strange lumps

  12. Colorimetric microtiter plate receptor-binding assay for the detection of freshwater and marine neurotoxins targeting the nicotinic acetylcholine receptors

    USGS Publications Warehouse

    Rubio, Fernando; Kamp, Lisa; Carpino, Justin; Faltin, Erin; Loftin, Keith A.; Molgó, Jordi; Aráoz, Rómulo

    2014-01-01

    Anatoxin-a and homoanatoxin-a, produced by cyanobacteria, are agonists of nicotinic acetylcholine receptors (nAChRs). Pinnatoxins, spirolides, and gymnodimines, produced by dinoflagellates, are antagonists of nAChRs. In this study we describe the development and validation of a competitive colorimetric, high throughput functional assay based on the mechanism of action of freshwater and marine toxins against nAChRs. Torpedo electrocyte membranes (rich in muscle-type nAChR) were immobilized and stabilized on the surface of 96-well microtiter plates. Biotinylated α-bungarotoxin (the tracer) and streptavidin-horseradish peroxidase (the detector) enabled the detection and quantitation of anatoxin-a in surface waters and cyclic imine toxins in shellfish extracts that were obtained from different locations across the US. The method compares favorably to LC/MS/MS and provides accurate results for anatoxin-a and cyclic imine toxins monitoring. Study of common constituents at the concentrations normally found in drinking and environmental waters, as well as the tolerance to pH, salt, solvents, organic and inorganic compounds did not significantly affect toxin detection. The assay allowed the simultaneous analysis of up to 25 samples within 3.5 h and it is well suited for on-site or laboratory monitoring of low levels of toxins in drinking, surface, and ground water as well as in shellfish extracts.

  13. Area-under-the-curve monitoring of cyclosporine therapy: Performance of different assay methods and their target concentrations

    SciTech Connect

    Grevel, J.; Napoli, K.L.; Gibbons, S.; Kahan, B.D. )

    1990-01-01

    The measurement of areas under the concentration-time curve (AUC) was recently introduced as an alternative to trough level monitoring of cyclosporine therapy. The AUC is divided by the oral dosing interval to calculate an average concentration. All measurements are performed at clinical steady state. The initial evaluation of AUC monitoring showed advantages over trough level monitoring with concentrations of cyclosporine measured in serum by the polyclonal radioimmunoassay of Sandoz. This assay technique is no longer available and the following assays were performed in parallel during up to 173 AUC determinations in 51 consecutive renal transplant patients: polyclonal fluorescence polarization immunoassay of Abbott in serum, specific and nonspecific monoclonal radioimmunoassays using {sup 3}H and {sup 125}I tracers in serum and whole blood, and high performance liquid chromatography in whole blood. Both trough levels and average concentrations at steady state measured by those different techniques were significantly correlated with the oral dose. The best correlation (r2 = 0.54) was shown by average concentrations measured in whole blood by the specific monoclonal radioimmunoassay of Sandoz ({sup 3}H tracer). This monitoring technique was also associated with the smallest absolute error between repeated observations in the same patient while the oral dose rate remained the same or was changed. Both allegedly specific monoclonal radioimmunoassays (with {sup 3}H and {sup 125}I tracer) measured significantly higher concentrations than the liquid chromatography.

  14. Colorimetric microtiter plate receptor-binding assay for the detection of freshwater and marine neurotoxins targeting the nicotinic acetylcholine receptors.

    PubMed

    Rubio, Fernando; Kamp, Lisa; Carpino, Justin; Faltin, Erin; Loftin, Keith; Molgó, Jordi; Aráoz, Rómulo

    2014-12-01

    Anatoxin-a and homoanatoxin-a, produced by cyanobacteria, are agonists of nicotinic acetylcholine receptors (nAChRs). Pinnatoxins, spirolides, and gymnodimines, produced by dinoflagellates, are antagonists of nAChRs. In this study we describe the development and validation of a competitive colorimetric, high throughput functional assay based on the mechanism of action of freshwater and marine toxins against nAChRs. Torpedo electrocyte membranes (rich in muscle-type nAChR) were immobilized and stabilized on the surface of 96-well microtiter plates. Biotinylated α-bungarotoxin (the tracer) and streptavidin-horseradish peroxidase (the detector) enabled the detection and quantitation of anatoxin-a in surface waters and cyclic imine toxins in shellfish extracts that were obtained from different locations across the US. The method compares favorably to LC/MS/MS and provides accurate results for anatoxin-a and cyclic imine toxins monitoring. Study of common constituents at the concentrations normally found in drinking and environmental waters, as well as the tolerance to pH, salt, solvents, organic and inorganic compounds did not significantly affect toxin detection. The assay allowed the simultaneous analysis of up to 25 samples within 3.5 h and it is well suited for on-site or laboratory monitoring of low levels of toxins in drinking, surface, and ground water as well as in shellfish extracts. PMID:25260255

  15. Gene-targeted embryonic stem cells: real-time PCR assay for estimation of the number of neomycin selection cassettes

    PubMed Central

    2011-01-01

    In the preparation of transgenic murine ES cells it is important to verify the construct has a single insertion, because an ectopic neomycin phosphortransferase positive selection cassette (NEO) may cause a position effect. During a recent work, where a knockin SCA28 mouse was prepared, we developed two assays based on Real-Time PCR using both SYBR Green and specific minor groove binder (MGB) probes to evaluate the copies of NEO using the comparative delta-delta Ct method versus the Rpp30 reference gene. We compared the results from Southern blot, routinely used to quantify NEO copies, with the two Real-Time PCR assays. Twenty-two clones containing the single NEO copy showed values of 0.98 ± 0.24 (mean ± 2 S.D.), and were clearly distinguishable from clones with two or more NEO copies. This method was found to be useful, easy, sensitive and fast and could substitute for the widely used, but laborious Southern blot method. PMID:22035318

  16. Multi-laboratory evaluations of the performance of Catellicoccus marimammalium PCR assays developed to target gull fecal sources

    EPA Science Inventory

    Here we report results from a multi-laboratory (n=11) evaluation of four different PCR methods targeting the 16S rRNA gene of Catellicoccus marimammalium used to detect fecal contamination from birds in coastal environments. The methods included conventional end-point PCR, a SYBR...

  17. Development of a Rapid, Sensitive, and Field-Deployable Razor Ex BioDetection System and Quantitative PCR Assay for Detection of Phymatotrichopsis omnivora Using Multiple Gene Targets

    PubMed Central

    Arif, M.; Marek, S. M.; Melcher, U.

    2013-01-01

    A validated, multigene-based method using real-time quantitative PCR (qPCR) and the Razor Ex BioDetection system was developed for detection of Phymatotrichopsis omnivora. This soilborne fungus causes Phymatotrichopsis root rot of cotton, alfalfa, and other dicot crops in the southwestern United States and northern Mexico, leading to significant crop losses and limiting the range of crops that can be grown in soils where the fungus is established. It is on multiple lists of regulated organisms. Because P. omnivora is difficult to isolate, accurate and sensitive culture-independent diagnostic tools are needed to confirm infections by this fungus. Specific PCR primers and probes were designed based on P. omnivora nucleotide sequences of the genes encoding rRNA internal transcribed spacers, beta-tubulin, and the second-largest subunit of RNA polymerase II (RPB2). PCR products were cloned and sequenced to confirm their identity. All primer sets allowed early detection of P. omnivora in infected but asymptomatic plants. A modified rapid DNA purification method, which facilitates a quick (∼30-min) on-site assay capability for P. omnivora detection, was developed. Combined use of three target genes increased the assay accuracy and broadened the range of detection. To our knowledge, this is the first report of a multigene-based, field-deployable, rapid, and reliable identification method for a fungal plant pathogen and should serve as a model for the development of field-deployable assays of other phytopathogens. PMID:23354717

  18. Development of a rapid, sensitive, and field-deployable razor ex BioDetection system and quantitative PCR assay for detection of Phymatotrichopsis omnivora using multiple gene targets.

    PubMed

    Arif, M; Fletcher, J; Marek, S M; Melcher, U; Ochoa-Corona, F M

    2013-04-01

    A validated, multigene-based method using real-time quantitative PCR (qPCR) and the Razor Ex BioDetection system was developed for detection of Phymatotrichopsis omnivora. This soilborne fungus causes Phymatotrichopsis root rot of cotton, alfalfa, and other dicot crops in the southwestern United States and northern Mexico, leading to significant crop losses and limiting the range of crops that can be grown in soils where the fungus is established. It is on multiple lists of regulated organisms. Because P. omnivora is difficult to isolate, accurate and sensitive culture-independent diagnostic tools are needed to confirm infections by this fungus. Specific PCR primers and probes were designed based on P. omnivora nucleotide sequences of the genes encoding rRNA internal transcribed spacers, beta-tubulin, and the second-largest subunit of RNA polymerase II (RPB2). PCR products were cloned and sequenced to confirm their identity. All primer sets allowed early detection of P. omnivora in infected but asymptomatic plants. A modified rapid DNA purification method, which facilitates a quick (∼30-min) on-site assay capability for P. omnivora detection, was developed. Combined use of three target genes increased the assay accuracy and broadened the range of detection. To our knowledge, this is the first report of a multigene-based, field-deployable, rapid, and reliable identification method for a fungal plant pathogen and should serve as a model for the development of field-deployable assays of other phytopathogens. PMID:23354717

  19. Development of a rapid, sensitive TaqMan real-time RT-PCR assay for the detection of Rose rosette virus using multiple gene targets.

    PubMed

    Babu, Binoy; Jeyaprakash, Ayyamperumal; Jones, Debra; Schubert, Timothy S; Baker, Carlye; Washburn, Brian K; Miller, Steven H; Poduch, Kristina; Knox, Gary W; Ochoa-Corona, Francisco M; Paret, Mathews L

    2016-09-01

    Rose rosette virus (RRV), belonging to the genus Emaravirus, is a highly destructive pathogen that causes rose rosette disease. The disease is a major concern for the rose industry in the U.S. due to the lack of highly sensitive methods for early detection of RRV. This is critical, as early identification of the infected plants and eradication is necessary in minimizing the risks associated with the spread of the disease. A highly reliable, specific and sensitive detection assay is thus required to test and confirm the presence of RRV in suspected plant samples. In this study a TaqMan real-time reverse transcription-polymerase chain reaction (RT-PCR) assay was developed for the detection of RRV from infected roses, utilizing multiple gene targets. Four pairs of primers and probes; two of them (RRV_2-1 and RRV_2-2) based on the consensus sequences of the glycoprotein gene (RNA2) and the other two (RRV_3-2 and RRV_3-5) based on the nucleocapsid gene (RNA3) were designed. The specificity of the primers and probes was evaluated against other representative viruses infecting roses, belonging to the genera Alfamovirus, Cucumovirus, Ilarvirus, Nepovirus, Tobamovirus, and Tospovirus and one Emaravirus (Wheat mosaic virus). Dilution assays using the in vitro transcripts (spiked with total RNA from healthy plants, and non-spiked) showed that all the primers and probes are highly sensitive in consistently detecting RRV with a detection limit of 1 fg. Testing of the infected plants over a period of time (three times in monthly intervals) indicated high reproducibility, with the primer/probe RRV_3-5 showing 100% positive detection, while RRV_2-1, RRV_2-2 and RRV_3-2 showed 90% positive detection. The developed real-time RT-PCR assay is reliable, highly sensitive, and can be easily used in diagnostic laboratories for testing and confirmation of RRV. PMID:27210549

  20. PET Imaging of Inflammation Biomarkers

    PubMed Central

    Wu, Chenxi; Li, Fang; Niu, Gang; Chen, Xiaoyuan

    2013-01-01

    Inflammation plays a significant role in many disease processes. Development in molecular imaging in recent years provides new insight into the diagnosis and treatment evaluation of various inflammatory diseases and diseases involving inflammatory process. Positron emission tomography using 18F-FDG has been successfully applied in clinical oncology and neurology and in the inflammation realm. In addition to glucose metabolism, a variety of targets for inflammation imaging are being discovered and utilized, some of which are considered superior to FDG for imaging inflammation. This review summarizes the potential inflammation imaging targets and corresponding PET tracers, and the applications of PET in major inflammatory diseases and tumor associated inflammation. Also, the current attempt in differentiating inflammation from tumor using PET is also discussed. PMID:23843893

  1. Potential of the microbial assay for risk assessment (MARA) for assessing ecotoxicological effects of herbicides to non-target organisms.

    PubMed

    Fai, Patricia Bi Asanga; Mbida, Mpoame; Demefack, Jean Marc; Yamssi, Cedric

    2015-11-01

    Many microbiotests that have been proposed for use in the risk assessment of environmental pollutants have the drawback of lacking relevant published data on various aspects of their test application possibilities and therefore do not receive the regulatory recognition which they may deserve. The MARA bioassay lacks published data for many relevant environmental pollutants, particularly pesticides and this may limit its use in regulatory framework. The present study has assessed the sensitivity of the MARA bioassay relative to other established bioassays (Daphnia magna and Oreochromis niloticus) to two widely used herbicide formulations: Roundup (having glyphosate as active ingredient) and Herbextra (with the active ingredient being 2,4-dichlorophenoxyacetic acid-2,4-D). Roundup was found to be more toxic than Herbextra in all three bioassays. The D. magna EC50 s obtained for Roundup and Herbextra were respectively 5.55 and 356.61 mg/l while the LC50 s for O. niloticus were 11.30 and 222,28 mg/l respectively. In the case of the MARA bioassay microbial toxic concentrations (MTCs) for individual species ranged from 6.85 to 468 mg/l with an overall mean MTC of 101.82 mg/l for glyphosate and from 74.67 to 13,333 mg/l for 2,4-D giving an overall mean MTC of 2855.88 mg/l. Although the overall MTCs from the MARA bioassay were much higher than the LC50 s and EC50 s from the fish and daphnia bioassays respectively, the most sensitive MARA organism for each of the herbicides had MTCs that were comparable to or lower than the corresponding endpoints from the other bioassays implying that the MARA assay is a potentially useful bioassay for risk assessment of pesticides. PMID:26362569

  2. Discovery of [¹¹C]MK-8193 as a PET tracer to measure target engagement of phosphodiesterase 10A (PDE10A) inhibitors.

    PubMed

    Cox, Christopher D; Hostetler, Eric D; Flores, Broc A; Evelhoch, Jeffrey L; Fan, Hong; Gantert, Liza; Holahan, Marie; Eng, Waisi; Joshi, Aniket; McGaughey, Georgia; Meng, Xiangjun; Purcell, Mona; Raheem, Izzat T; Riffel, Kerry; Yan, Youwei; Renger, John J; Smith, Sean M; Coleman, Paul J

    2015-11-01

    Phosphodiesterase 10A (PDE10A) inhibition has recently been identified as a potential mechanism to treat multiple symptoms that manifest in schizophrenia. In order to facilitate preclinical development and support key proof-of-concept clinical trials of novel PDE10A inhibitors, it is critical to discover positron emission tomography (PET) tracers that enable plasma concentration/PDE10A occupancy relationships to be established across species with structurally diverse PDE10A inhibitors. In this Letter, we describe how a high-throughput screening hit was optimized to provide [(11)C]MK-8193 (8j), a PET tracer that supports the determination of plasma concentration/PDE10A occupancy relationships for structurally diverse series of PDE10A inhibitors in both rat and rhesus monkey. PMID:26077491

  3. Total ApoE and ApoE4 Isoform Assays in an Alzheimer's Disease Case-control Study by Targeted Mass Spectrometry (n = 669): A Pilot Assay for Methionine-containing Proteotypic Peptides*

    PubMed Central

    Simon, Romain; Girod, Marion; Fonbonne, Catherine; Salvador, Arnaud; Clément, Yohann; Lantéri, Pierre; Amouyel, Philippe; Lambert, Jean Charles; Lemoine, Jérôme

    2012-01-01

    Allelic polymorphism of the apolipoprotein E (ApoE) gene (ApoE ε2, ApoE ε3 and ApoE ε4 alleles) gives rise to three protein isoforms (ApoE2, ApoE3 and ApoE4) that differ by 1 or 2 amino acids. Inheritance of the ApoE ε4 allele is a risk factor for developing Alzheimer's disease (AD). The potential diagnostic value of ApoE protein levels in biological fluids (i.e. cerebrospinal fluid, plasma and serum) for distinguishing between AD patients and healthy elderly subjects is subject to great controversy. Although a recent study reported subnormal total ApoE and ApoE4 levels in the plasma of AD patients, other studies have found normal or even elevated protein levels (versus controls). Because all previously reported assays were based on immunoenzymatic techniques, we decided to develop an orthogonal assay based on targeted mass spectrometry by tracking (i) a proteotypic peptide common to all ApoE isoforms and (ii) a peptide that is specific for the ε4 allele. After trypsin digestion, the ApoE4-specific peptide contains an oxidation-prone methionine residue. The endogenous methionine oxidation level was evaluated in a small cohort (n = 68) of heterozygous ε3ε4 carriers containing both healthy controls and AD patients. As expected, the proportion of oxidized residues varied from 0 to 10%, with an average of 5%. We therefore developed a standardized strategy for the unbiased, absolute quantification of ApoE4, based on performic acid oxidization of methionine. Once the sample workflow had been thoroughly validated, it was applied to the concomitant quantification of total ApoE and ApoE4 isoform in a large case-control study (n = 669). The final measurements were consistent with most previously reported ApoE concentration values and confirm the influence of the different alleles on the protein expression level. Our results illustrate (i) the reliability of selected reaction monitoring-based assays and (ii) the value of the oxidization step for unbiased monitoring of

  4. Assaying Pharmacodynamic Endpoints with Targeted Therapy: Flavopiridol and 17AAG Induced Dephosphorylation of Histone H1.5 in Acute Myeloid Leukemia

    PubMed Central

    Wang, Liwen; Harshman, Sean W.; Liu, Shujun; Ren, Chen; Xu, Hua; Sallans, Larry; Grever, Michael; Byrd, John C.; Marcucci, Guido; Freitas, Michael A.

    2011-01-01

    Histone H1 is commonly used to assay kinase activity in vitro. As many promising targeted therapies affect kinase activity of specific enzymes involved in cancer transformation, H1 phosphorylation can serve as potential pharmacodynamic marker for drug activity within the cell. In this report we utilized a phosphoproteomic workflow to characterize histone H1 phosphorylation changes associated with two targeted therapies in the Kasumi-1 Acute Myeloid Leukemia (AML) cell line. The phosphoproteomic workflow was first validated with standard casein phosphoproteins and then applied to the direct analysis of histone H1 from Kasumi-1 nuclear lysates. Ten H1 phosphorylation sites were identified on the H1 variants, H1.2, H1.3, H1.4, H1.5 and H1.x. Liquid chromatography mass spectrometry profiling of intact H1s demonstrated global dephosphorylation of H1.5 associated with therapy by the cyclin dependent kinase inhibitor, flavopiridol, and the Hsp90 inhibitor, 17AAG (17-(Allylamino)-17-demethoxygeldanamycin). In contrast, independent treatments with a nucleotide analog, proteosome inhibitor and histone deacetylase inhibitor did not exhibit decreased H1.5 phosphorylation. The data presented herein demonstrate that potential of histones to assess the cellular response of reagents that have direct and indirect effects on kinase activity that alters histone phosphorylation. As such, this approach may be a highly informative marker for response to targeted therapies influencing histone phosphorylation. PMID:21110323

  5. Patient-Specific Dosimetry Using Pretherapy [124I]m-iodobenzylguanidine ([124I]mIBG) Dynamic PET/CT Imaging Before [131I]mIBG Targeted Radionuclide Therapy for Neuroblastoma

    PubMed Central

    Huang, Shih-ying; Bolch, Wesley E.; Lee, Choonsik; Van Brocklin, Henry F.; Pampaloni, Miguel H.; Hawkins, Randall A.; Sznewajs, Aimee; DuBois, Steven G.; Matthay, Katherine K.; Seo, Youngho

    2014-01-01

    Purpose Iodine-131-m-iodobenzylguanidine ([131I]mIBG) targeted radionuclide therapy (TRT) is a standard treatment for recurrent or refractory neuroblastoma with response rates of 30–40%. The aim of this study is to demonstrate patient-specific dosimetry using quantitative [124I]mIBG PET/CT imaging with a Geant4-based Monte Carlo method for better treatment planning. Procedures A Monte Carlo dosimetry method was developed using the Geant4 toolkit with voxelized anatomical geometry and source distribution as input. The pre-segmented hybrid computational human phantoms developed by the University of Florida and the National Cancer Institute (UF/NCI) were used as a surrogate to characterize the anatomy of a given patient. S-values for I-131 were estimated by the phantoms coupled with Geant4 and compared with those estimated by OLINDA|EXM and MCNPX for the newborn model. To obtain patient-specific biodistribution of [131I]mIBG, a 10-year-old girl with relapsed neuroblastoma was imaged with [124I]mIBG PET/CT at four time points prior to the planned [131I]mIBG TRT. The organ and tumor absorbed dose of the clinical case were estimated with the Geant4 method using the modified UF/NCI 10-year-old phantom with tumors and the patient-specific residence time. Results For the newborn model, the Geant4 S-values were consistent with the MCNPX S- values. The S-value ratio of the Geant4 method to OLINDA|EXM ranged from 0.08 to 6.5 of all major organs. The [131I]mIBG residence time quantified from the pretherapy [124I]mIBG PET/CT imaging of the 10-year-old patient was mostly comparable to those previously reported. Organ absorbed dose for the salivary glands were 98.0 Gy, heart wall, 36.5 Gy, and liver, 34.3 Gy; while tumor absorbed dose ranged from 143.9 Gy to 1641.3 Gy in different sites. Conclusions Patient-specific dosimetry for [131I]mIBG targeted radionuclide therapy was accomplished using pretherapy [124I]mIBG PET/CT imaging and a Geant4-based Monte Carlo dosimetry method

  6. Improvement of the performance of targeted LC-MS assays through enrichment of histidine-containing peptides.

    PubMed

    Mesmin, Cédric; Domon, Bruno

    2014-12-01

    Mass spectrometric-based quantification using targeted methods has matured during the past decade and is now commonly used in proteomics. However, the reliability of protein quantification in complex matrixes using selected reaction monitoring is often impaired by interfering signals arising from coelution of nontargeted components. Sample preparation methods resulting in the reduction of the number of peptides present in the mixture minimizes this effect. One solution consists in the selective capture of peptides containing infrequent amino acids. The enrichment of histidine-containing peptides via immobilized metal-ion affinity chromatography loaded with Cu(2+) ions (IMAC-Cu) was applied in a quantitative workflow and found to be a simple and cost effective method for the reduction of sample complexity with high recovery and selectivity. When applied to a series of depleted human plasma digests, the method decreased nonspecific signals, resulting in a more precise and robust protein quantification. The method was also shown to be an alternative to HSA/IgG depletion during plasma protein analysis. This method, used in conjunction with recent improvements in the instrument's peak capacity, addresses a bottleneck generally encountered in quantitative proteomics studies by providing the robustness and throughput required for the analysis of large sample series without compromising the number of proteins monitored. PMID:25321649

  7. High Specificity of a Quantitative PCR Assay Targeting a Saxitoxin Gene for Monitoring Toxic Algae Associated with Paralytic Shellfish Toxins in the Yellow Sea

    PubMed Central

    Gao, Yan; Murray, Shauna A.; Chen, Jian-Hua; Kang, Zhen-Jun; Zhang, Qing-Chun; Kong, Fan-Zhou; Zhou, Ming-Jiang

    2015-01-01

    The identification of core genes involved in the biosynthesis of saxitoxin (STX) offers a great opportunity to detect toxic algae associated with paralytic shellfish toxins (PST). In the Yellow Sea (YS) in China, both toxic and nontoxic Alexandrium species are present, which makes it a difficult issue to specifically monitor PST-producing toxic algae. In this study, a quantitative PCR (qPCR) assay targeting sxtA4, a domain in the sxt gene cluster that encodes a unique enzyme involved in STX biosynthesis, was applied to analyze samples collected from the YS in spring of 2012. The abundance of two toxic species within the Alexandrium tamarense species complex, i.e., A. fundyense and A. pacificum, was also determined with TaqMan-based qPCR assays, and PSTs in net-concentrated phytoplankton samples were analyzed with high-performance liquid chromatography coupled with a fluorescence detector. It was found that the distribution of the sxtA4 gene in the YS was consistent with the toxic algae and PSTs, and the quantitation results of sxtA4 correlated well with the abundance of the two toxic species (r = 0.857). These results suggested that the two toxic species were major PST producers during the sampling season and that sxtA-based qPCR is a promising method to detect toxic algae associated with PSTs in the YS. The correlation between PST levels and sxtA-based qPCR results, however, was less significant (r = 0.552), implying that sxtA-based qPCR is not accurate enough to reflect the toxicity of PST-producing toxic algae. The combination of an sxtA-based qPCR assay and chemical means might be a promising method for monitoring toxic algal blooms. PMID:26231652

  8. High Specificity of a Quantitative PCR Assay Targeting a Saxitoxin Gene for Monitoring Toxic Algae Associated with Paralytic Shellfish Toxins in the Yellow Sea.

    PubMed

    Gao, Yan; Yu, Ren-Cheng; Murray, Shauna A; Chen, Jian-Hua; Kang, Zhen-Jun; Zhang, Qing-Chun; Kong, Fan-Zhou; Zhou, Ming-Jiang

    2015-10-01

    The identification of core genes involved in the biosynthesis of saxitoxin (STX) offers a great opportunity to detect toxic algae associated with paralytic shellfish toxins (PST). In the Yellow Sea (YS) in China, both toxic and nontoxic Alexandrium species are present, which makes it a difficult issue to specifically monitor PST-producing toxic algae. In this study, a quantitative PCR (qPCR) assay targeting sxtA4, a domain in the sxt gene cluster that encodes a unique enzyme involved in STX biosynthesis, was applied to analyze samples collected from the YS in spring of 2012. The abundance of two toxic species within the Alexandrium tamarense species complex, i.e., A. fundyense and A. pacificum, was also determined with TaqMan-based qPCR assays, and PSTs in net-concentrated phytoplankton samples were analyzed with high-performance liquid chromatography coupled with a fluorescence detector. It was found that the distribution of the sxtA4 gene in the YS was consistent with the toxic algae and PSTs, and the quantitation results of sxtA4 correlated well with the abundance of the two toxic species (r=0.857). These results suggested that the two toxic species were major PST producers during the sampling season and that sxtA-based qPCR is a promising method to detect toxic algae associated with PSTs in the YS. The correlation between PST levels and sxtA-based qPCR results, however, was less significant (r=0.552), implying that sxtA-based qPCR is not accurate enough to reflect the toxicity of PST-producing toxic algae. The combination of an sxtA-based qPCR assay and chemical means might be a promising method for monitoring toxic algal blooms. PMID:26231652

  9. A multiplexed targeted assay for high-throughput quantitative analysis of serum methylamines by ultra performance liquid chromatography coupled to high resolution mass spectrometry.

    PubMed

    Kadar, Hanane; Dubus, Justine; Dutot, Jérémie; Hedjazi, Lyamine; Srinivasa, Suman; Fitch, Kathleen V; Grinspoon, Steven K; Nicholson, Jeremy K; Dumas, Marc-Emmanuel; Gauguier, Dominique

    2016-05-01

    Methylamines are biologically-active metabolites present in serum and urine samples, which play complex roles in metabolic diseases. Methylamines can be detected by proton nuclear magnetic resonance (NMR), but specific methods remain to be developed for their routine assay in human serum in clinical settings. Here we developed and validated a novel reliable "methylamine panel" method for simultaneous quantitative analysis of trimethylamine (TMA), its major detoxification metabolite trimethylamine-N-oxide (TMAO), and precursors choline, betaine and l-carnitine in human serum using Ultra Performance Liquid Chromatography (UPLC) coupled to High Resolution Mass Spectrometry (HRMS). Metabolite separation was carried out on a HILIC stationary phase. For all metabolites, the assay was linear in the range of 0.25-12.5 μmol/L and enabled to reach limit of detection of about 0.10 μmol/L. Relative standard deviations were below 16% for the three levels of concentrations. We demonstrated the strong reliability and robustness of the method, which was applied to serum samples from healthy individuals to establish the range of concentrations of the metabolites and their correlation relationships and detect gender differences. Our data provide original information for implementing in a clinical environment a MS-based diagnostic method with potential for targeted metabolic screening of patients at risk of cardiometabolic diseases. PMID:27036856

  10. Development of a Sequence-Characterized Amplified Region Marker-Targeted Quantitative PCR Assay for Strain-Specific Detection of Oenococcus oeni during Wine Malolactic Fermentation▿

    PubMed Central

    Solieri, Lisa; Giudici, Paolo

    2010-01-01

    Control over malolactic fermentation (MLF) is a difficult goal in winemaking and needs rapid methods to monitor Oenococcus oeni malolactic starters (MLS) in a stressful environment such as wine. In this study, we describe a novel quantitative PCR (QPCR) assay enabling the detection of an O. oeni strain during MLF without culturing. O. oeni strain LB221 was used as a model to develop a strain-specific sequence-characterized amplified region (SCAR) marker derived from a discriminatory OPA20-based randomly amplified polymorphic DNA (RAPD) band. The 5′ and 3′ flanking regions and the copy number of the SCAR marker were characterized using inverse PCR and Southern blotting, respectively. Primer pairs targeting the SCAR sequence enabled strain-specific detection without cross amplification of other O. oeni strains or wine species of lactic acid bacteria (LAB), acetic acid bacteria (AAB), and yeasts. The SCAR-QPCR assay was linear over a range of cell concentrations (7 log units) and detected as few as 2.2 × 102 CFU per ml of red wine with good quantification effectiveness, as shown by the correlation of QPCR and plate counting results. Therefore, the cultivation-independent monitoring of a single O. oeni strain in wine based on a SCAR marker represents a rapid and effective strain-specific approach. This strategy can be adopted to develop easy and rapid detection techniques for monitoring the implantation of inoculated O. oeni MLS on the indigenous LAB population, reducing the risk of unsuccessful MLF. PMID:20935116

  11. Pet Problems at Home: Pet Problems in the Community.

    ERIC Educational Resources Information Center

    Soltow, Willow

    1984-01-01

    Discusses problems of pets in the community, examining the community's role related to disruptive pets and pet overpopulation. Also discusses pet problems at home, offering advice on selecting a pet, meeting a pet's needs, and disciplining pets. Includes a list of books, films/filmstrips, teaching materials, and various instructional strategies.…

  12. A triple-amplification colorimetric assay for antibiotics based on magnetic aptamer-enzyme co-immobilized platinum nanoprobes and exonuclease-assisted target recycling.

    PubMed

    Miao, Yangbao; Gan, Ning; Ren, Hong-Xia; Li, Tianhua; Cao, Yuting; Hu, Futao; Yan, Zhongdan; Chen, Yinji

    2015-11-21

    Herein, an ultrasensitive and selective colorimetric assay for antibiotics, using chloramphenicol (CAP) as the model analyte, was developed based on magnetic aptamer-HRP-platinum composite probes and exonuclease-assisted target recycling. The composite probes were prepared through immunoreactions between the double stranded DNA antibody (anti-DNA) labeled on core-shell Fe3O4@Au nanoparticles (AuMNP-anti-DNA) as the capture probe, and the double stranded aptamer (aptamer hybrid with its complementary oligonucleotides) labeled on Pt@HRP nanoparticles as the nanotracer (ds-Apt-HRP-PtNPs). When the CAP samples were incubated with the probes for 30 min at room temperature, they could be captured by the aptamer to form a nanotracer-CAP complex, which was then released into the supernatant after magnetic separation. This is because the anti-DNA on the capture probes cannot recognize the single strand aptamer-CAP complex. The exonuclease I (Exo I) added into the supernatant can further digest the aptamer-CAP from the 3'-end of the aptamer and the CAP in the aptamer-CAP complex can be released again, which can further participate in a new cycling process to react with the probes. Pt and HRP in the nanotracer could both catalyze and dual amplify the absorbance at 650 nm ascribed to the 3,3',5,5'-tetramethylbenzidine (TMB)-H2O2 system. Moreover, Exo I can assist the target recycling, which can further amplify the signal. Thus, the triple amplified signal can be quantified by ultraviolet-visible spectroscopy. The experimental results showed that the CAP detection possessed a linear range of 0.001-10 ng mL(-1) and a detection limit of 0.0003 ng mL(-1) (S/N = 3). The assay was successfully employed to detect CAP in milk, which is much more facile, time saving, and sensitive than the commercial ELISA kits. PMID:26442572

  13. Development of a PCR Assay Based on a Single-Base Pair Substitution for the Detection of Aeromonas caviae by Targeting the gyrB Gene.

    PubMed

    Payattikul, Penpan; Longyant, Siwaporn; Sithigorngul, Paisarn; Chaivisuthangkura, Parin

    2015-09-01

    Aeromonas caviae is a bacterial pathogen that causes various infectious diseases in both humans and animals. To facilitate its detection, we developed species-specific primer sets targeting polymorphisms in the gyrB gene for use in a PCR assay. The technique was able to detect 100% (29/29) of the A. caviae strains tested using either of two sets of primers (designated ACF1-ACR and ACF3-ACR), which produced 293-bp and 206-bp amplicons, respectively. Another set of primers (designated ACF2-ACR) yielded a 237-bp amplicon and exhibited 90% (26/29) positive results with respect to A. caviae. None of the primer sets exhibited cross-reactivity with 12 non-A. caviae isolates and 52 other non-Aeromonas bacteria. The detection limit using the ACF2-ACR and ACF3-ACR primer sets in pure culture was 1.6 × 10(3) CFU/mL, or 6 CFU per reaction, whereas that of the ACF1-ACR primer set was 1.6 × 10(4) CFU/mL, or 60 CFU per reaction. In the case of spiked Nile Tilapia Oreochromis niloticus, the sensitivity of all primer sets without enrichment was 1.8 × 10(4) CFU/g, or 30 CFU per reaction. Primer set ACF3-ACR was the best for a PCR assay targeting the gyrB gene, and the PCR technique developed was rapid, specific, and sensitive for the identification of A. caviae. PMID:26223267

  14. Pet Bonding and Pet Bereavement among Adolescents.

    ERIC Educational Resources Information Center

    Brown, Brenda H.; And Others

    1996-01-01

    Studied adolescent-pet bonding and bereavement following pet loss (n=55). Hypothesized that highly-bonded adolescents experience more intense grief when a pet dies than do those less bonded; degree of bonding is greater for girls than for boys; and intensity of bereavement is greater for girls than for boys. Results supported the hypotheses. (RB)

  15. First-In-Human Study Demonstrating Tumor-Angiogenesis by PET/CT Imaging with 68Ga-NODAGA-THERANOST, a High-Affinity Peptidomimetic for αvβ3 Integrin Receptor Targeting

    PubMed Central

    Baum, Richard P.; Kulkarni, Harshad R.; Müller, Dirk; Danthi, Narasimhan; Kim, Young-Seung; Brechbiel, Martin W.

    2015-01-01

    Abstract 68Ga-NODAGA-THERANOST™ is an αvβ3 integrin antagonist and the first radiolabeled peptidomimetic to reach clinical development for targeting integrin receptors. In this first-in-human study, the feasibility of integrin receptor peptidomimetic positron emission tomography/computed tomography (PET/CT) imaging was confirmed in patients with non-small-cell lung cancer and breast cancer. Methods: Patients underwent PET/CT imaging with 68Ga NODAGA-THERANOST. PET images were analyzed qualitatively and quantitatively and compared to 2-deoxy-2-(18F) fluoro-d-glucose (18F-FDG) findings. Images were obtained 60 minutes postinjection of 300–500 MBq of 68Ga-NODAGA-THERANOST. Results: 68Ga-NODAGA-THERANOST revealed high tumor-to-background ratios (SUVmax=4.8) and uptake at neoangiogenesis sites. Reconstructed fused images distinguished cancers with high malignancy potential and enabled enhanced bone metastasis detection. 18F-FDG-positive lung and lymph node metastases did not show uptake, indicating the absence of neovascularization. Conclusions: 68Ga-NODAGA-THERANOST was found to be safe and effective, exhibiting in this study rapid blood clearance, stability, rapid renal excretion, favorable biodistribution and PK/PD, low irradiation burden (μSv/MBq/μg), and convenient radiolabeling. This radioligand might enable theranostics, that is, a combination of diagnostics followed by the appropriate therapeutics, namely antiangiogenic therapy, image-guided presurgical assessment, treatment response evaluation, prediction of pathologic response, neoadjuvant-peptidomimetic-radiochemotherapy, and personalized medicine strategies. Further clinical trials evaluating 68Ga-NODAGA-THERANOST are warranted. PMID:25945808

  16. PET/SPECT imaging agents for neurodegenerative diseases

    PubMed Central

    Zhu, Lin; Ploessl, Karl; Kung, Hank F.

    2014-01-01

    Single photon emission computed tomography (SPECT) or positron emission computed tomography (PET) imaging agents for neurodegenerative disease have a significant impact on clinical diagnosis and patient care. The examples of Parkinson’s Disease (PD) and Alzheimer’s Disease (AD) imaging agents described in this paper provide a general view on how imaging agents, ie radioactive drugs, are selected, chemically prepared and applied in humans. Imaging the living human brain can provide unique information on the pathology and progression of neurodegenerative diseases, such as AD and PD. The imaging method will also facilitate preclinical and clinical trials of new drugs offering specific information related to drug binding sites in the brain. In the future, chemists will continue to play important roles in identifying specific targets, synthesizing target-specific probes for screening and ultimately testing them by in vitro and in vivo assays. PMID:24676152

  17. A comparison of two real-time polymerase chain reaction assays using hybridization probes targeting either 16S ribosomal RNA or a subsurface lipoprotein gene for detecting leptospires in canine urine.

    PubMed

    Gentilini, Fabio; Zanoni, Renato Giulio; Zambon, Elisa; Turba, Maria Elena

    2015-11-01

    Leptospires are excreted in the urine of infected animals, and the prompt detection of leptospiral DNA using polymerase chain reaction (PCR) is increasingly being used. However, contradictory data has emerged concerning the diagnostic accuracy of the most popular PCR assays that target either the 16S ribosomal RNA (rrs) or the subsurface lipoprotein (LipL32) genes. In order to clarify the effect of the gene target, a novel hydrolysis probe-based, quantitative real-time PCR (qPCR) assay targeting the LipL32 gene was developed, validated, and then compared directly to the previously described rrs hydrolysis probe-based qPCR using a convenience collection of canine urine samples. The novel LipL32 qPCR assay was linear from 5.9 × 10(6) to 59 genome equivalents per reaction. Both the LipL32 and the rrs qPCR assays showed a limit of detection of 10 target copies per reaction indicating an approximately equivalent analytical sensitivity. Both assays amplified all 20 pathogenic leptospiral strains tested but did not amplify a representative collection of bacteria commonly found in voided canine urine. When the field samples were assayed, 1 and 5 out of 184 samples yielded an amplification signal in the LipL32 and rrs assays, respectively. Nevertheless, when the limit of detection was considered as the cutoff for interpreting findings, the 4 discordant cases were judged as negative. In conclusion, our study confirmed that both LipL32 and rrs are suitable targets for qPCR for the detection of leptospiral DNA in canine urine. However, the rrs target requires the mandatory use of a cutoff value in order to correctly interpret spurious amplifications. PMID:26450835

  18. 64Cu antibody-targeting of the T-cell receptor and subsequent internalization enables in vivo tracking of lymphocytes by PET.

    PubMed

    Griessinger, Christoph M; Maurer, Andreas; Kesenheimer, Christian; Kehlbach, Rainer; Reischl, Gerald; Ehrlichmann, Walter; Bukala, Daniel; Harant, Maren; Cay, Funda; Brück, Jürgen; Nordin, Renate; Kohlhofer, Ursula; Rammensee, Hans-Georg; Quintanilla-Martinez, Leticia; Schaller, Martin; Röcken, Martin; Pichler, Bernd J; Kneilling, Manfred

    2015-01-27

    T cells are key players in inflammation, autoimmune diseases, and immunotherapy. Thus, holistic and noninvasive in vivo characterizations of the temporal distribution and homing dynamics of lymphocytes in mammals are of special interest. Herein, we show that PET-based T-cell labeling facilitates quantitative, highly sensitive, and holistic monitoring of T-cell homing patterns in vivo. We developed a new T-cell receptor (TCR)-specific labeling approach for the intracellular labeling of mouse T cells. We found that continuous TCR plasma membrane turnover and the endocytosis of the specific (64)Cu-monoclonal antibody (mAb)-TCR complex enables a stable labeling of T cells. The TCR-mAb complex was internalized within 24 h, whereas antigen recognition was not impaired. Harmful effects of the label on the viability, DNA-damage and apoptosis-necrosis induction, could be minimized while yielding a high contrast in in vivo PET images. We were able to follow and quantify the specific homing of systemically applied (64)Cu-labeled chicken ovalbumin (cOVA)-TCR transgenic T cells into the pulmonary and perithymic lymph nodes (LNs) of mice with cOVA-induced airway delayed-type hypersensitivity reaction (DTHR) but not into pulmonary and perithymic LNs of naïve control mice or mice diseased from turkey or pheasant OVA-induced DTHR. Our protocol provides consequent advancements in the detection of small accumulations of immune cells in single LNs and specific homing to the sites of inflammation by PET using the internalization of TCR-specific mAbs as a specific label of T cells. Thus, our labeling approach is applicable to other cells with constant membrane receptor turnover. PMID:25587131

  19. Targeted Next Generation Sequencing as a Reliable Diagnostic Assay for the Detection of Somatic Mutations in Tumours Using Minimal DNA Amounts from Formalin Fixed Paraffin Embedded Material

    PubMed Central

    Koudijs, Marco J.; Nijman, Ies; Hinrichs, John W. J.; Cuppen, Edwin; van Lieshout, Stef; Loberg, Robert D.; de Jonge, Maja; Voest, Emile E.; de Weger, Roel A.; Steeghs, Neeltje; Langenberg, Marlies H. G.; Sleijfer, Stefan; Willems, Stefan M.; Lolkema, Martijn P.

    2016-01-01

    Background Targeted Next Generation Sequencing (NGS) offers a way to implement testing of multiple genetic aberrations in diagnostic pathology practice, which is necessary for personalized cancer treatment. However, no standards regarding input material have been defined. This study therefore aimed to determine the effect of the type of input material (e.g. formalin fixed paraffin embedded (FFPE) versus fresh frozen (FF) tissue) on NGS derived results. Moreover, this study aimed to explore a standardized analysis pipeline to support consistent clinical decision-making. Method We used the Ion Torrent PGM sequencing platform in combination with the Ion AmpliSeq Cancer Hotspot Panel v2 to sequence frequently mutated regions in 50 cancer related genes, and validated the NGS detected variants in 250 FFPE samples using standard diagnostic assays. Next, 386 tumour samples were sequenced to explore the effect of input material on variant detection variables. For variant calling, Ion Torrent analysis software was supplemented with additional variant annotation and filtering. Results Both FFPE and FF tissue could be sequenced reliably with a sensitivity of 99.1%. Validation showed a 98.5% concordance between NGS and conventional sequencing techniques, where NGS provided both the advantage of low input DNA concentration and the detection of low-frequency variants. The reliability of mutation analysis could be further improved with manual inspection of sequence data. Conclusion Targeted NGS can be reliably implemented in cancer diagnostics using both FFPE and FF tissue when using appropriate analysis settings, even with low input DNA. PMID:26919633

  20. Target-induced nanocatalyst deactivation facilitated by core@shell nanostructures for signal-amplified headspace-colorimetric assay of dissolved hydrogen sulfide.

    PubMed

    Gao, Zhuangqiang; Tang, Dianyong; Tang, Dianping; Niessner, Reinhard; Knopp, Dietmar

    2015-10-01

    Colorimetric assay platforms for dissolved hydrogen sulfide (H2S) have been developed for more than 100 years, but most still suffer from relatively low sensitivity. One promising route out of this predicament relies on the design of efficient signal amplification methods. Herein, we rationally designed an unprecedented H2S-induced deactivation of (gold core)@(ultrathin platinum shell) nanocatalysts (Au@TPt-NCs) as a highly efficient signal amplification method for ultrasensitive headspace-colorimetric assay of dissolved H2S. Upon target introduction, Au@TPt-NCs were deactivated to different degrees dependent on H2S levels, and the degrees could be indicated by using a Au@TPt-NCs-triggered catalytic system as a signal amplifier, thus paving a way for H2S sensing. The combination of experimental studies and density functional theory (DFT) studies revealed that the Au@TPt-NCs with only 2-monolayer equivalents of Pt (θPt = 2) were superior for H2S-induced nanocatalyst deactivation owing to their enhanced peroxidase-like catalytic activity and deactivation efficiency stemmed from the unique synergistic structural/electronic effects between Au nanocores and ultrathin Pt nanoshells. Importantly, our analytical results showed that the designed method was indeed highly sensitive for sensing H2S with a wide linear range of 10-100 nM, a slope of 0.013 in the regression equation, and a low detection limit of 7.5 nM. Also the selectivity, reproducibility, and precision were excellent. Furthermore, the method was validated for the analysis of H2S-spiked real samples, and the recovery in all cases was 91.6-106.7%. With the merits of high sensitivity and selectivity, simplification, low cost, and visual readout with the naked eye, the colorimetric method has the potential to be utilized as an effective detection kit for point-of-care testing. PMID:26327230

  1. PET/MRI and PET/CT in Lung Lesions and Thoracic Malignancies.

    PubMed

    Flechsig, Paul; Mehndiratta, Amit; Haberkorn, Uwe; Kratochwil, Clemens; Giesel, Frederik L

    2015-07-01

    More than one decade ago, introduction of integrated PET/CT scanners changed oncologic imaging and oncologic patient management profoundly. With these systems, the metabolic information acquired by PET can be anatomically localized even to small structures such as small primary tumors, lymph nodes, and soft tissue masses owing to the high-resolution multidetector CT scanners. This has made PET/CT a most reliable method for tumor detection, characterization, staging, and response monitoring. The importance of an integrated functional and morphologic approach to better understand the biology of oncologic disease and to improve therapy planning is underlined by the increasing number of PET/CT systems worldwide, leading to an increasing number of scientific publications in the field. The paradigmatic indication of integrated PET/CT is staging of patients with lung cancer, as PET/CT allows for precise pretherapeutic staging and also posttreatment restaging according to the TNM criteria. The growing numbers of targeted therapy strategies in the fields of surgery, chemotherapy, and radiation therapy, which are adapted to dedicated tumor stages, require the exact classifications of each patient's tumor stage. In this context, whole-body examinations using integrated (18)F-FDG-PET/CT have been shown to reduce the side effects of futile invasive procedures and reduce additional costly staging procedures. In this review article, the diagnostic and therapeutic effects of PET/CT examinations are highlighted and compared with some competitive techniques such as scintigraphy, MRI, and, where possible, integrated PET/MRI. PMID:26050655

  2. Hybrid PET/MR imaging: physics and technical considerations.

    PubMed

    Shah, Shetal N; Huang, Steve S

    2015-08-01

    In just over a decade, hybrid imaging with FDG PET/CT has become a standard bearer in the management of cancer patients. An exquisitely sensitive whole-body imaging modality, it combines the ability to detect subtle biologic changes with FDG PET and the anatomic information offered by CT scans. With advances in MR technology and advent of novel targeted PET radiotracers, hybrid PET/MRI is an evolutionary technique that is poised to revolutionize hybrid imaging. It offers unparalleled spatial resolution and functional multi-parametric data combined with biologic information in the non-invasive detection and characterization of diseases, without the deleterious effects of ionizing radiation. This article reviews the basic principles of FDG PET and MR imaging, discusses the salient technical developments of hybrid PET/MR systems, and provides an introduction to FDG PET/MR image acquisition. PMID:25985965

  3. Discovery of Phosphodiesterase 10A (PDE10A) PET Tracer AMG 580 to Support Clinical Studies.

    PubMed

    Hu, Essa; Chen, Ning; Kunz, Roxanne K; Hwang, Dah-Ren; Michelsen, Klaus; Davis, Carl; Ma, Ji; Shi, Jianxia; Lester-Zeiner, Dianna; Hungate, Randall; Treanor, James; Chen, Hang; Allen, Jennifer R

    2016-07-14

    We report the discovery of PDE10A PET tracer AMG 580 developed to support proof of concept studies with PDE10A inhibitors in the clinic. To find a tracer with higher binding potential (BPND) in NHP than our previously reported tracer 1, we implemented a surface plasmon resonance assay to measure the binding off-rate to identify candidates with slower washout rate in vivo. Five candidates (2-6) from two structurally distinct scaffolds were identified that possessed both the in vitro characteristics that would favor central penetration and the structural features necessary for PET isotope radiolabeling. Two cinnolines (2, 3) and one keto-benzimidazole (5) exhibited PDE10A target specificity and brain uptake comparable to or better than 1 in the in vivo LC-MS/MS kinetics distribution study in SD rats. In NHP PET imaging study, [(18)F]-5 produced a significantly improved BPND of 3.1 and was nominated as PDE10A PET tracer clinical candidate for further studies. PMID:27437084

  4. Development of an HTS assay for the search of anti-influenza agents targeting the interaction of viral RNA with the NS1 protein.

    PubMed

    Maroto, Marta; Fernandez, Yolanda; Ortin, Juan; Pelaez, Fernando; Cabello, M Angerles

    2008-08-01

    The NS1 protein is a nonstructural protein encoded by the influenza A virus. It is responsible for many alterations produced in the cellular metabolism upon infection by the virus and for modulation of virus virulence. The NS1 protein is able to perform a large variety of functions due to its ability to bind various types of RNA molecules, from both viral and nonviral origin, and to interact with several cell factors. With the aim of exploring whether the binding of NS1 protein to viral RNA (vRNA) could constitute a novel target for the search of anti-influenza drugs, a filter-binding assay measuring the specific interaction between the recombinant His-NS1 protein from influenza A virus and a radiolabeled model vRNA ( 32P-vNSZ) was adapted to a format suitable for screening and easy automation. Flashplate technology (PerkinElmer, Waltham, MA), either in 96- or 384-well plates, was used. The Flashplate wells were precoated with the recombinant His-NS1 protein, and the binding of His-NS1 to a 35S-vNSZ probe was measured. A pilot screening of a collection of 27,520 mixtures of synthetic chemical compounds was run for inhibitors of NS1 binding to vRNA. We found 3 compounds in which the inhibition of NS1 binding to vRNA, observed at submicromolar concentrations, was correlated with a reduction of the cytopathic effect during the infection of cell cultures with influenza virus. These results support the hypothesis that the binding of NS1 to vRNA could be a novel target for the development of anti-influenza drugs. PMID:18594021

  5. Trends in PET imaging

    SciTech Connect

    Moses, William W.

    2000-11-01

    Positron Emission Tomography (PET) imaging is a well established method for obtaining information on the status of certain organs within the human body or in animals. This paper presents an overview of recent trends PET instrumentation. Significant effort is being expended to develop new PET detector modules, especially those capable of measuring depth of interaction. This is aided by recent advances in scintillator and pixellated photodetector technology. The other significant area of effort is development of special purpose PET cameras (such as for imaging breast cancer or small animals) or cameras that have the ability to image in more than one modality (such as PET / SPECT or PET / X-Ray CT).

  6. Laboratory utility of coproscopy, copro immunoassays and copro nPCR assay targeting Hsp90 gene for detection of Cryptosporidium in children, Cairo, Egypt.

    PubMed

    Ghallab, Marwa M I; Aziz, Inas Z Abdel; Shoeib, Eman Y; El-Badry, Ayman A

    2016-09-01

    Cryptosporidium is a significant cause of diarrhea worldwide especially in children. Infection may end fatally in immunocompromised patients. Multi-attribute analysis was used to determine the lab utility of 4 diagnostics; coproscopy of AF stained fecal smear, fecal immunoassays by ICT and ELISA and copro-nPCR assay targeting Hsp90 gene, for detection of Cryptosporidium in stool of 250 Egyptian children (150 diarrheic and 100 non-diarrhaeic children). Also, to determine Cryptosporidium molecular prevalence. Cryptosporidium was an important enteric pathogen among both diarrheic and non-diarrheic study children with a clearly high prevalence of 16.4 % (n = 41). Conventional methods had perfect specificity (100 %) but couldn`t be used as a consistent single detection method due to their lowered sensitivities. Multi-attribute analysis ranked nPCR the highest test for lab use. Being the test with the best diagnostic yield, nPCR is a reliable diagnostic test and is going to replace conventional methods for reliable detection of Cryptosporidium. PMID:27605806

  7. Development of a Medium-Throughput Targeted LCMS Assay to Detect Endogenous Cellular Levels of Malonyl-CoA to Screen Fatty Acid Synthase Inhibitors.

    PubMed

    Hopcroft, Philip J; Fisher, David I

    2016-02-01

    The fatty acid synthase (FAS) enzyme in mammalian cells is a large multidomain protein responsible for de novo synthesis of fatty acids. The steps catalyzed by FAS involve the condensation of acetyl-CoA and malonyl-CoA moieties in the presence of NADPH until palmitate is formed. Inhibition of FAS causes an accumulation of intracellular malonyl-CoA, as this metabolite is essentially committed to fatty acid synthesis once formed. Detection of intracellular metabolites for screening can be problematic due to a lack of appropriate tools, but here we describe a targeted liquid chromatography-mass spectroscopy (LCMS) method to directly measure endogenous levels of malonyl-CoA to drive a drug development structure-activity relationship (SAR) screening cascade. Our process involves preparation of samples at 96-well scale, normalization postpermeabilization via use of a whole-well imaging platform, and the LCMS detection methodology. The assay is amenable to multiplexing cellular endpoints, has a typical Z' of >0.6, and has high reproducibility of EC50 values. PMID:26586251

  8. Label-free assay for the assessment of nonspecific binding of positron emission tomography tracer candidates.

    PubMed

    Assmus, Frauke; Seelig, Anna; Gobbi, Luca; Borroni, Edilio; Glaentzlin, Patricia; Fischer, Holger

    2015-11-15

    Positron emission tomography (PET) is a valuable non-invasive technique for the visualization of drug tissue distribution and receptor occupancy at the target site in living animals and men. Many potential PET tracers, however, fail due to an unfavorably high non-specific binding (NSB) to non-target proteins and phospholipid membranes which compromises the sensitivity of PET. Hence, there is a high demand to assess the extent of NSB as early as possible in the PET tracer development process, preferentially before ligands are radiolabeled and elaborate imaging studies are performed. The purpose of this study was to establish a novel Lipid Membrane Binding Assay (LIMBA) for assessing the tendency of potential tracers to bind non-specifically to brain tissue. The assay works with unlabeled compounds and allows the medium-throughput measurement of brain tissue/water distribution coefficients, logDbrain (pH7.4), at minimal expense of animal tissue. To validate LIMBA, logDbrain (pH7.4) values were measured and compared with NSB estimates derived from in vivo PET studies in human brain (n=10 tracers, literature data), and in vitro autoradiography studies in rat and mouse brain slices (n=30 tritiated radioligands). Good agreement between logDbrain (pH7.4) and the volume of distribution in brain of non-specifically bound tracer in PET was achieved, pertaining to compounds classified as non-substrates of P-glycoprotein (R(2)≥0.88). The ability of LIMBA for the prediction of NSB was further supported by the strong correlation between logDbrain (pH7.4) and NSB in brain autoradiography (R(2)≥0.76), whereas octanol/water distribution coefficients, logDoct (pH7.4) were less predictive. In conclusion, LIMBA provides a fast and reliable tool for identifying compounds with unfavorably high NSB in brain tissue. The data may be used in conjunction with other parameters like target affinity, density and membrane permeability for the selection of most promising compounds to be

  9. Heart PET scan

    MedlinePlus

    ... nuclear medicine scan; Heart positron emission tomography; Myocardial PET scan ... A PET scan requires a small amount of radioactive material (tracer). This tracer is given through a vein (IV), ...

  10. Breast PET scan

    MedlinePlus

    ... medlineplus.gov/ency/article/007469.htm Breast PET scan To use the sharing features on this page, ... enable JavaScript. A breast positron emission tomography (PET) scan is an imaging test that uses a radioactive ...

  11. Advanced Tracers in PET Imaging of Cardiovascular Disease

    PubMed Central

    Zhang, Wei; Wu, Hua; Liu, Gang

    2014-01-01

    Cardiovascular disease is the leading cause of death worldwide. Molecular imaging with targeted tracers by positron emission tomography (PET) allows for the noninvasive detection and characterization of biological changes at the molecular level, leading to earlier disease detection, objective monitoring of therapies, and better prognostication of cardiovascular diseases progression. Here we review, the current role of PET in cardiovascular disease, with emphasize on tracers developed for PET imaging of cardiovascular diseases. PMID:25389529

  12. 2-(2’-((Dimethylamino)methyl)-4’-(3-[18F]fluoropropoxy)-phenylthio)benzenamine for PET Imaging of Serotonin Transporters

    PubMed Central

    Wang, Julie L.; Parhi, Ajit K.; Oya, Shunichi; Lieberman, Brian; Kung, Mei-Ping; Kung, Hank F.

    2008-01-01

    Introduction A new 18F ligand, 2-(2’-((dimethylamino)methyl)-4’-(3-[18F]fluoropropoxy)-phenylthio)benzenamine ([18F]1), for positron emission tomography (PET) imaging of serotonin transporters (SERT) was evaluated. Methods Binding affinity was determined through in vitro binding assays with LLC-PK1 cells overexpressing SERT, NET, or DAT (LLC-SERT, LLC-NET, and LLC-DAT) and with rat cortical homogenates. Localization and selectivity of [18F]1_binding in vivo was evaluated by biodistribution, autoradiography, and A-PET imaging studies in rats. Results This compound displayed excellent binding affinity for SERT in vitro with Ki = 0.33 and 0.24 nM in LLC- SERT and rat cortical homogenates, respectively. Biodistribution studies with [18F]1 showed good brain uptake (1.61% dose/g at 2 min post-injection), high uptake into the hypothalamus (1.22% dose/g at 30 min), and a high target to non-target (hypothalamus to cerebellum) ratio of 9.66 at 180 min post-injection. Pretreatment with a SERT selective inhibitor considerably inhibited [18F]1_binding in biodistribution studies. Ex vivo autoradiography reveals [18F]1 localization to brain regions with high SERT density, and this binding was blocked by pretreatment with SERT selective inhibitors. Small animal PET (A-PET) imaging in rats provided clear images of tracer localization in the thalamus, midbrain, and striatum. In A-PET chasing experiments, injecting a SERT selective inhibitor 75 min post-tracer injection causes a dramatic reduction in regional radioactivity and the target to non-target ratio. Conclusion The results of the biological studies and the ease of radiosynthesis with moderately good radiochemical yield (RCY = 10–35%) make [18F]1 an excellent candidate for SERT PET imaging. PMID:18482682

  13. Synthesis of Clinical-Grade [(18)F]-Fluoroestradiol as a Surrogate PET Biomarker for the Evaluation of Estrogen Receptor-Targeting Therapeutic Drug.

    PubMed

    Dixit, Manish; Shi, Jianfeng; Wei, Ling; Afari, George; Bhattacharyya, Sibaprasad

    2013-01-01

    16 α -[(18)F]-fluoroestradiol ([(18)F]FES), a steroid-based positron emission tomography (PET) tracer, has emerged as a dependable tracer for the evaluation and management of estrogen receptor-positive (ER+) breast cancer patients. We have developed a fully automatic, one-pot procedure for the synthesis of [(18)F]FES using the Eckert & Ziegler (E & Z) radiomodular system. After [(18)F]fluorination, the intermediate was hydrolyzed with 2.0 M HCl twice and neutralized with sodium bicarbonate. After high-performance liquid chromatography (HPLC) purification, the decay-corrected radiochemical yield and purity of [(18)F]FES were 40 ± 5.0% (n = 12) and >97%, respectively. The product was stable up to 10 h. Total synthesis time including HPLC purification was 80 min. This new, fully automated rapid synthetic procedure provided high and reproducible yields of [(18)F]FES. Quality control (QC) tests showed that the [(18)F]FES produced by this method met all specifications for human injection. PMID:23762549

  14. Birds Kept as Pets

    MedlinePlus

    ... restricts the importation of pet birds from certain countries and enforces a 30-day quarantine for all imported birds except those that come from Canada. People interested in importing pet birds should visit the USDA non-US Origin Pet Bird Importation website . Choosing a bird Match ...

  15. Performance and Verification of a Real-Time PCR Assay Targeting the gyrA Gene for Prediction of Ciprofloxacin Resistance in Neisseria gonorrhoeae.

    PubMed

    Hemarajata, P; Yang, S; Soge, O O; Humphries, R M; Klausner, J D

    2016-03-01

    In the United States, 19.2% of Neisseria gonorrhoeae isolates are resistant to ciprofloxacin. We evaluated a real-time PCR assay to predict ciprofloxacin susceptibility using residual DNA from the Roche Cobas 4800 CT/NG assay. The results of the assay were 100% concordant with agar dilution susceptibility test results for 100 clinical isolates. Among 76 clinical urine and swab specimens positive for N. gonorrhoeae by the Cobas assay, 71% could be genotyped. The test took 1.5 h to perform, allowing the physician to receive results in time to make informed clinical decisions. PMID:26739156

  16. Proteomic Discovery and Development of a Multiplexed Targeted MRM-LC-MS/MS Assay for Urine Biomarkers of Extracellular Matrix Disruption in Mucopolysaccharidoses I, II, and VI.

    PubMed

    Heywood, Wendy E; Camuzeaux, Stephane; Doykov, Ivan; Patel, Nina; Preece, Rhian-Lauren; Footitt, Emma; Cleary, Maureen; Clayton, Peter; Grunewald, Stephanie; Abulhoul, Lara; Chakrapani, Anupam; Sebire, Neil J; Hindmarsh, Peter; de Koning, Tom J; Heales, Simon; Burke, Derek; Gissen, Paul; Mills, Kevin

    2015-12-15

    The mucopolysaccharidoses (MPS) are lysosomal storage disorders that result from defects in the catabolism of glycosaminoglycans. Impaired muscle, bone, and connective tissue are typical clinical features of MPS due to disruption of the extracellular matrix. Markers of MPS disease pathology are needed to determine disease severity and monitor effects of existing and emerging new treatments on disease mechanisms. Urine samples from a small cohort of MPS-I, -II, and -VI patients (n = 12) were analyzed using label-free quantative proteomics. Fifty-three proteins including many associated with extracellular matrix organization were differently expressed. A targeted multiplexed peptide MRM LC-MS/MS assay was used on a larger validation cohort of patient samples (MPS-I n = 18, MPS-II n = 12, MPS-VI n = 6, control n = 20). MPS-I and -II groups were further subdivided according to disease severity. None of the markers assessed were altered significantly in the mild disease groups compared to controls. β-galactosidase, a lysosomal protein, was elevated 3.6-5.7-fold significantly (p < 0.05) in all disease groups apart from mild MPS-I and -II. Collagen type Iα, fatty-acid-binding-protein 5, nidogen-1, cartilage oligomeric matrix protein, and insulin-like growth factor binding protein 7 concentrations were elevated in severe MPS I and II groups. Cartilage oligomeric matrix protein, insulin-like growth factor binding protein 7, and β-galactosidase were able to distinguish the severe neurological form of MPS-II from the milder non-neurological form. Protein Heg1 was significantly raised only in MPS-VI. This work describes the discovery of new biomarkers of MPS that represent disease pathology and allows the stratification of MPS-II patients according to disease severity. PMID:26537538

  17. A cultivation-independent PCR-RFLP assay targeting oprF gene for detection and identification of Pseudomonas spp. in samples from fibrocystic pediatric patients.

    PubMed

    Lagares, Antonio; Agaras, Betina; Bettiol, Marisa P; Gatti, Blanca M; Valverde, Claudio

    2015-07-01

    Species-specific genetic markers are crucial to develop faithful and sensitive molecular methods for the detection and identification of Pseudomonas aeruginosa (Pa). We have previously set up a PCR-RFLP protocol targeting oprF, the gene encoding the genus-specific outer membrane porin F, whose strong conservation and marked sequence diversity allowed detection and differentiation of environmental isolates (Agaras et al., 2012). Here, we evaluated the ability of the PCR-RFLP assay to genotype clinical isolates previously identified as Pa by conventional microbiological methods within a collection of 62 presumptive Pa isolates from different pediatric clinical samples and different sections of the Hospital de Niños "Sor María Ludovica" from La Plata, Argentina. All isolates, but one, gave an oprF amplicon consistent with that from reference Pa strains. The sequence of the smaller-sized amplicon revealed that the isolate was in fact a mendocina Pseudomonas strain. The oprF RFLP pattern generated with TaqI or HaeIII nucleases matched those of reference Pa strains for 59 isolates (96%). The other two Pa isolates (4%) revealed a different RFLP pattern based on HaeIII digestion, although oprF sequencing confirmed that Pa identification was correct. We next tested the effectiveness of the PCR-RFLP to detect pseudomonads on clinical samples of pediatric fibrocystic patients directly without sample cultivation. The expected amplicon and its cognate RFLP profile were obtained for all samples in which Pa was previously detected by cultivation-dependent methods. Altogether, these results provide the basis for the application of the oprF PCR-RFLP protocol to directly detect and identify Pa and other non-Pa pseudomonads in fibrocystic clinical samples. PMID:25960432

  18. Ex vivo cytotoxic drug evaluation by DiSC assay to expedite identification of clinical targets: results with 8-chloro-cAMP.

    PubMed Central

    Bosanquet, A. G.; Burlton, A. R.; Bell, P. B.; Harris, A. L.

    1997-01-01

    There is a pressing need to reduce the time and cost of developing new cytotoxic agents and to accurately identify clinically active agents at an early stage. In this study, the differential staining cytotoxicity (DiSC) assay was used to assess the efficacy of the novel antitumour cAMP analogue, 8-chloro-cAMP (8-Cl-cAMP) (and its metabolite 8-Cl-adenosine) against 107 fresh specimens of human neoplastic and normal cells. Diagnoses included chronic and acute leukaemias, myeloma, non-Hodgkin's lymphoma (NHL) and miscellaneous solid tumours. The aim was to identify targets for subsequent phase I, II and III trials. 8-Cl-cAMP was tested at 4-985 microM, along with standard chemotherapeutic drugs. 8-Cl-cAMP and its metabolite caused no morphologically observable cell differentiation but induced dose-dependent cytotoxicity. Compared with untreated patients, previously treated chronic lymphocytic leukaemia (CLL) patients showed no increase in ex vivo resistance to 8-Cl-cAMP (P = 0.878); minimal cross-resistance with other cytotoxic drugs was detected. Compared with normal cells (mean LC90 = 1803 microM), 8-Cl-cAMP showed significant ex vivo activity against CLL (117.0 microM; P < 0.0001) and NHL (140.0 microM; P < 0.0001), of which eight were mantle cell NHL (84.7 microM), and greatest activity against cells from patients with acute myeloid leukaemia (AML; mean LC90 = 24.3 microM; in vitro therapeutic index 74-fold, P < 0.0001). Solid tumour specimens were comparatively resistant to 8-Cl-cAMP. The results highlight the clinical potential of 8-Cl-cAMP, point to several new phase I, II and III trial possibilities and provide a rationale for the inclusion of ex vivo cytotoxic drug evaluation in the drug development process. PMID:9275029

  19. Validation of a High-Throughput Screening Assay for Identification of Adjunctive and Directly Acting Antimicrobials Targeting Carbapenem-Resistant Enterobacteriaceae.

    PubMed

    Smith, Kenneth P; Kirby, James E

    2016-04-01

    We describe development and validation of a high-throughput screen (HTS) for identifying small molecules that restore the efficacy of carbapenems (adjunctives) and/or directly inhibit growth of carbapenem-resistant Enterobacteriaceae (CRE). Our HTS assay is based on a screen-counterscreen approach using a representative multidrug-resistant CRE strain, Klebsiella pneumoniae BIDMC12A. Specifically, we tested the ability of small molecules to inhibit bacterial growth in the presence (screen) or absence (counterscreen) of meropenem, a representative carbapenem antibiotic. Primary screening of 11,698 known bioactive compounds identified 14 with adjunctive activity and 79 with direct antimicrobial effect. Secondary screening identified triclosan as a strongly synergistic meropenem adjunctive (fractional inhibitory concentration = 0.48) and confirmed azidothymidine (AZT) (minimal inhibitory concentration [MIC] = 4 μg mL(-1)), NH125 (MIC = 4 μg mL(-1)), diphenyleneiodonium chloride (MIC = 8 μg mL(-1)), and spectinomycin (MIC = 32 μg mL(-1)) as potent direct antimicrobials. Spectrum of activity of AZT and spectinomycin was tested against a collection of 103 representative Enterobacteriaceae strains (≈50% CRE). AZT, a nucleoside analog used to treat human immunodeficiency virus, demonstrated an MIC50 of 2 μg mL(-1). Spectinomycin, an antibiotic used to treat gonorrhea, had an MIC50 of 32 μg mL(-1). Therefore, a significant percentage of CRE strains appeared relatively susceptible to these antimicrobials. These data identified AZT and spectinomycin as available agents warranting further study for potential treatment of multidrug-resistant CRE infection. Our results provide proof of principle and impetus for performing a large-scale HTS for discovery of novel, small-molecule adjunctives and antibacterial agents directly targeting CRE. PMID:27045615

  20. Molecular assays for targeting human and bovine enteric viruses in coastal waters and their application for library-independent source tracking

    USGS Publications Warehouse

    Fong, T.-T.; Griffin, Dale W.; Lipp, E.K.

    2005-01-01

    Rapid population growth and urban development along waterways and coastal areas have led to decreasing water quality. To examine the effects of upstream anthropogenic activities on microbiological water quality, methods for source-specific testing are required. In this study, molecular assays targeting human enteroviruses (HEV), bovine enteroviruses (BEV), and human adenoviruses (HAdV) were developed and used to identify major sources of fecal contamination in the lower Altamaha River, Georgia. Two-liter grab samples were collected monthly from five tidally influenced stations between July and December 2002. Samples were analyzed by reverse transcription- and nested-PCR. PCR results were confirmed by dot blot hybridization. Eleven and 17 of the 30 surface water samples tested positive for HAdV and HEV, respectively. Two-thirds of the samples tested positive for either HEV or HAdV, and the viruses occurred simultaneously in 26% of samples. BEV were detected in 11 of 30 surface water samples. Binary logistic regression analysis showed that the presence of both human and bovine enteric viruses was not significantly related to either fecal coliform or total coliform levels. The presence of these viruses was directly related to dissolved oxygen and streamflow but inversely related to water temperature, rainfall in the 30 days preceding sampling, and chlorophyll-?? concentrations. The stringent host specificity of enteric viruses makes them good library-independent indicators for identification of water pollution sources. Viral pathogen detection by PCR is a highly sensitive and easy-to-use tool for rapid assessment of water quality and fecal contamination when public health risk characterization is not necessary. Copyright ?? 2005, American Society for Microbiology. All Rights Reserved.

  1. (18)F-FDG PET/CT for Monitoring the Response of Breast Cancer to miR-143-Based Therapeutics by Targeting Tumor Glycolysis.

    PubMed

    Miao, Ying; Zhang, Ling-Fei; Guo, Rui; Liang, Sheng; Zhang, Min; Shi, Shuo; Shang-Guan, Cheng-Fang; Liu, Mo-Fang; Li, Biao

    2016-01-01

    Increased glucose utilization is a hallmark of cancer, and tumor metabolism is emerging as anticancer target for therapeutic intervention. Triple-negative breast cancers TNBC are highly glycolytic and show poor clinical outcomes. We previously identified hexokinase 2, the major glycolytic enzyme, as a target gene of miR-143 in TNBC. Here, we developed a therapeutic formulation using cholesterol-modified miR-143 agomir encapsulated in a neutral lipid-based delivery agent that blocked tumor growth and glucose metabolism in TNBC tumor-bearing mice when administered systemically. The antioncogenic effects were accompanied by a reduction in the direct target hexokinase 2 and [(18)F]-fluorodeoxyglucose ((18)F-FDG) uptake based on positron emission tomography/computed tomography. Treatment with miR-143 formulation has minimal toxic effects and mice tolerated it well. Thus, we demonstrated that miR-143 is a robust inhibitor of the Warburg effect and an effective therapeutic target for TNBC. In addition, (18)F-FDG positron emission tomography/computed tomography can be used to specifically monitor the response of TNBC to miR-143-based therapeutics by targeting tumor glycolysis. PMID:27574783

  2. Comparative Study of a Real-Time PCR Assay Targeting senX3-regX3 versus Other Molecular Strategies Commonly Used in the Diagnosis of Tuberculosis

    PubMed Central

    Sanjuan-Jimenez, Rocio; Toro-Peinado, Inmaculada; Bermudez, Pilar; Colmenero, Juan D.; Morata, Pilar

    2015-01-01

    Background Nucleic acid amplification tests are increasingly used for the rapid diagnosis of tuberculosis. We undertook a comparative study of the efficiency and diagnostic yield of a real-time PCR senX3-regX3 based assay versus the classical IS6110 target and the new commercial methods. Methods This single-blind prospective comparative study included 145 consecutive samples: 76 from patients with culture-confirmed tuberculosis (86.8% pulmonary and 13.2% extrapulmonary tuberculosis: 48.7% smear-positive and 51.3% smear-negative) and 69 control samples (24 from patients diagnosed with non-tuberculous mycobacteria infections and 45 from patients with suspected tuberculosis which was eventually ruled out). All samples were tested by two CE-marked assays (Xpert®MTB/RIF and AnyplexTM plus MTB/NTM) and two in-house assays targeting senX3-regX3 and the IS6110 gene. Results The detection limit ranged from 1.00E+01 fg for Anyplex, senX3-regX3 and IS6110 to 1.00E+04 fg for Xpert. All three Xpert, senX3-regX3 and IS6110 assays detected all 37 smear-positive cases. Conversely, Anyplex was positive in 34 (91.9%) smear-positive cases. In patients with smear-negative tuberculosis, differences were observed between the assays; Xpert detected 22 (56.41%) of the 39 smear-negative samples, Anyplex 24 (61.53%), senX3-regX3 28 (71.79%) and IS6110 35 (89.74%). Xpert and senX3-regX3 were negative in all control samples; however, the false positive rate was 8.7% and 13% for Anyplex and IS6110, respectively. The overall sensitivity was 77.6%, 85.7%, 77.3% and 94.7% and the specificity was 100%, 100%, 90.8% and 87.0% for the Xpert, senX3-regX3, Anyplex and IS6110 assays, respectively. Conclusion Real-time PCR assays targeting IS6110 lack the desired specificity. The Xpert MTB/RIF and in-house senX3-regX3 assays are both sensitive and specific for the detection of MTBC in both pulmonary and extrapulmonary samples. Therefore, the real time PCR senX3-regX3 based assay could be a useful and

  3. FDG and (82)Rb PET/MRI features of brain metastasis of breast cancer.

    PubMed

    Lu, Yang

    2015-06-01

    A 64-year-old woman with stage IV breast cancer underwent an FDG and Rb PET brain studies. The PET brain images were fused with MRI brain T1 post-contrast images. The known enhancing left superoposterior frontal brain metastasis is positive on both FDG Rb PET/MRI images. The Rb PET/MRI showed better target-to-noise ratio, but showed nonspecific uptake in the superior sagittal sinus. PMID:25674864

  4. The fluorescent two-hybrid assay to screen for protein-protein interaction inhibitors in live cells: targeting the interaction of p53 with Mdm2 and Mdm4.

    PubMed

    Yurlova, Larisa; Derks, Maarten; Buchfellner, Andrea; Hickson, Ian; Janssen, Marc; Morrison, Denise; Stansfield, Ian; Brown, Christopher J; Ghadessy, Farid J; Lane, David P; Rothbauer, Ulrich; Zolghadr, Kourosh; Krausz, Eberhard

    2014-04-01

    Protein-protein interactions (PPIs) are attractive but challenging targets for drug discovery. To overcome numerous limitations of the currently available cell-based PPI assays, we have recently established a fully reversible microscopy-assisted fluorescent two-hybrid (F2H) assay. The F2H assay offers a fast and straightforward readout: an interaction-dependent co-localization of two distinguishable fluorescent signals at a defined spot in the nucleus of mammalian cells. We developed two reversible F2H assays for the interactions between the tumor suppressor p53 and its negative regulators, Mdm2 and Mdm4. We then performed a pilot F2H screen with a subset of compounds, including small molecules (such as Nutlin-3) and stapled peptides. We identified five cell-penetrating compounds as potent p53-Mdm2 inhibitors. However, none exhibited intracellular activity on p53-Mdm4. Live cell data generated by the F2H assays enable the characterization of stapled peptides based on their ability to penetrate cells and disrupt p53-Mdm2 interaction as well as p53-Mdm4 interaction. Here, we show that the F2H assays enable side-by-side analysis of substances' dual Mdm2-Mdm4 activity. In addition, they are suitable for testing various types of compounds (e.g., small molecules and peptidic inhibitors) and concurrently provide initial data on cellular toxicity. Furthermore, F2H assays readily allow real-time visualization of PPI dynamics in living cells. PMID:24476585

  5. An Improved Multiplex Real-Time SYBR Green PCR Assay for Analysis of 24 Target Genes from 16 Bacterial Species in Fecal DNA Samples from Patients with Foodborne Illnesses.

    PubMed

    Kawase, Jun; Etoh, Yoshiki; Ikeda, Tetsuya; Yamaguchi, Keiji; Watahiki, Masanori; Shima, Tomoko; Kameyama, Mitsuhiro; Horikawa, Kazumi; Fukushima, Hiroshi; Goto, Ryoichi; Shirabe, Komei

    2016-05-20

    Here, we developed a new version of our original screening system (Rapid Foodborne Bacterial Screening 24; RFBS24), which can simultaneously detect 24 genes of foodborne pathogens in fecal DNA samples. This new version (RFBS24 ver. 5) detected all known stx2 subtypes, enterotoxigenic Escherichia coli (STh genotype), and Vibrio parahaemolyticus (trh2), which were not detected by the original RFBS24 assay. The detection limits of RFBS24 ver. 5 were approximately 5.6 × 10(-2)-5.6 × 10(-5) (ng DNA)/reaction, significantly lower (10- to 100-fold) than those of the original RFBS24 for the 22 target genes analyzed here. We also tested the new assay on fecal DNA samples from patients infected with Salmonella, Campylobacter, or enterohemorrhagic E. coli. The number of bacterial target genes detected by RFBS24 ver. 5 was greater than that detected by RFBS24. RFBS24 ver. 5 combined with an Ultra Clean Fecal DNA Isolation Kit showed adequate performance (sensitivity and specificity 89% and 100%, respectively, for Salmonella spp. and 100% and 83%, respectively, for Campylobacter jejuni) in terms of rapid detection of a causative pathogen during foodborne-illness outbreaks. Thus, RFBS24 ver. 5 is more useful than the previous assay system for detection of foodborne pathogens and offers quick simultaneous analysis of many targets and thus facilitates rapid dissemination of information to public health officials. PMID:26166499

  6. Immunochromatographic assay on thread.

    PubMed

    Zhou, Gina; Mao, Xun; Juncker, David

    2012-09-18

    Lateral-flow immunochromatographic assays are low-cost, simple-to-use, rapid tests for point-of-care screening of infectious diseases, drugs of abuse, and pregnancy. However, lateral flow assays are generally not quantitative, give a yes/no answer, and lack multiplexing. Threads have recently been proposed as a support for transporting and mixing liquids in lateral-flow immunochromatographic assays, but their use for quantitative high-sensitivity immunoassays has yet to be demonstrated. Here, we introduce the immunochromatographic assay on thread (ICAT) in a cartridge format that is suitable for multiplexing. The ICAT is a sandwich assay performed on a cotton thread knotted to a nylon fiber bundle, both of which are precoated with recognition antibodies against one target analyte. Upon sample application, the assay results become visible to the eye within a few minutes and are quantified using a flatbed scanner. Assay conditions were optimized, the binding curves for C-reactive protein (CRP) in buffer and diluted serum were established and a limit of detection of 377 pM was obtained. The possibility of multiplexing was demonstrated using three knotted threads coated with antibodies against CRP, osteopontin, and leptin proteins. The performance of the ICAT was compared with that of the paper-based and conventional assays. The results suggest that thread is a suitable support for making low-cost, sensitive, simple-to-use, and multiplexed diagnostic tests. PMID:22889381

  7. Simultaneous {sup 68}Ga-DOTATOC-PET/MRI for IMRT Treatment Planning for Meningioma: First Experience

    SciTech Connect

    Thorwarth, Daniela; Henke, Guido; Mueller, Arndt-Christian; Reimold, Matthias; Beyer, Thomas; Boss, Andreas; Kolb, Armin; Pichler, Bernd; Pfannenberg, Christina

    2011-09-01

    Purpose: To evaluate intensity-modulated radiotherapy (IMRT) treatment planning based on simultaneous positron-emission tomography and magnetic resonance imaging (PET/MRI) of meningioma. Methods and Materials: A meningioma patient was examined prior to radiotherapy with dedicated planning computed tomography (CT), MRI, PET/CT with gallium-68-labeled DOTATOC ({sup 68}Ga-DOTATOC), and simultaneous {sup 68}Ga-DOTATOC-PET/MRI. The first gross target volume (GTV) was defined based on a combination of separate MR and {sup 68}Ga-DOTATOC-PET/CT imaging (GTV{sub PET/CT+MR}). Then, the simultaneous PET/MR images were used to delineate a second GTV (GTV{sub PET/MR}) by following exactly the same delineation strategy. After an isotropic expansion of those volumes by a 4-mm safety margin, the resulting planning target volumes (PTVs) were compared by calculating the intersection volume and the relative complements. A cross-evaluation of IMRT plans was performed, where the treatment plan created for the PTV{sub PET/CT+MR} was applied to the PET/MR-based PTV{sub PET/MR}. Results: Generally, target volumes for IMRT treatment planning did not differ between MRI plus {sup 68}Ga-DOTATOC-PET/CT and simultaneous PET/MR imaging. Only in certain regions of the GTV were differences observed. The overall volume of the PET/MR-based PTV was approximately the same as that obtained from PET/CT data. A small region of infiltrative tumor growth next to the main tumor mass was better visualized with combined PET/MR due to smaller PET voxel sizes and improved recovery. An IMRT treatment plan was optimized for the PTV{sub PET/CT+MR}. The evaluation of this plan with respect to the PTV{sub PET/MR} showed parts of the target volume that would not have received the full radiation dose after delineation of the tumor, based on simultaneous PET/MR. Conclusion: This case showed that differences in target volumes delineated on the basis of separate MR and PET/CT and simultaneous PET/MR may be observed that

  8. Efficiency gains in tracer identification for nuclear imaging: can in vivo LC-MS/MS evaluation of small molecules screen for successful PET tracers?

    PubMed

    Joshi, Elizabeth M; Need, Anne; Schaus, John; Chen, Zhaogen; Benesh, Dana; Mitch, Charles; Morton, Stuart; Raub, Thomas J; Phebus, Lee; Barth, Vanessa

    2014-12-17

    Positron emission tomography (PET) imaging has become a useful noninvasive technique to explore molecular biology within living systems; however, the utility of this method is limited by the availability of suitable radiotracers to probe specific targets and disease biology. Methods to identify potential areas of improvement in the ability to predict small molecule performance as tracers prior to radiolabeling would speed the discovery of novel tracers. In this retrospective analysis, we characterized the brain penetration or peak SUV (standardized uptake value), binding potential (BP), and brain exposure kinetics across a series of known, nonradiolabeled PET ligands using in vivo LC-MS/MS (liquid chromatography coupled to mass spectrometry) and correlated these parameters with the reported PET ligand performance in nonhuman primates and humans available in the literature. The PET tracers studied included those reported to label G protein-coupled receptors (GPCRs), intracellular enzymes, and transporters. Additionally, data for each tracer was obtained from a mouse brain uptake assay (MBUA), previously published, where blood-brain barrier (BBB) penetration and clearance parameters were assessed and compared against similar data collected on a broad compound set of central nervous system (CNS) therapeutic compounds. The BP and SUV identified via nonradiolabeled LC-MS/MS, while different from the published values observed in the literature PET tracer data, allowed for an identification of initial criteria values we sought to facilitate increased potential for success from our early discovery screening paradigm. Our analysis showed that successful, as well as novel, clinical PET tracers exhibited BP of greater than 1.5 and peak SUVs greater than approximately 150% at 5 min post dose in rodents. The brain kinetics appeared similar between both techniques despite differences in tracer dose, suggesting linearity across these dose ranges. The assessment of tracers in a

  9. Dynamic neurotransmitter interactions measured with PET

    SciTech Connect

    Schiffer, W.K.; Dewey, S.L.

    2001-04-02

    Positron emission tomography (PET) has become a valuable interdisciplinary tool for understanding physiological, biochemical and pharmacological functions at a molecular level in living humans, whether in a healthy or diseased state. The utility of tracing chemical activity through the body transcends the fields of cardiology, oncology, neurology and psychiatry. In this, PET techniques span radiochemistry and radiopharmaceutical development to instrumentation, image analysis, anatomy and modeling. PET has made substantial contributions in each of these fields by providing a,venue for mapping dynamic functions of healthy and unhealthy human anatomy. As diverse as the disciplines it bridges, PET has provided insight into an equally significant variety of psychiatric disorders. Using the unique quantitative ability of PET, researchers are now better able to non-invasively characterize normally occurring neurotransmitter interactions in the brain. With the knowledge that these interactions provide the fundamental basis for brain response, many investigators have recently focused their efforts on an examination of the communication between these chemicals in both healthy volunteers and individuals suffering from diseases classically defined as neurotransmitter specific in nature. In addition, PET can measure the biochemical dynamics of acute and sustained drug abuse. Thus, PET studies of neurotransmitter interactions enable investigators to describe a multitude of specific functional interactions in the human brain. This information can then be applied to understanding side effects that occur in response to acute and chronic drug therapy, and to designing new drugs that target multiple systems as opposed to single receptor types. Knowledge derived from PET studies can be applied to drug discovery, research and development (for review, see (Fowler et al., 1999) and (Burns et al., 1999)). Here, we will cover the most substantial contributions of PET to understanding

  10. The Design, Synthesis and Potential Utility of Fluorescence Probes that Target DFG-out Conformation of p38[alpha] for High Throughput Screening Binding Assay

    SciTech Connect

    Tecle, Haile; Feru, Frederic; Liu, Hu; Kuhn, Cyrille; Rennie, Glen; Morris, Mark; Shao, Jiangxing; Cheng, Alan C.; Gikunju, Diana; Miret, Juan; Coli, Rocco; Xi, Simon; Clugston, Susan L.; Low, Simon; Kazmirski, Steven; Ding, Yuan-Hua; Cao, Qing; Johnson, Theresa L.; Deshmukh, Gayatri D.; DiNitto, Jonathan P.; Wu, Joe C.; English, Jessie M.; Pfizer

    2010-10-18

    The design, synthesis and utility of fluorescence probes that bind to the DFG-out conformation of p38{alpha} kinase are described. Probes that demonstrate good affinity for p38{alpha}, have been identified and one of the probes, PF-04438255, has been successfully used in an high throughput screening (HTS) assay to identify two novel non-classical p38{alpha} inhibitors. In addition, a cascade activity assay was utilized to validate the selective binding of these non-classical kinase inhibitors to the unactive form of the enzyme.

  11. PET/CT AND RADIOIMMUNOTHERAPY OF PROSTATE CANCER

    PubMed Central

    Bouchelouche, Kirsten; Capala, Jacek; Oehr, Peter

    2009-01-01

    Purpose of review Traditional morphologically based imaging modalities are now being complemented by positron emission tomography (PET)/computerized tomography (CT) in prostate cancer. Metastatic prostate cancer is an attractive target for radioimmunotherapy (RIT) since no effective therapies are available. This review highlights the most important achievements within the last year in PET/CT and RIT of prostate cancer. Recent findings Conflicting results exist on the use of choline for detection of malignant disease in the prostate gland. The role of PET/CT in N-staging remains to be elucidated further. However, 18F-choline and 11C-choline PET/CT have been demonstrated to be useful for detection of recurrence. 18F-choline and 18F-fluoride PET/CT are useful for detection of bone metastases. Prostate tumor antigens may be used as targets for RIT. Prostate specific membrane antigen (PSMA) is currently under focus of a number of diagnostic and therapeutic strategies. J591, a monoclonal antibody, that targets the extracellular domain of PSMA, shows promising results. HER2 receptors may also have a potential as target for PET/CT imaging and RIT of advanced prostate cancer. Summary PET/CT in prostate cancer has proven to play a significant role, in particular for detection of prostate cancer recurrence and bone metastases. Radioimmunotherapy of metastatic prostate cancer warrant further investigations. PMID:19535981

  12. Application of Targeted Functional Assays to Assess a Putative Vascular Disruption Developmental Toxicity Pathway Informed By ToxCast High-Throughput Screening Data

    EPA Science Inventory

    Chemical perturbation of vascular development is a putative toxicity pathway which may result in developmental toxicity. EPA’s high-throughput screening (HTS) ToxCast program contains assays which measure cellular signals and biological processes critical for blood vessel develop...

  13. A Multiplexed Cell-Based Assay for the Identification of Modulators of Pre-Membrane Processing as a Target against Dengue Virus

    PubMed Central

    Stolp, Zachary D.; Smurthwaite, Cameron A.; Reed, Connor; Williams, Wesley; Dharmawan, Andre; Djaballah, Hakim

    2015-01-01

    The DenV pre-membrane protein (prM) is a crucial chaperone for the viral envelope protein, preventing premature fusion with vesicles during viral export. prM molecules in immature particles are cleaved by host proteases, leading to mature fusogenic virions. Blockade of prM cleavage would restrict fusion and represents a novel druggable opportunity against DenV. We have thus established a cell-based platform to monitor prM processing that relies on an engineered two-tag scaffold that travels to the cell surface through the secretory pathway. The assay discriminates between a single cell-surface tag when prM is cleaved and two tags when it is not, as detected through fluorescent-coupled antibodies by flow cytometry. The assay, miniaturized into a 96-well plate format, was multiplexed with the HIV-1 envelope boundary, also cleaved in the same pathway. A pilot screen against 1280 compounds was executed, leading to the identification of a potential active and corroborating the robustness of our assay for large-scale screening. We describe for the first time a cell-based assay that monitors DenV prM processing within the classical secretory pathway, which was exploited to identify a potential novel drug against DenV. PMID:25724189

  14. A Multiplexed Cell-Based Assay for the Identification of Modulators of Pre-Membrane Processing as a Target against Dengue Virus.

    PubMed

    Stolp, Zachary D; Smurthwaite, Cameron A; Reed, Connor; Williams, Wesley; Dharmawan, Andre; Djaballah, Hakim; Wolkowicz, Roland

    2015-06-01

    The DenV pre-membrane protein (prM) is a crucial chaperone for the viral envelope protein, preventing premature fusion with vesicles during viral export. prM molecules in immature particles are cleaved by host proteases, leading to mature fusogenic virions. Blockade of prM cleavage would restrict fusion and represents a novel druggable opportunity against DenV. We have thus established a cell-based platform to monitor prM processing that relies on an engineered two-tag scaffold that travels to the cell surface through the secretory pathway. The assay discriminates between a single cell-surface tag when prM is cleaved and two tags when it is not, as detected through fluorescent-coupled antibodies by flow cytometry. The assay, miniaturized into a 96-well plate format, was multiplexed with the HIV-1 envelope boundary, also cleaved in the same pathway. A pilot screen against 1280 compounds was executed, leading to the identification of a potential active and corroborating the robustness of our assay for large-scale screening. We describe for the first time a cell-based assay that monitors DenV prM processing within the classical secretory pathway, which was exploited to identify a potential novel drug against DenV. PMID:25724189

  15. Micro-PET/CT Monitoring of Herpes Thymidine Kinase Suicide Gene Therapy in a Prostate Cancer Xenograft: The Advantage of a Cell-specific Transcriptional Targeting Approach

    PubMed Central

    Johnson, Mai; Sato, Makoto; Burton, Jeremy; Gambhir, Sanjiv S.; Carey, Michael; Wu, Lily

    2010-01-01

    Cancer gene therapy based on tissue-restricted expression of cytotoxic gene should achieve superior therapeutic index over an unrestricted method. This study compared the therapeutic effects of a highly augmented, prostate-specific gene expression method to a strong constitutive promoter-driven approach. Molecular imaging was coupled to gene therapy to ascertain real-time therapeutic activity. The imaging reporter gene (luciferase) and the cytotoxic gene (herpes simplex thymidine kinase) were delivered by adenoviral vectors injected directly into human prostate tumors grafted in SCID mice. Serial bioluminescence imaging, positron emission tomography, and computed tomography revealed restriction of gene expression to the tumors when prostate-specific vector was employed. In contrast, administration of constitutive active vector resulted in strong signals in the liver. Liver serology, tissue histology, and frail condition of animals confirmed liver toxicity suffered by the constitutive active cohorts, whereas the prostate-targeted group was unaffected. The extent of tumor killing was analyzed by apoptotic staining and human prostate marker (prostate-specific antigen). Overall, the augmented prostate-specific expression system was superior to the constitutive approach in safeguarding against systemic toxicity, while achieving effective tumor killing. Integrating noninvasive imaging into cytotoxic gene therapy will provide a useful strategy to monitor gene expression and therapeutic efficacy in future clinical protocols. PMID:16285908

  16. Improving Instruction through PET.

    ERIC Educational Resources Information Center

    Evans, Pamela Roland

    1982-01-01

    Outlines the content and training methods used in the Program for Effective Teaching (PET), the successful staff development program of Newport News (Virginia). PET promotes application of five instructional skills: selecting learning objectives, teaching to the objectives, establishing learner focus, monitoring learner progress, and enhancing…

  17. My Pet Rock

    ERIC Educational Resources Information Center

    Lark, Adam; Kramp, Robyne; Nurnberger-Haag, Julie

    2008-01-01

    Many teachers and students have experienced the classic pet rock experiment in conjunction with a geology unit. A teacher has students bring in a "pet" rock found outside of school, and the students run geologic tests on the rock. The tests include determining relative hardness using Mohs scale, checking for magnetization, and assessing luster.…

  18. Healthy pets, healthy people.

    PubMed

    Wong, S K; Feinstein, L H; Heidmann, P

    1999-08-01

    Zoonoses, diseases that can be transmitted from animals to humans, can pose serious health risks to immunocompromised people. Although pets can carry zoonoses, owning and caring for animals can benefit human health. Information exists about preventing transmission of zoonoses, but not all physicians and veterinarians provide adequate and accurate information to immunocompromised pet owners. This disease prevention/health promotion project provides physicians and veterinarians with information, created specifically to share with patients and clients, about the health risks and benefits of pet ownership. Further, "Healthy Pets, Healthy People" encourages communication between veterinarians, physicians, clients, and patients and can serve as a model program for a nation-wide effort to aid health professionals in making recommendations about pet ownership for immunocompromised people. PMID:10434969

  19. TAL Effector Specificity for base 0 of the DNA Target Is Altered in a Complex, Effector- and Assay-Dependent Manner by Substitutions for the Tryptophan in Cryptic Repeat –1

    PubMed Central

    Doyle, Erin L.; Hummel, Aaron W.; Demorest, Zachary L.; Starker, Colby G.; Voytas, Daniel F.; Bradley, Philip; Bogdanove, Adam J.

    2013-01-01

    TAL effectors are re-targetable transcription factors used for tailored gene regulation and, as TAL effector-nuclease fusions (TALENs), for genome engineering. Their hallmark feature is a customizable central string of polymorphic amino acid repeats that interact one-to-one with individual DNA bases to specify the target. Sequences targeted by TAL effector repeats in nature are nearly all directly preceded by a thymine (T) that is required for maximal activity, and target sites for custom TAL effector constructs have typically been selected with this constraint. Multiple crystal structures suggest that this requirement for T at base 0 is encoded by a tryptophan residue (W232) in a cryptic repeat N-terminal to the central repeats that exhibits energetically favorable van der Waals contacts with the T. We generated variants based on TAL effector PthXo1 with all single amino acid substitutions for W232. In a transcriptional activation assay, many substitutions altered or relaxed the specificity for T and a few were as active as wild type. Some showed higher activity. However, when replicated in a different TAL effector, the effects of the substitutions differed. Further, the effects differed when tested in the context of a TALEN in a DNA cleavage assay, and in a TAL effector-DNA binding assay. Substitution of the N-terminal region of the PthXo1 construct with that of one of the TAL effector-like proteins of Ralstonia solanacearum, which have arginine in place of the tryptophan, resulted in specificity for guanine as the 5’ base but low activity, and several substitutions for the arginine, including tryptophan, destroyed activity altogether. Thus, the effects on specificity and activity generated by substitutions at the W232 (or equivalent) position are complex and context dependent. Generating TAL effector scaffolds with high activity that robustly accommodate sites without a T at position 0 may require larger scale re-engineering. PMID:24312634

  20. Using a Non-Image-Based Medium-Throughput Assay for Screening Compounds Targeting N-myristoylation in Intracellular Leishmania Amastigotes

    PubMed Central

    Paape, Daniel; Bell, Andrew S.; Heal, William P.; Hutton, Jennie A.; Leatherbarrow, Robin J.; Tate, Edward W.; Smith, Deborah F.

    2014-01-01

    We have refined a medium-throughput assay to screen hit compounds for activity against N-myristoylation in intracellular amastigotes of Leishmania donovani. Using clinically-relevant stages of wild type parasites and an Alamar blue-based detection method, parasite survival following drug treatment of infected macrophages is monitored after macrophage lysis and transformation of freed amastigotes into replicative extracellular promastigotes. The latter transformation step is essential to amplify the signal for determination of parasite burden, a factor dependent on equivalent proliferation rate between samples. Validation of the assay has been achieved using the anti-leishmanial gold standard drugs, amphotericin B and miltefosine, with EC50 values correlating well with published values. This assay has been used, in parallel with enzyme activity data and direct assay on isolated extracellular amastigotes, to test lead-like and hit-like inhibitors of Leishmania N-myristoyl transferase (NMT). These were derived both from validated in vivo inhibitors of Trypanosoma brucei NMT and a recent high-throughput screen against L. donovani NMT. Despite being a potent inhibitor of L. donovani NMT, the activity of the lead T. brucei NMT inhibitor (DDD85646) against L. donovani amastigotes is relatively poor. Encouragingly, analogues of DDD85646 show improved translation of enzyme to cellular activity. In testing the high-throughput L. donovani hits, we observed macrophage cytotoxicity with compounds from two of the four NMT-selective series identified, while all four series displayed low enzyme to cellular translation, also seen here with the T. brucei NMT inhibitors. Improvements in potency and physicochemical properties will be required to deliver attractive lead-like Leishmania NMT inhibitors. PMID:25522361

  1. Rapid Genotyping of Cytomegalovirus in Dried Blood Spots by Multiplex Real-Time PCR Assays Targeting the Envelope Glycoprotein gB and gH Genes

    PubMed Central

    Wessels, Els; Korver, Anna M. H.; van der Eijk, Annemiek A.; Rusman, Lisette G.; Kroes, Aloys C. M.; Vossen, Ann C. T. M.

    2012-01-01

    Genotyping of cytomegalovirus (CMV) is useful to examine potential differences in the pathogenicity of strains and to demonstrate coinfection with multiple strains involved in CMV disease in adults and congenitally infected newborns. Studies on genotyping of CMV in dried blood spots (DBS) are rare and have been hampered by the small amount of dried blood available. In this study, two multiplex real-time PCR assays for rapid gB and gH genotyping of CMV in DBS were developed. Validation of the assays with 39 CMV-positive plasma samples of transplant recipients and 21 urine specimens of congenitally infected newborns was successful in genotyping 100% of the samples, with gB1 and gB3 being the most prevalent genotypes. Multiple gB and gH genotypes were detected in 36% and 33% of the plasma samples, respectively. One urine sample from a newborn with symptomatic congenital CMV was positive for gB1 and gB2. DBS of congenitally infected newborns (n = 41) were tested using 9 μl of dried blood, and genotypes were detected in 81% (gB) and 73% (gH) of the samples, with gB3 being the most prevalent genotype. No clear association of specific genotypes with clinical outcome was observed. In conclusion, the CMV gB and gH PCR assays were found to be rapid, sensitive for detecting mixed infections, and suitable for direct usage on DBS. These assays are efficient tools for genotyping of CMV in DBS of congenitally infected newborns. PMID:22116158

  2. A versatile scalable PET processing system

    SciTech Connect

    H. Dong, A. Weisenberger, J. McKisson, Xi Wenze, C. Cuevas, J. Wilson, L. Zukerman

    2011-06-01

    Positron Emission Tomography (PET) historically has major clinical and preclinical applications in cancerous oncology, neurology, and cardiovascular diseases. Recently, in a new direction, an application specific PET system is being developed at Thomas Jefferson National Accelerator Facility (Jefferson Lab) in collaboration with Duke University, University of Maryland at Baltimore (UMAB), and West Virginia University (WVU) targeted for plant eco-physiology research. The new plant imaging PET system is versatile and scalable such that it could adapt to several plant imaging needs - imaging many important plant organs including leaves, roots, and stems. The mechanical arrangement of the detectors is designed to accommodate the unpredictable and random distribution in space of the plant organs without requiring the plant be disturbed. Prototyping such a system requires a new data acquisition system (DAQ) and data processing system which are adaptable to the requirements of these unique and versatile detectors.

  3. Cardiac applications of PET.

    PubMed

    Sarikaya, Ismet

    2015-10-01

    Routine use of cardiac positron emission tomography (PET) applications has been increasing but has not replaced cardiac single-photon emission computerized tomography (SPECT) studies yet. The majority of cardiac PET tracers, with the exception of fluorine-18 fluorodeoxyglucose (18F-FDG), are not widely available, as they require either an onsite cyclotron or a costly generator for their production. 18F-FDG PET imaging has high sensitivity for the detection of hibernating/viable myocardium and has replaced Tl-201 SPECT imaging in centers equipped with a PET/CT camera. PET myocardial perfusion imaging with various tracers such as Rb-82, N-13 ammonia, and O-15 H2O has higher sensitivity and specificity than myocardial perfusion SPECT for the detection of coronary artery disease (CAD). In particular, quantitative PET measurements of myocardial perfusion help identify subclinical coronary stenosis, better define the extent and severity of CAD, and detect ischemia when there is balanced reduction in myocardial perfusion due to three-vessel or main stem CAD. Fusion images of PET perfusion and CT coronary artery calcium scoring or CT coronary angiography provide additional complementary information and improve the detection of CAD. PET studies with novel 18F-labeled perfusion tracers such as 18F-flurpiridaz and 18F-FBnTP have yielded high sensitivity and specificity in the diagnosis of CAD. These tracers are still being tested in humans, and, if approved for clinical use, they will be commercially and widely available. In addition to viability studies, 18F-FDG PET can also be utilized to detect inflammation/infection in various conditions such as endocarditis, sarcoidosis, and atherosclerosis. Some recent series have obtained encouraging results for the detection of endocarditis in patients with intracardiac devices and prosthetic valves. PET tracers for cardiac neuronal imaging, such as C-11 HED, help assess the severity of heart failure and post-transplant cardiac

  4. PET Imaging in Huntington’s Disease

    PubMed Central

    Roussakis, Andreas-Antonios; Piccini, Paola

    2015-01-01

    To date, little is known about how neurodegeneration and neuroinflammation propagate in Huntington’s disease (HD). Unfortunately, no treatment is available to cure or reverse the progressive decline of function caused by the disease, thus considering HD a fatal disease. Mutation gene carriers typically remain asymptomatic for many years although alterations in the basal ganglia and cortex occur early on in mutant HD gene–carriers. Positron Emission Tomography (PET) is a functional imaging technique of nuclear medicine which enables in vivo visualization of numerous biological molecules expressed in several human tissues. Brain PET is most powerful to study in vivo neuronal and glial cells function as well as cerebral blood flow in a plethora of neurodegenerative disorders including Parkinson’s disease, Alzheimer’s and HD. In absence of HD–specific biomarkers for monitoring disease progression, previous PET studies in HD were merely focused on the study of dopaminergic terminals, cerebral blood flow and glucose metabolism in manifest and premanifest HD–gene carriers. More recently, research interest has been exploring novel PET targets in HD including the state of phosphodiesterse expression and the role of activated microglia. Hence, a better understanding of the HD pathogenesis mechanisms may lead to the development of targeted therapies. PET imaging follow–up studies with novel selective PET radiotracers such as 11C-IMA–107 and 11C-PBR28 may provide insight on disease progression and identify prognostic biomarkers, elucidate the underlying HD pathology and assess novel pharmaceutical agents and over time. PMID:26683130

  5. The ADNI PET Core

    PubMed Central

    Jagust, William J.; Bandy, Dan; Chen, Kewei; Foster, Norman L.; Landau, Susan M.; Mathis, Chester A.; Price, Julie C.; Reiman, Eric M.; Skovronsky, Daniel; Koeppe, Robert A.

    2010-01-01

    Background This is a progress report of the Alzheimer's Disease Neuroimaging Initiative (ADNI) PET Core. Methods The Core has supervised the acquisition, quality control, and analysis of longitudinal [18F]fluorodeoxyglucose PET (FDG-PET) data in approximately half of the ADNI cohort. In an “add on” study, approximately 100 subjects also underwent scanning with [11C]PIB-PET for amyloid imaging. The Core developed quality control procedures and standardized image acquisition by developing an imaging protocol that has been widely adopted in academic and pharmaceutical industry studies. Data processing provides users with scans that have identical orientation and resolution characteristics despite acquisition on multiple scanner models. The Core labs have used a number of different approaches to characterize differences between subject groups (AD, MCI, controls), to examine longitudinal change over time in glucose metabolism and amyloid deposition, and to assess the use of FDG-PET as a potential outcome measure in clinical trials. Results ADNI data indicate that FDG-PET increases statistical power over traditional cognitive measures, might aid subject selection, and could substantially reduce the sample size in a clinical trial. PIB-PET data showed expected group differences, and identified subjects with significant annual increases in amyloid load across the subject groups. The next activities of the PET core in ADNI will entail developing standardized protocols for amyloid imaging using the [18F]-labeled amyloid imaging agent AV45, which can be delivered to virtually all ADNI sites. Conclusions ADNI has demonstrated the feasibility and utility of multicenter PET studies and is helping to clarify the role of biomarkers in the study of aging and dementia. PMID:20451870

  6. Comparison of liquid chromatography and supercritical fluid chromatography coupled to compact single quadrupole mass spectrometer for targeted in vitro metabolism assay.

    PubMed

    Spaggiari, Dany; Mehl, Florence; Desfontaine, Vincent; Grand-Guillaume Perrenoud, Alexandre; Fekete, Szabolcs; Rudaz, Serge; Guillarme, Davy

    2014-12-01

    The goal of this study was to evaluate the combination of powerful chromatographic methods and compact single quadrupole MS device for simple in vitro cytochrome P450 (CYP) inhibition assay, instead of more expensive triple quadrupole MS/MS detectors. For this purpose, two modern chromatographic approaches (ultra-high pressure liquid chromatography (UHPLC) and ultra-high performance supercritical fluid chromatography (UHPSFC)) were tested in combination with simple MS detector. To show the applicability for an in vitro CYP-mediated metabolism assay using the cocktail approach, a method was first developed in UHPLC-MS to separate a mixture of 8 probe substrates and 8 CYP-specific metabolites. A screening procedure was initially applied to determine the best combination of a column, an organic modifier and a mobile-phase pH, followed by fine tuning of the conditions (i.e., gradient profile, temperature and pH) using HPLC modelling software. A similar sequential method development procedure was also evaluated for UHPSFC-MS. For method development, where peak tracking is necessary, the use of single quadrupole MS was found to be extremely valuable for following the investigated analytes. Ultimately, a baseline separation of the 16 compounds was achieved in both UHPLC-MS and UHPSFC-MS with an analysis time of less than 7 min. In a second series of experiments, sensitivity was evaluated, and LOQ values were between 2 and 100 ng/mL in UHPLC-MS, while they ranged from 2 to 200 ng/mL in UHPSFC-MS. Based on the concentrations employed for the current in vitro phase I metabolism assay, these LOQ values were appropriate for this type of application. Finally, the two analytical methods were applied to in vitro CYP-dependent metabolism testing. Two well-known phytochemical inhibitors, yohimbine and resveratrol, were investigated, and reliable conclusions were drawn from these experiments with both UHPLC-MS and UHPSFC-MS. At the end, the proposed strategy of optimized

  7. Specific detection of Xanthomonas oryzae pv. oryzicola in infected rice plant by use of PCR assay targeting a membrane fusion protein gene.

    PubMed

    Kang, Man Jung; Shim, Jae Kyung; Cho, Min Seok; Seol, Young Joo; Hahn, Jang Ho; Hwang, Duk Ju; Park, Dong Suk

    2008-09-01

    Successful control of Xanthomonas oryzae pv. oryzicola, the causal agent of bacterial leaf streak, requires a specific and reliable diagnostic tool. A pathovar-specific PCR assay was developed for the rapid and accurate detection of the plant pathogenic bacterium Xanthomonas oryzae pv. oryzicola in diseased plant. Based on differences in a membrane fusion protein gene of Xanthomonas oryzae pv. oryzicola and other microorganisms, which was generated from NCBI (http://www.ncbi.nlm.nih.gov/) and CMR (http://cmr.tigr.org/) BLAST searches, one pair of pathovar-specific primers, XOCMF/XOCMR, was synthesized. Primers XOCMF and XOCMR from a membrane fusion protein gene were used to amplify a 488-bp DNA fragment. The PCR product was only produced from 4 isolates of Xanthomonas oryzae pv. oryzicola among 37 isolates of other pathovars and species of Xanthomonas, Pectobacterium, Pseudomonas, Burkholderia, Escherichia coli, and Fusarium oxysporum f.sp. dianthi. The results suggested that the assay detected the pathogen more rapidly and accurately than standard isolation methods. PMID:18852502

  8. ThermoPhyl: a software tool for selecting phylogenetically optimized conventional and quantitative-PCRtaxon-targeted assays for usewith complex samples

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The ability to specifically and sensitively target genotypes of interest is critical for the success of many PCR-based analyses of environmental or clinical samples that contain multiple templates. Next-generation sequence data clearly show that such samples can harbour hundreds to thousands of oper...

  9. Heart PET scan

    MedlinePlus

    Heart nuclear medicine scan; Heart positron emission tomography; Myocardial PET scan ... Udelson JE, Dilsizian V, Bonow RO. Nuclear cardiology. In: Mann DL, ... A Textbook of Cardiovascular Medicine . 10th ed. Philadelphia, ...

  10. Healthy Pets and People

    MedlinePlus

    ... Pregnant women should avoid adopting or handling stray cats, especially kittens. They particularly should not clean litter ... may be sick. Many pets, such as dogs, cats, reptiles, rodents, and birds, carry germs that can ...

  11. Pets and Parasites

    MedlinePlus

    ... make me sick? Household pets such as dogs, cats, birds and reptiles can carry diseases or parasites ... might be used as litter boxes by neighborhood cats. Keep your children out of the dirt in ...

  12. Household Hazards to Pets

    MedlinePlus

    ... health by becoming aware of the most common health hazards found in many pet-owning households. Hazards in the Kitchen Foods Many foods are perfectly safe for humans, but could be harmful or potentially deadly to ...

  13. Brain PET scan

    MedlinePlus

    ... tests, such as magnetic resonance imaging ( MRI ) and computed tomography ( CT ) scans only reveal the structure of the ... a PET/CT. Alternative Names ... PT, Rijntjes M, Weiller C. Neuroimaging: Functional neuroimaging. In: Daroff RB, Fenichel GM, Jankovic ...

  14. Use of DNA melting simulation software for in silico diagnostic assay design: targeting regions with complex melting curves and confirmation by real-time PCR using intercalating dyes

    PubMed Central

    Rasmussen, John P; Saint, Christopher P; Monis, Paul T

    2007-01-01

    Background DNA melting curve analysis using double-stranded DNA-specific dyes such as SYTO9 produce complex and reproducible melting profiles, resulting in the detection of multiple melting peaks from a single amplicon and allowing the discrimination of different species. We compare the melting curves of several Naegleria and Cryptosporidium amplicons generated in vitro with in silico DNA melting simulations using the programs POLAND and MELTSIM., then test the utility of these programs for assay design using a genetic marker for toxin production in cyanobacteria. Results The SYTO9 melting curve profiles of three species of Naegleria and two species of Cryptosporidium were similar to POLAND and MELTSIM melting simulations, excepting some differences in the relative peak heights and the absolute melting temperatures of these peaks. MELTSIM and POLAND were used to screen sequences from a putative toxin gene in two different species of cyanobacteria and identify regions exhibiting diagnostic melting profiles. For one of these diagnostic regions the POLAND and MELTSIM melting simulations were observed to be different, with POLAND more accurately predicting the melting curve generated in vitro. Upon further investigation of this region with MELTSIM, inconsistencies between the melting simulation for forward and reverse complement sequences were observed. The assay was used to accurately type twenty seven cyanobacterial DNA extracts in vitro. Conclusion Whilst neither POLAND nor MELTSIM simulation programs were capable of exactly predicting DNA dissociation in the presence of an intercalating dye, the programs were successfully used as tools to identify regions where melting curve differences could be exploited for diagnostic melting curve assay design. Refinements in the simulation parameters would be required to account for the effect of the intercalating dye and salt concentrations used in real-time PCR. The agreement between the melting curve simulations for

  15. PET studies in epilepsy

    PubMed Central

    Sarikaya, Ismet

    2015-01-01

    Various PET studies, such as measurements of glucose, serotonin and oxygen metabolism, cerebral blood flow and receptor bindings are availabe for epilepsy. 18Fluoro-2-deoxyglucose (18F-FDG) PET imaging of brain glucose metabolism is a well established and widely available technique. Studies have demonstrated that the sensitivity of interictal FDG-PET is higher than interictal SPECT and similar to ictal SPECT for the lateralization and localization of epileptogenic foci in presurgical patients refractory to medical treatments who have noncontributory EEG and MRI. In addition to localizing epileptogenic focus, FDG-PET provide additional important information on the functional status of the rest of the brain. The main limitation of interictal FDG-PET is that it cannot precisely define the surgical margin as the area of hypometabolism usually extends beyond the epileptogenic zone. Various neurotransmitters (GABA, glutamate, opiates, serotonin, dopamine, acethylcholine, and adenosine) and receptor subtypes are involved in epilepsy. PET receptor imaging studies performed in limited centers help to understand the role of neurotransmitters in epileptogenesis, identify epileptic foci and investigate new treatment approaches. PET receptor imaging studies have demonstrated reduced 11C-flumazenil (GABAA-cBDZ) and 18F-MPPF (5-HT1A serotonin) and increased 11C-cerfentanil (mu opiate) and 11C-MeNTI (delta opiate) bindings in the area of seizure. 11C-flumazenil has been reported to be more sensitive than FDG-PET for identifying epileptic foci. The area of abnormality on GABAAcBDZ and opiate receptor images is usually smaller and more circumscribed than the area of hypometabolism on FDG images. Studies have demonstrated that 11C-alpha-methyl-L-tryptophan PET (to study synthesis of serotonin) can detect the epileptic focus within malformations of cortical development and helps in differentiating epileptogenic from non-epileptogenic tubers in patients with tuberous sclerosis complex

  16. PET/CT Artifacts

    PubMed Central

    Blodgett, Todd M.; Mehta, Ajeet S.; Mehta, Amar S.; Laymon, Charles M.; Carney, Jonathan; Townsend, David W.

    2014-01-01

    There are several artifacts encountered in PET/CT imaging, including attenuation correction (AC) artifacts associated with using CT for attenuation correction. Several artifacts can mimic a 2-deoxy-2-[18F] fluoro-D-glucose (FDG) avid malignant lesions and therefore recognition of these artifacts is clinically relevant. Our goal was to identify and characterize these artifacts and also discuss some protocol variables that may affect image quality in PET/CT. PMID:21237418

  17. A facile, sensitive, and highly specific trinitrophenol assay based on target-induced synergetic effects of acid induction and electron transfer towards DNA-templated copper nanoclusters.

    PubMed

    Li, Haiyin; Chang, Jiafu; Hou, Ting; Ge, Lei; Li, Feng

    2016-11-01

    Reliable, selective and sensitive approaches for trinitrophenol (TNP) detection are highly desirable with respect to national security and environmental protection. Herein, a simple and novel fluorescent strategy for highly sensitive and specific TNP assay has been successfully developed, which is based on the quenching of the fluorescent poly(thymine)-templated copper nanoclusters (DNA-CuNCs), through the synergetic effects of acid induction and electron transfer. Upon the addition of TNP, donor-acceptor complexes between the electron-deficient nitro-groups in TNP and the electron-donating DNA templates are formed, resulting in the close proximity between TNP and CuNCs. Moreover, the acidity of TNP contributes to the pH decrease of the system. These factors combine to dramatically quench the fluorescence of DNA-CuNCs, providing a "signal-off" strategy for TNP sensing. The as-proposed strategy demonstrates high sensitivity for TNP assay, and a detection limit of 0.03μM is obtained, which is lower than those reported by using organic fluorescent materials. More significantly, this approach shows outstanding selectivity over a number of TNP analogues, such as 2,4,6-trinitrotoluene (TNT), 2,4-dinitrotoluene (DNT), 2,4-dinitrophenol (DNP), 3-nitrophenol (NP), nitrobenzene (NB), phenol (BP), and toluene (BT). Compared with previous studies, this method does not need complex DNA sequence design, fluorescent dye labeling, or sophisticated organic reactions, rendering the strategy with additional advantages of simplicity and cost-effectiveness. In addition, the as-proposed strategy has been adopted for the detection of TNP in natural water samples, indicating its great potential to be applied in the fields of public safety and environmental monitoring. PMID:27591641

  18. Assessment of α-Fetoprotein Targeted HSV1-tk Expression in Hepatocellular Carcinoma with In Vivo Imaging

    PubMed Central

    Park, Ju Hui; Kim, Kwang Il; Lee, Kyo Chul; Lee, Yong Jin; Lee, Tae Sup; Chung, Wee Sup; Lim, Sang Moo

    2015-01-01

    Abstract Tumor-specific enhancer/promoter is applicable for targeting gene expression in tumors and helpful for tumor-targeting imaging and therapy. We aimed to acquire α-fetoprotein (AFP)-producing hepatocellular carcinoma (HCC) specific images using adenovirus containing HSV1-tk gene controlled by AFP enhancer/promoter and evaluate in vivo ganciclovir (GCV)-medicated therapeutic effects on AFP-targeted HSV1-tk expression with 18F-FDG positron emission tomography (PET). Recombinant adenovirus expressing HSV1-tk under AFP enhancer/promoter was produced (AdAFP-TK) and the expression levels were evaluated by RT-PCR and 125I-IVDU uptake. GCV-mediated HSV1-tk cytotoxicity was determined by MTT assay. After the mixture of AdAFP-fLuc and AdAFP-TK was administrated, bioluminescent images (BLIs) and 18F-FHBG PET images were obtained in tumor-bearing mice. In vivo therapeutic effects of AdAFP-TK and GCV in the HuH-7 xenograft model were monitored by 18F-FDG PET. When infected with AdAFP-TK, cell viability in HuH-7 was reduced, but those in HT-29 and SK-Hep-1 were not significantly decreased at any GCV concentration less than 100 μM. AFP-targeted fLuc and HSV1-tk expression were clearly visualized by BLI and 18F-FHBG PET images in AFP-producing HCC, respectively. In vivo GCV-mediated tumor growth inhibition by AFP-targeted HSV1-tk expression was monitored by 18F-FDG PET. Recombinant AdAFP-TK could be applied for AFP-targeted HCC gene therapy and imaging in AFP-producing HCC. PMID:25545853

  19. Pet-related infections.

    PubMed

    Rabinowitz, Peter M; Gordon, Zimra; Odofin, Lynda

    2007-11-01

    Human contact with cats, dogs, and other pets results in several million infections each year in the United States, ranging from self-limited skin conditions to life-threatening systemic illnesses. Toxoplasmosis is one of the most common pet-related parasitic infections. Although toxoplasmosis is usually asymptomatic or mild, it may cause serious congenital infection if a woman is exposed during pregnancy, particularly in the first trimester. Common pet-borne fungal infections include tinea corporis/capitis (ringworm); campylobacteriosis and salmonellosis are among the most common bacterial infections associated with pet ownership. Less commonly, pets can transmit arthropod-borne and viral illnesses (e.g., scabies, rabies). Infection in a pet can provide sentinel warning of local vectors and endemic conditions, such as Lyme disease risk. Treatment is infection-specific, although many infections are self-limited. Prevention involves common sense measures such as adequate hand washing, proper disposal of animal waste, and ensuring that infected animals are diagnosed and treated. Special precautions are indicated for immunocompromised persons. Increased communication between primary care physicians and veterinarians could improve treatment and prevention of these conditions. PMID:18019874

  20. A Pragmatic Approach to HIV-1 Drug Resistance Determination in Resource-Limited Settings by Use of a Novel Genotyping Assay Targeting the Reverse Transcriptase-Encoding Region Only

    PubMed Central

    Bronze, Michelle; Wallis, Carole L.; Stuyver, Lieven; Steegen, Kim; Balinda, Sheila; Kityo, Cissy; Stevens, Wendy; Rinke de Wit, Tobias F.; Schuurman, Rob

    2013-01-01

    In resource-limited settings (RLS), reverse transcriptase (RT) inhibitors form the backbone of first-line treatment regimens. We have developed a simplified HIV-1 drug resistance genotyping assay targeting the region of RT harboring all major RT inhibitor resistance mutation positions, thus providing all relevant susceptibility data for first-line failures, coupled with minimal cost and labor. The assay comprises a one-step RT-PCR amplification reaction, followed by sequencing using one forward and one reverse primer, generating double-stranded coverage of RT amino acids (aa) 41 to 238. The assay was optimized for all major HIV-1 group M subtypes in plasma and dried blood spot (DBS) samples using a panel of reference viruses for HIV-1 subtypes A to D, F to H, and circulating recombinant form 01_AE (CRF01_AE) and applied to 212 clinical plasma samples and 25 DBS samples from HIV-1-infected individuals from Africa and Europe. The assay was subsequently transferred to Uganda and applied locally on clinical plasma samples. All major HIV-1 subtypes could be detected with an analytical sensitivity of 5.00E+3 RNA copies/ml for plasma and DBS. Application of the assay on 212 clinical samples from African subjects comprising subtypes A to D, F to H (rare), CRF01_AE, and CRF02_AG at a viral load (VL) range of 6.71E+2 to 1.00E+7 (median, 1.48E+5) RNA copies/ml was 94.8% (n = 201) successful. Application on clinical samples in Uganda demonstrated a comparable success rate. Genotyping of clinical DBS samples, all subtype C with a VL range of 1.02E+3 to 4.49E+5 (median, 1.42E+4) RNA copies/ml, was 84.0% successful. The described assay greatly reduces hands-on time and the costs required for genotyping and is ideal for use in RLS, as demonstrated in a reference laboratory in Uganda and its successful application on DBS samples. PMID:23536405

  1. A pragmatic approach to HIV-1 drug resistance determination in resource-limited settings by use of a novel genotyping assay targeting the reverse transcriptase-encoding region only.

    PubMed

    Aitken, Susan C; Bronze, Michelle; Wallis, Carole L; Stuyver, Lieven; Steegen, Kim; Balinda, Sheila; Kityo, Cissy; Stevens, Wendy; Rinke de Wit, Tobias F; Schuurman, Rob

    2013-06-01

    In resource-limited settings (RLS), reverse transcriptase (RT) inhibitors form the backbone of first-line treatment regimens. We have developed a simplified HIV-1 drug resistance genotyping assay targeting the region of RT harboring all major RT inhibitor resistance mutation positions, thus providing all relevant susceptibility data for first-line failures, coupled with minimal cost and labor. The assay comprises a one-step RT-PCR amplification reaction, followed by sequencing using one forward and one reverse primer, generating double-stranded coverage of RT amino acids (aa) 41 to 238. The assay was optimized for all major HIV-1 group M subtypes in plasma and dried blood spot (DBS) samples using a panel of reference viruses for HIV-1 subtypes A to D, F to H, and circulating recombinant form 01_AE (CRF01_AE) and applied to 212 clinical plasma samples and 25 DBS samples from HIV-1-infected individuals from Africa and Europe. The assay was subsequently transferred to Uganda and applied locally on clinical plasma samples. All major HIV-1 subtypes could be detected with an analytical sensitivity of 5.00E+3 RNA copies/ml for plasma and DBS. Application of the assay on 212 clinical samples from African subjects comprising subtypes A to D, F to H (rare), CRF01_AE, and CRF02_AG at a viral load (VL) range of 6.71E+2 to 1.00E+7 (median, 1.48E+5) RNA copies/ml was 94.8% (n = 201) successful. Application on clinical samples in Uganda demonstrated a comparable success rate. Genotyping of clinical DBS samples, all subtype C with a VL range of 1.02E+3 to 4.49E+5 (median, 1.42E+4) RNA copies/ml, was 84.0% successful. The described assay greatly reduces hands-on time and the costs required for genotyping and is ideal for use in RLS, as demonstrated in a reference laboratory in Uganda and its successful application on DBS samples. PMID:23536405

  2. PET imaging of in vivo caspase-3/7 activity following myocardial ischemia-reperfusion injury with the radiolabeled isatin sulfonamide analogue [18F]WC-4-116

    PubMed Central

    Thukkani, Arun K; Shoghi, Kooresh I; Zhou, Dong; Xu, Jinbin; Chu, Wenhua; Novak, Eric; Chen, Delphine L; Gropler, Robert J; Mach, Robert H

    2016-01-01

    The utility of [18F]WC-4-116, a PET tracer for imaging caspase-3 activation, was evaluated in an animal model of myocardial apoptosis. [18F]WC-4-116 was injected into rats at 3 hours after a 30 min period of ischemia induced by temporary occlusion of the left anterior descending coronary artery in Sprague-Dawley rats. [18F]WC-4-116 uptake was quantified by 1) autoradiography, 2) microPET imaging studies, and 3) post-PET biodistribution studies. MicroPET imaging also assessed uptake of the non-caspase-3-targeted tracer [18F]ICMT-18 at 3 hours postischemia. Enzyme assays and Western blotting assessed caspase-3 activation in both at-risk and not-at-risk regions. Caspase-3 enzyme activity increased in the at-risk but not in the not-at-risk myocardium. Quantitative autoradiographic analysis of [18F]WC-4-116 demonstrated nearly 2-fold higher uptake in the ischemia-reperfusion (IR) versus sham animals. [18F]WC-4-116 microPET imaging studies demonstrated that the IR animals was similarly elevated in relation to sham. [18F]ICMT-18 uptake did not increase in at-risk myocardium despite evidence of caspase-3 activation. Biodistribution studies with [18F]WC-4-116 confirmed the microPET findings. These data indicate that the caspase-3-PET tracer [18F]WC-4-116 can noninvasively image in vivo caspase activity during myocardial apoptosis and may be useful for clinical imaging in humans. PMID:27186438

  3. PET imaging of in vivo caspase-3/7 activity following myocardial ischemia-reperfusion injury with the radiolabeled isatin sulfonamide analogue [(18)F]WC-4-116.

    PubMed

    Thukkani, Arun K; Shoghi, Kooresh I; Zhou, Dong; Xu, Jinbin; Chu, Wenhua; Novak, Eric; Chen, Delphine L; Gropler, Robert J; Mach, Robert H

    2016-01-01

    The utility of [(18)F]WC-4-116, a PET tracer for imaging caspase-3 activation, was evaluated in an animal model of myocardial apoptosis. [(18)F]WC-4-116 was injected into rats at 3 hours after a 30 min period of ischemia induced by temporary occlusion of the left anterior descending coronary artery in Sprague-Dawley rats. [(18)F]WC-4-116 uptake was quantified by 1) autoradiography, 2) microPET imaging studies, and 3) post-PET biodistribution studies. MicroPET imaging also assessed uptake of the non-caspase-3-targeted tracer [(18)F]ICMT-18 at 3 hours postischemia. Enzyme assays and Western blotting assessed caspase-3 activation in both at-risk and not-at-risk regions. Caspase-3 enzyme activity increased in the at-risk but not in the not-at-risk myocardium. Quantitative autoradiographic analysis of [(18)F]WC-4-116 demonstrated nearly 2-fold higher uptake in the ischemia-reperfusion (IR) versus sham animals. [(18)F]WC-4-116 microPET imaging studies demonstrated that the IR animals was similarly elevated in relation to sham. [(18)F]ICMT-18 uptake did not increase in at-risk myocardium despite evidence of caspase-3 activation. Biodistribution studies with [(18)F]WC-4-116 confirmed the microPET findings. These data indicate that the caspase-3-PET tracer [(18)F]WC-4-116 can noninvasively image in vivo caspase activity during myocardial apoptosis and may be useful for clinical imaging in humans. PMID:27186438

  4. Clinical Application of in-room PET for in vivo Treatment Monitoring in Proton Radiotherapy

    PubMed Central

    Min, Chul Hee; Zhu, Xuping; Winey, Brian A.; Grogg, Kira; Testa, Mauro; Fakhri, Georges El; Bortfeld, Thomas R.; Paganetti, Harald; Shih, Helen A.

    2013-01-01

    Purpose/Objective(s) The purpose of this study is to evaluate the potential of using an in-room PET for treatment verification in proton therapy and to derive suitable PET scan times. Materials/Methods Nine patients undergoing passive scattering proton therapy were scanned immediately after treatment with an in-room PET scanner. The scanner was positioned next to the treatment head after treatment. The Monte Carlo (MC) method was employed to reproduce PET activities for each patient. To assess the proton beam range uncertainty we designed a novel concept where the measured PET activity surface distal to the target at the end of range was compared with MC predictions. The repositioning of patients for the PET scan took on average about 2 minutes. The PET images were reconstructed considering varying scan times to test the scan time dependency of the method. Results The measured PET images show overall good spatial correlations with MC predictions. Some discrepancies could be attributed to uncertainties in the local elemental composition and biological washout. For 8 patients treated with a single field, the average range differences between PET measurements and CT-image-based MC results were less than 5 mm (< 3 mm for 6 of 8 patients) and root-mean-square deviations (RMSD) were 4-11 mm with PET-CT image co-registration errors of about 2 mm. Our results also show that a short-length PET scan of 5 minutes can yield similar results compared to a 20 minutes PET scan. Conclusions Our first clinical trials of 9 patients using an in-room PET system demonstrated its potential for in vivo treatment monitoring in proton therapy. For a quantitative range prediction with arbitrary shape of target volume, we suggest employing the distal PET activity surface. PMID:23391817

  5. Contribution of FDOPA PET to radiotherapy planning for advanced glioma

    NASA Astrophysics Data System (ADS)

    Dowson, Nicholas; Fay, Michael; Thomas, Paul; Jeffree, Rosalind; McDowall, Robert; Winter, Craig; Coulthard, Alan; Smith, Jye; Gal, Yaniv; Bourgeat, Pierrick; Salvado, Olivier; Crozier, Stuart; Rose, Stephen

    2014-03-01

    Despite radical treatment with surgery, radiotherapy and chemotherapy, advanced gliomas recur within months. Geographic misses in radiotherapy planning may play a role in this seemingly ineluctable recurrence. Planning is typically performed on post-contrast MRIs, which are known to underreport tumour volume relative to FDOPA PET scans. FDOPA PET fused with contrast enhanced MRI has demonstrated greater sensitivity and specificity than MRI alone. One sign of potential misses would be differences between gross target volumes (GTVs) defined using MRI alone and when fused with PET. This work examined whether such a discrepancy may occur. Materials and Methods: For six patients, a 75 minute PET scan using 3,4-dihydroxy-6-18F-fluoro-L-phynel-alanine (18F-FDOPA) was taken within 2 days of gadolinium enhanced MRI scans. In addition to standard radiotherapy planning by an experienced radiotherapy oncologist, a second gross target volume (GTV) was defined by an experienced nuclear medicine specialist for fused PET and MRI, while blinded to the radiotherapy plans. The volumes from standard radiotherapy planning were compared to the PET defined GTV. Results: The comparison indicated radiotherapy planning would change in several cases if FDOPA PET data was available. PET-defined contours were external to 95% prescribed dose for several patients. However, due to the radiotherapy margins, the discrepancies were relatively small in size and all received a dose of 50 Gray or more. Conclusions: Given the limited size of the discrepancies it is uncertain that geographic misses played a major role in patient outcome. Even so, the existence of discrepancies indicates that FDOPA PET could assist in better defining margins when planning radiotherapy for advanced glioma, which could be important for highly conformal radiotherapy plans.

  6. Identification of Antiviral Agents Targeting Hepatitis B Virus Promoter from Extracts of Indonesian Marine Organisms by a Novel Cell-Based Screening Assay

    PubMed Central

    Yamashita, Atsuya; Fujimoto, Yuusuke; Tamaki, Mayumi; Setiawan, Andi; Tanaka, Tomohisa; Okuyama-Dobashi, Kaori; Kasai, Hirotake; Watashi, Koichi; Wakita, Takaji; Toyama, Masaaki; Baba, Masanori; de Voogd, Nicole J.; Maekawa, Shinya; Enomoto, Nobuyuki; Tanaka, Junichi; Moriishi, Kohji

    2015-01-01

    The current treatments of chronic hepatitis B (CHB) face a limited choice of vaccine, antibody and antiviral agents. The development of additional antiviral agents is still needed for improvement of CHB therapy. In this study, we established a screening system in order to identify compounds inhibiting the core promoter activity of hepatitis B virus (HBV). We prepared 80 extracts of marine organisms from the coral reefs of Indonesia and screened them by using this system. Eventually, two extracts showed high inhibitory activity (>95%) and low cytotoxicity (66% to 77%). Solvent fractionation, column chromatography and NMR analysis revealed that 3,5-dibromo-2-(2,4-dibromophenoxy)-phenol (compound 1) and 3,4,5-tribromo-2-(2,4-dibromophenoxy)-phenol (compound 2), which are classified as polybrominated diphenyl ethers (PBDEs), were identified as anti-HBV agents in the extracts. Compounds 1 and 2 inhibited HBV core promoter activity as well as HBV production from HepG2.2.15.7 cells in a dose-dependent manner. The EC50 values of compounds 1 and 2 were 0.23 and 0.80 µM, respectively, while selectivity indexes of compound 1 and 2 were 18.2 and 12.8, respectively. These results suggest that our cell-based HBV core promoter assay system is useful to determine anti-HBV compounds, and that two PBDE compounds are expected to be candidates of lead compounds for the development of anti-HBV drugs. PMID:26561821

  7. Identification of Antiviral Agents Targeting Hepatitis B Virus Promoter from Extracts of Indonesian Marine Organisms by a Novel Cell-Based Screening Assay.

    PubMed

    Yamashita, Atsuya; Fujimoto, Yuusuke; Tamaki, Mayumi; Setiawan, Andi; Tanaka, Tomohisa; Okuyama-Dobashi, Kaori; Kasai, Hirotake; Watashi, Koichi; Wakita, Takaji; Toyama, Masaaki; Baba, Masanori; de Voogd, Nicole J; Maekawa, Shinya; Enomoto, Nobuyuki; Tanaka, Junichi; Moriishi, Kohji

    2015-11-01

    The current treatments of chronic hepatitis B (CHB) face a limited choice of vaccine, antibody and antiviral agents. The development of additional antiviral agents is still needed for improvement of CHB therapy. In this study, we established a screening system in order to identify compounds inhibiting the core promoter activity of hepatitis B virus (HBV). We prepared 80 extracts of marine organisms from the coral reefs of Indonesia and screened them by using this system. Eventually, two extracts showed high inhibitory activity (>95%) and low cytotoxicity (66% to 77%). Solvent fractionation, column chromatography and NMR analysis revealed that 3,5-dibromo-2-(2,4-dibromophenoxy)-phenol (compound 1) and 3,4,5-tribromo-2-(2,4-dibromophenoxy)-phenol (compound 2), which are classified as polybrominated diphenyl ethers (PBDEs), were identified as anti-HBV agents in the extracts. Compounds 1 and 2 inhibited HBV core promoter activity as well as HBV production from HepG2.2.15.7 cells in a dose-dependent manner. The EC50 values of compounds 1 and 2 were 0.23 and 0.80 µM, respectively, while selectivity indexes of compound 1 and 2 were 18.2 and 12.8, respectively. These results suggest that our cell-based HBV core promoter assay system is useful to determine anti-HBV compounds, and that two PBDE compounds are expected to be candidates of lead compounds for the development of anti-HBV drugs. PMID:26561821

  8. Characterization of the cloned full-length and a truncated human target of rapamycin: Activity, specificity, and enzyme inhibition as studied by a high capacity assay

    SciTech Connect

    Toral-Barza, Lourdes; Zhang Weiguo; Lamison, Craig; LaRocque, James; Gibbons, James; Yu, Ker . E-mail: yuk@wyeth.com

    2005-06-24

    The mammalian target of rapamycin (mTOR/TOR) is implicated in cancer and other human disorders and thus an important target for therapeutic intervention. To study human TOR in vitro, we have produced in large scale both the full-length TOR (289 kDa) and a truncated TOR (132 kDa) from HEK293 cells. Both enzymes demonstrated a robust and specific catalytic activity towards the physiological substrate proteins, p70 S6 ribosomal protein kinase 1 (p70S6K1) and eIF4E binding protein 1 (4EBP1), as measured by phosphor-specific antibodies in Western blotting. We developed a high capacity dissociation-enhanced lanthanide fluorescence immunoassay (DELFIA) for analysis of kinetic parameters. The Michaelis constant (K {sub m}) values of TOR for ATP and the His6-S6K substrate were shown to be 50 and 0.8 {mu}M, respectively. Dose-response and inhibition mechanisms of several known inhibitors, the rapamycin-FKBP12 complex, wortmannin and LY294002, were also studied in DELFIA. Our data indicate that TOR exhibits kinetic features of those shared by traditional serine/threonine kinases and demonstrate the feasibility for TOR enzyme screen in searching for new inhibitors.

  9. Molecular Imaging of Prostate Cancer: PET Radiotracers

    PubMed Central

    Jadvar, Hossein

    2012-01-01

    OBJECTIVE Recent advances in the fundamental understanding of the complex biology of prostate cancer have provided an increasing number of potential targets for imaging and treatment. The imaging evaluation of prostate cancer needs to be tailored to the various phases of this remarkably heterogeneous disease. CONCLUSION In this article, I review the current state of affairs on a range of PET radiotracers for potential use in the imaging evaluation of men with prostate cancer. PMID:22826388

  10. A novel photoinduced electron transfer (PET) primer technique for rapid real-time PCR detection of Cryptosporidium spp

    SciTech Connect

    Jothikumar, N. Hill, Vincent R.

    2013-06-28

    Highlights: •Uses a single-labeled fluorescent primer for real-time PCR. •The detection sensitivity of PET PCR was comparable to TaqMan PCR. •Melt curve analysis can be performed to confirm target amplicon production. •Conventional PCR primers can be converted to PET PCR primers. -- Abstract: We report the development of a fluorescently labeled oligonucleotide primer that can be used to monitor real-time PCR. The primer has two parts, the 3′-end of the primer is complimentary to the target and a universal 17-mer stem loop at the 5′-end forms a hairpin structure. A fluorescent dye is attached to 5′-end of either the forward or reverse primer. The presence of guanosine residues at the first and second position of the 3′ dangling end effectively quenches the fluorescence due to the photo electron transfer (PET) mechanism. During the synthesis of nucleic acid, the hairpin structure is linearized and the fluorescence of the incorporated primer increases several-fold due to release of the fluorescently labeled tail and the absence of guanosine quenching. As amplicons are synthesized during nucleic acid amplification, the fluorescence increase in the reaction mixture can be measured with commercially available real-time PCR instruments. In addition, a melting procedure can be performed to denature the double-stranded amplicons, thereby generating fluorescence peaks that can differentiate primer dimers and other non-specific amplicons if formed during the reaction. We demonstrated the application of PET-PCR for the rapid detection and quantification of Cryptosporidium parvum DNA. Comparison with a previously published TaqMan® assay demonstrated that the two real-time PCR assays exhibited similar sensitivity for a dynamic range of detection of 6000–0.6 oocysts per reaction. PET PCR primers are simple to design and less-expensive than dual-labeled probe PCR methods, and should be of interest for use by laboratories operating in resource

  11. The ADNI PET Core: 2015

    PubMed Central

    Jagust, William J.; Landau, Susan M.; Koeppe, Robert A.; Reiman, Eric M.; Chen, Kewei; Mathis, Chester A.; Price, Julie C.; Foster, Norman L.; Wang, Angela Y.

    2015-01-01

    INTRODUCTION This paper reviews the work done in the ADNI PET core over the past 5 years, largely concerning techniques, methods, and results related to amyloid imaging in ADNI. METHODS The PET Core has utilized [18F]florbetapir routinely on ADNI participants, with over 1600 scans available for download. Four different laboratories are involved in data analysis, and have examined factors such as longitudinal florbetapir analysis, use of FDG-PET in clinical trials, and relationships between different biomarkers and cognition. RESULTS Converging evidence from the PET Core has indicated that cross-sectional and longitudinal florbetapir analyses require different reference regions. Studies have also examined the relationship between florbetapir data obtained immediately after injection, which reflects perfusion, and FDG-PET results. Finally, standardization has included the translation of florbetapir PET data to a centiloid scale. CONCLUSION The PET Core has demonstrated a variety of methods for standardization of biomarkers such as florbetapir PET in a multicenter setting. PMID:26194311

  12. Pet Loss: Implications for Counselors.

    ERIC Educational Resources Information Center

    Sharkin, Bruce S.; Bahrick, Audrey S.

    1990-01-01

    Attempts to increase awareness of counselors about topic of pet loss. Discusses how counselors can be actively involved through practice, consultation, and research to help people deal with emotional impact of pet loss. (Author/NB)

  13. Helicase Assays

    PubMed Central

    Wang, Xin; Li, Jing; Diaz, Jason; You, Jianxin

    2016-01-01

    Helicases are a class of enzymes which are motor proteins using energy derived from ATP hydrolysis to move directionally along a nucliec acid phosphodiester backbone (such as DNA, RNA and DNA-RNA hybrids) and separate two annealed nucleic acid strands. Many cellular processes, such as transcription, DNA replication, recombination and DNA repair involve helicase activity. Here, we provide a protocol to analyze helicase activities in vitro. In this protocol, the DNA helicase protein Merkel cell polyomavirus large T-antigen was expressed in the mammalian cell line HEK293 and immoblized on an IgG resin. The helicase assay is performing while the protein is immoblized on IgG resin.

  14. The development, past achievements, and future directions of brain PET

    PubMed Central

    Jones, Terry; Rabiner, Eugenii A

    2012-01-01

    The early developments of brain positron emission tomography (PET), including the methodological advances that have driven progress, are outlined. The considerable past achievements of brain PET have been summarized in collaboration with contributing experts in specific clinical applications including cerebrovascular disease, movement disorders, dementia, epilepsy, schizophrenia, addiction, depression and anxiety, brain tumors, drug development, and the normal healthy brain. Despite a history of improving methodology and considerable achievements, brain PET research activity is not growing and appears to have diminished. Assessments of the reasons for decline are presented and strategies proposed for reinvigorating brain PET research. Central to this is widening the access to advanced PET procedures through the introduction of lower cost cyclotron and radiochemistry technologies. The support and expertize of the existing major PET centers, and the recruitment of new biologists, bio-mathematicians and chemists to the field would be important for such a revival. New future applications need to be identified, the scope of targets imaged broadened, and the developed expertize exploited in other areas of medical research. Such reinvigoration of the field would enable PET to continue making significant contributions to advance the understanding of the normal and diseased brain and support the development of advanced treatments. PMID:22434067

  15. The development, past achievements, and future directions of brain PET.

    PubMed

    Jones, Terry; Rabiner, Eugenii A

    2012-07-01

    The early developments of brain positron emission tomography (PET), including the methodological advances that have driven progress, are outlined. The considerable past achievements of brain PET have been summarized in collaboration with contributing experts in specific clinical applications including cerebrovascular disease, movement disorders, dementia, epilepsy, schizophrenia, addiction, depression and anxiety, brain tumors, drug development, and the normal healthy brain. Despite a history of improving methodology and considerable achievements, brain PET research activity is not growing and appears to have diminished. Assessments of the reasons for decline are presented and strategies proposed for reinvigorating brain PET research. Central to this is widening the access to advanced PET procedures through the introduction of lower cost cyclotron and radiochemistry technologies. The support and expertize of the existing major PET centers, and the recruitment of new biologists, bio-mathematicians and chemists to the field would be important for such a revival. New future applications need to be identified, the scope of targets imaged broadened, and the developed expertize exploited in other areas of medical research. Such reinvigoration of the field would enable PET to continue making significant contributions to advance the understanding of the normal and diseased brain and support the development of advanced treatments. PMID:22434067

  16. Angiogenesis Assays.

    PubMed

    Nambiar, Dhanya K; Kujur, Praveen K; Singh, Rana P

    2016-01-01

    Neoangiogenesis constitutes one of the first steps of tumor progression beyond a critical size of tumor growth, which supplies a dormant mass of cancerous cells with the required nutrient supply and gaseous exchange through blood vessels essentially needed for their sustained and aggressive growth. In order to understand any biological process, it becomes imperative that we use models, which could mimic the actual biological system as closely as possible. Hence, finding the most appropriate model is always a vital part of any experimental design. Angiogenesis research has also been much affected due to lack of simple, reliable, and relevant models which could be easily quantitated. The angiogenesis models have been used extensively for studying the action of various molecules for agonist or antagonistic behaviour and associated mechanisms. Here, we have described two protocols or models which have been popularly utilized for studying angiogenic parameters. Rat aortic ring assay tends to bridge the gap between in vitro and in vivo models. The chorioallantoic membrane (CAM) assay is one of the most utilized in vivo model system for angiogenesis-related studies. The CAM is highly vascularized tissue of the avian embryo and serves as a good model to study the effects of various test compounds on neoangiogenesis. PMID:26608294

  17. PET/CT imaging artifacts.

    PubMed

    Sureshbabu, Waheeda; Mawlawi, Osama

    2005-09-01

    The purpose of this paper is to introduce the principles of PET/CT imaging and describe the artifacts associated with it. PET/CT is a new imaging modality that integrates functional (PET) and structural (CT) information into a single scanning session, allowing excellent fusion of the PET and CT images and thus improving lesion localization and interpretation accuracy. Moreover, the CT data can also be used for attenuation correction, ultimately leading to high patient throughput. These combined advantages have rendered PET/CT a preferred imaging modality over dedicated PET. Although PET/CT imaging offers many advantages, this dual-modality imaging also poses some challenges. CT-based attenuation correction can induce artifacts and quantitative errors that can affect the PET emission images. For instance, the use of contrast medium and the presence of metallic implants can be associated with focal radiotracer uptake. Furthermore, the patient's breathing can introduce mismatches between the CT attenuation map and the PET emission data, and the discrepancy between the CT and PET fields of view can lead to truncation artifacts. After reading this article, the technologist should be able to describe the principles of PET/CT imaging, identify at least 3 types of image artifacts, and describe the differences between PET/CT artifacts of different causes: metallic implants, respiratory motion, contrast medium, and truncation. PMID:16145223

  18. An Educational PET Camera Model

    ERIC Educational Resources Information Center

    Johansson, K. E.; Nilsson, Ch.; Tegner, P. E.

    2006-01-01

    Positron emission tomography (PET) cameras are now in widespread use in hospitals. A model of a PET camera has been installed in Stockholm House of Science and is used to explain the principles of PET to school pupils as described here.

  19. Transporter assays and assay ontologies: useful tools for drug discovery.

    PubMed

    Zdrazil, Barbara; Chichester, Christine; Zander Balderud, Linda; Engkvist, Ola; Gaulton, Anna; Overington, John P

    2014-06-01

    Transport proteins represent an eminent class of drug targets and ADMET (absorption, distribution, metabolism, excretion, toxicity) associated genes. There exists a large number of distinct activity assays for transport proteins, depending on not only the measurement needed (e.g. transport activity, strength of ligand–protein interaction), but also due to heterogeneous assay setups used by different research groups. Efforts to systematically organize this (divergent) bioassay data have large potential impact in Public-Private partnership and conventional commercial drug discovery. In this short review, we highlight some of the frequently used high-throughput assays for transport proteins, and we discuss emerging assay ontologies and their application to this field. Focusing on human P-glycoprotein (Multidrug resistance protein 1; gene name: ABCB1, MDR1), we exemplify how annotation of bioassay data per target class could improve and add to existing ontologies, and we propose to include an additional layer of metadata supporting data fusion across different bioassays. PMID:25027375

  20. Immuno-PET Imaging of HER3 in a Model in which HER3 Signaling Plays a Critical Role

    PubMed Central

    Yuan, Qinghua; Furukawa, Takako; Tashiro, Takahiro; Okita, Kouki; Jin, Zhao-Hui; Aung, Winn; Sugyo, Aya; Nagatsu, Kotaro; Endo, Hiroko; Tsuji, Atsushi B.; Zhang, Ming-Rong; Masuko, Takashi; Inoue, Masahiro; Fujibayashi, Yasuhisa; Saga, Tsuneo

    2015-01-01

    HER3 is overexpressed in various carcinomas including colorectal cancer (CRC), which is associated with poor prognosis, and is involved in the development of therapy resistance. Thus, an in vivo imaging technique is needed to evaluate the expression of HER3, an important therapeutic and diagnostic target. Here, we report successful HER3 PET imaging using a newly generated anti-human HER3 monoclonal antibody, Mab#58, and a mouse model of a HER3-overexpressing xenograft tumor. Furthermore, we assessed the role of HER3 signaling in CRC cancer tissue-originated spheroid (CTOS) and applied HER3 imaging to detect endogenous HER3 in CTOS-derived xenografts. Cell binding assays of 89Zr-labeled Mab#58 using the HER3-overexpressing cell line HER3/RH7777 demonstrated that [89Zr]Mab#58 specifically bound to HER3/RH7777 cells (Kd = 2.7 nM). In vivo biodistribution study in mice bearing HER3/RH7777 and its parent cell xenografts showed that tumor accumulation of [89Zr]Mab#58 in HER3/RH7777 xenografts was significantly higher than that in the control from day 1 to day 4, tending to increase from day 1 to day 4 and reaching 12.2 ± 4.5%ID/g. Radioactivity in other tissues, including the control xenograft, decreased or remained unchanged from day 1 to day 6. Positron emission tomography (PET) in the same model enabled clear visualization of HER3/RH7777 xenografts but not of RH7777 xenografts. CTOS growth assay and signaling assay revealed that CRC CTOS were dependent on HER3 signaling for their growth. In PET studies of mice bearing a CRC CTOS xenograft, the tumor was clearly visualized with [89Zr]Mab#58 but not with the 89Zr-labeled control antibody. Thus, tumor expression of HER3 was successfully visualized by PET with 89Zr-labeled anti-HER3 antibody in CTOS xenograft-bearing mice, a model that retains the properties of the patient tumor. Non-invasive targeting of HER3 by antibodies is feasible, and it is expected to be useful for cancer diagnosis and treatment. PMID:26571416

  1. PET in Cerebrovascular Disease

    PubMed Central

    Powers, William J.; Zazulia, Allyson R.

    2010-01-01

    SYNOPSIS Investigation of the interplay between the cerebral circulation and brain cellular function is fundamental to understanding both the pathophysiology and treatment of stroke. Currently, PET is the only technique that provides accurate, quantitative in vivo regional measurements of both cerebral circulation and cellular metabolism in human subjects. We review normal human cerebral blood flow and metabolism and human PET studies of ischemic stroke, carotid artery disease, vascular dementia, intracerebral hemorrhage and aneurysmal subarachnoid hemorrhage and discuss how these studies have added to our understanding of the pathophysiology of human cerebrovascular disease. PMID:20543975

  2. Detection of Shigella by a PCR Assay Targeting the ipaH Gene Suggests Increased Prevalence of Shigellosis in Nha Trang, Vietnam

    PubMed Central

    Thiem, Vu Dinh; Sethabutr, Orntipa; von Seidlein, Lorenz; Van Tung, Tran; Canh, Do Gia; Chien, Bui Trong; Tho, Le Huu; Lee, Hyejon; Houng, Huo-Shu; Hale, Thomas L.; Clemens, John D.; Mason, Carl; Trach, Dang Duc

    2004-01-01

    Shigella spp. are exquisitely fastidious gram-negative organisms which frequently escape detection by traditional culture methods. To get a more complete understanding of the disease burden caused by Shigella in Nha Trang, Vietnam, real-time PCR was used to detect Shigella DNA. Randomly selected rectal swab specimens from 60 Shigella culture-positive patients and 500 Shigella culture-negative patients detected by population-based surveillance of patients seeking care for diarrhea were processed by real-time PCR. The target of the primer pair is the invasion plasmid antigen H gene sequence (ipaH), carried by all four Shigella species and enteroinvasive Escherichia coli. Shigella spp. could be isolated from the rectal swabs of 547 of 19,206 (3%) patients with diarrhea. IpaH was detected in 55 of 60 (93%) Shigella culture-positive specimens, whereas it was detected in 87 of 245 (36%) culture-negative patients free of dysentery (P < 0.001). The number of PCR cycles required to detect a PCR product was highest for culture-negative, nonbloody diarrheal specimens (mean number of cycles to detection, 36.6) and was lowest for children with culture-positive, bloody diarrheal specimens (mean number of cycles, 25.3) (P < 0.001). The data from real-time PCR amplification indicate that the culture-proven prevalence of Shigella among patients with diarrhea may underestimate the prevalence of Shigella infections. The clinical presentation of shigellosis may be directly related to the bacterial load. PMID:15131166

  3. Development and Use of Assay Conditions Suited to Screening for and Profiling of SET-Domain-Targeted Inhibitors of the MLL/SET1 Family of Lysine Methyltransferases

    PubMed Central

    Ferry, Joseph J.; Smith, Robert F.; Denney, Natalie; Walsh, Colin P.; McCauley, Lauren; Qian, Jie; Ma, Haiching; Horiuchi, Kurumi Y.

    2015-01-01

    Abstract Methylation of histone H3 lysine-4 (H3K4) is an important, regulatory, epigenetic post-translational modification associated with actively transcribed genes. In humans, the principal mediators of this modification are part of the MLL/SET1 family of methyltransferases, which comprises six members, MLLs1–4 and SET1A/SET1B. Aberrations in the structure, expression, and regulation of these enzymes are implicated in various disease states, making them important potential targets for drug discovery, particularly for oncology indications. The MLL/SET1 family members are most enzymatically active when part of a “core complex,” the catalytic SET-domain-containing subunits bound to a subcomplex consisting of the proteins WDR5, RbBP5, Ash2L and a homodimer of DPY-30 (WRAD2). The necessity of MLL/SET1 members to bind WRAD2 for full activity is the basis of a particular drug development strategy, which seeks to disrupt the interaction between the MLL/SET1 subunits and WDR5. This strategy is not without its theoretical and practical drawbacks, some of which relate to the ease with which complexes of Escherichia coli-expressed MLL/SET1 and WRAD2 fall apart. As an alternative strategy, we explore ways to stabilize the complex, focusing on the use of an excess of WRAD2 to drive the binding equilibria toward complex formation while maintaining low concentrations of the catalytic subunits. The purpose of this approach is to seek inhibitors that bind the SET domain, an approach proven successful with the related, but inherently more stable, enhancer of zeste homolog 2 (EZH2) complex. PMID:26065558

  4. PET/MR in dementia and other neurodegenerative diseases.

    PubMed

    Barthel, Henryk; Schroeter, Matthias L; Hoffmann, Karl-Titus; Sabri, Osama

    2015-05-01

    The spectrum of neurodegenerative diseases covers the dementias, parkinsonian syndromes, Huntington disease, amyotrophic lateral sclerosis, and prion diseases. In these entities, brain MRI is often used in clinical routine to exclude other pathologies and to demonstrate specific atrophy patterns. [18F]FDG PET delivers early and sensitive readouts of neural tissue loss, and more specific PET tracers currently in use clinically target β-amyloid plaques or dopaminergic deficiency. The recent integration of PET into MR technology offers a new chance to improve early and differential diagnosis of many neurodegenerative diseases. Initial evidence in the literature is available to support this notion. New emerging PET tracers, such as tracers that bind to tau or α-synuclein aggregates, as well as MR techniques, like diffusion-tensor imaging, resting-state functional MRI, and arterial spin labeling, have the potential to broaden the diagnostic capabilities of combined PET/MRI to image dementias, Parkinson disease, and other neurodegenerative diseases. The ultimate goal is to establish combined PET/MRI as a first-line imaging technique to provide, in a one-stop-shop fashion with improved patient comfort, all biomarker information required to increase diagnostic confidence toward specific diagnoses. The technical challenge of accurate PET data attenuation correction within PET/MRI systems needs yet to be solved. Apart from the projected clinical routine applications, future research would need to answer the questions of whether combined brain PET/MRI is able to improve basic research of neurodegenerative diseases and antineurodegeneration drug testing. PMID:25841277

  5. Quantitative assessment of human and pet exposure to Salmonella associated with dry pet foods.

    PubMed

    Lambertini, Elisabetta; Buchanan, Robert L; Narrod, Clare; Ford, Randall M; Baker, Robert C; Pradhan, Abani K

    2016-01-01

    Recent Salmonella outbreaks associated with dry pet foods and treats highlight the importance of these foods as previously overlooked exposure vehicles for both pets and humans. In the last decade efforts have been made to raise the safety of this class of products, for instance by upgrading production equipment, cleaning protocols, and finished product testing. However, no comprehensive or quantitative risk profile is available for pet foods, thus limiting the ability to establish safety standards and assess the effectiveness of current and proposed Salmonella control measures. This study sought to develop an ingredients-to-consumer quantitative microbial exposure assessment model to: 1) estimate pet and human exposure to Salmonella via dry pet food, and 2) assess the impact of industry and household-level mitigation strategies on exposure. Data on prevalence and concentration of Salmonella in pet food ingredients, production process parameters, bacterial ecology, and contact transfer in the household were obtained through literature review, industry data, and targeted research. A probabilistic Monte Carlo modeling framework was developed to simulate the production process and basic household exposure routes. Under the range of assumptions adopted in this model, human exposure due to handling pet food is null to minimal if contamination occurs exclusively before extrusion. Exposure increases considerably if recontamination occurs post-extrusion during coating with fat, although mean ingested doses remain modest even at high fat contamination levels, due to the low percent of fat in the finished product. Exposure is highly variable, with the distribution of doses ingested by adult pet owners spanning 3Log CFU per exposure event. Child exposure due to ingestion of 1g of pet food leads to significantly higher doses than adult doses associated with handling the food. Recontamination after extrusion and coating, e.g., via dust or equipment surfaces, may also lead to

  6. Loop-mediated isothermal amplification assay targeting the blaCTX-M9 gene for detection of extended spectrum β-lactamase-producing Escherichia coli and Klebsiella pneumoniae.

    PubMed

    Thirapanmethee, Krit; Pothisamutyothin, Kanokporn; Nathisuwan, Surakit; Chomnawang, Mullika T; Wiwat, Chanpen

    2014-12-01

    Extended-spectrum β-lactamases (ESBLs) produced by Enterobacteriaceae are one of the resistance mechanisms to most β-lactam antibiotics. ESBLs are currently a major problem in both hospitals and community settings worldwide. Rapid and reliable means of detecting ESBL-producing bacteria is necessary for identification, prevention and treatment. Loop-mediated isothermal amplification (LAMP) is a technique that rapidly amplifies DNA with high specificity and sensitivity under isothermal conditions. This study was aimed to develop a convenient, accurate and inexpensive method for detecting ESBL-producing bacteria by a LAMP technique. ESBLs-producing Escherichia coli and Klebsiella pneumoniae were isolated from a tertiary hospital in Bangkok, Thailand and reconfirmed by double-disk synergy test. A set of four specific oligonucleotide primers of LAMP for detection of bla(CTX-M9) gene was designed based on bla(CTX-M9) from E. coli (GenBank Accession No. AJ416345). The LAMP reaction was amplified under isothermal temperature at 63°C for 60 min. Ladder-like patterns of band sizes from 226 bp of the bla(CTX-M9) DNA target was observed. The LAMP product was further analyzed by restriction digestion with MboI and TaqI endonucleases. The fragments generated were approximately 168, 177 and 250 bp in size for MboI digestion and 165, 193, 229, 281 and 314 bp for TaqI digestion, which is in agreement with the predicted sizes. The sensitivity of the LAMP technique to bla(CTX-M9) was greater than that of the PCR method by at least 10,000-fold. These results showed that the LAMP primers specifically amplified only the bla(CTX-M9) gene. Moreover, the presence of LAMP amplicon was simply determined by adding SYBR Green I in the reaction. In conclusion, this technique for detection of ESBLs is convenient, reliable and easy to perform routinely in hospitals or laboratory units in developing countries. PMID:25284314

  7. [Pets, veterinarians, and multicultural society].

    PubMed

    Klumpers, M; Endenburg, N

    2009-01-15

    Dutch society comprises a growing percentage of non-Western ethnic minority groups. Little is known about pet ownership among these groups. This study explores some aspects of pet ownership, and the position of veterinarians, among the four largest non-Western ethnic minority groups in the Netherlands. Information was gathered through street interviews with people from a Moroccan, Turkish, Surinamese, or Antillean (including Aruban) background. Five hundred people where interviewed, including 41 pet owners. Results showed that people from non-Western ethnic minorities kept pets less often than Dutch people, with fish and birds being the most frequently kept pets. The number of visits to the veterinary clinic was comparable to that of Dutch pet owners; however, reasons given for the last visit were different. People from non-Western ethnic minorities mostly visited a veterinarian if their pet was ill whereas Dutch people visited the veterinarian if their pet needed to be vaccinated. People from non-Western ethnic minorities were positive about veterinarians, considering that they had sufficient knowledge about and concern for their pets. Moreover, veterinarians were trusted and provided understandable information--the respondents felt that they could go to their veterinarian with any question or problem regarding their pets. Although most respondents considered a visit to the veterinarian expensive, they were more than willing to invest in their pet's health. PMID:19235301

  8. PET/Computed Tomography in the Individualization of Treatment of Prostate Cancer.

    PubMed

    Ceci, Francesco; Castellucci, Paolo; Graziani, Tiziano; Schiavina, Riccardo; Fanti, Stefano

    2015-10-01

    Choline PET/computed tomographic (CT) imaging represents the most diffused PET imaging techniques to investigate patients with prostate cancer (PCa). It may show the site of tumor recurrence in a single step examination, earlier than other conventional imaging techniques. In this context, the availability of a diagnostic test capable of differentiating between potentially curable local recurrence and metastatic disease implying palliative approaches may play an important role in those patients in whom targeted therapies could be performed according to choline PET/CT results. This review analyzes the value of choline PET/CT imaging in the evaluation of treatment of patients with PCa. PMID:26384595

  9. MR/PET or PET/MRI: does it matter?

    PubMed

    Beyer, Thomas; Moser, Ewald

    2013-02-01

    After the very successful clinical introduction of combined PET/CT imaging a decade ago, a hardware combination of PET and MR is following suit. Today, three different approaches towards integrated PET/MR have been proposed: (1) a triple-modality system with a 3T MRI and a time-of-flight PET/CT installed in adjacent rooms, (2) a tandem system with a 3T MRI and a time-of-flight PET/CT in a co-planar installation with a joint patient handling system, and (3) a fully-integrated system with a whole-body PET system mounted inside a 3T MRI system. This special issue of MAGMA brings together contributions from key experts in the field of PET/MR, PET/CT and CT. The various papers share the author's perspectives on the state-of-the-art PET/MR imaging with any of the three approaches mentioned above. In addition to several reviews discussing advantages and challenges of combining PET and MRI for clinical diagnostics, first clinical data are also presented. We expect this special issue to nurture future improvements in hardware, clinical protocols, and efficient post-processing strategies to further assess the diagnostic value of combined PET/MR imaging. It remains to be seen whether a so-called "killer application" for PET/MRI will surface. In that case PET/MR is likely to excel in pre-clinical and selected research applications for now. This special issue helps the readers to stay on track of this exciting development. PMID:23385880

  10. ImmunoPET Imaging of CD146 Expression in Malignant Brain Tumors.

    PubMed

    Hernandez, Reinier; Sun, Haiyan; England, Christopher G; Valdovinos, Hector F; Barnhart, Todd E; Yang, Yunan; Cai, Weibo

    2016-07-01

    Recently, the overexpression of CD146 and its potential as a therapeutic target in high-grade gliomas, the most lethal type of brain cancer, was uncovered. In this study, we describe the generation of (89)Zr-Df-YY146, a novel (89)Zr-labeled monoclonal antibody (mAb) for the targeting and quantification of CD146 expression in a mouse model of glioblastoma, using noninvasive immunoPET imaging. YY146, a high affinity anti-CD146 mAb, was conjugated to deferoxamine (Df) for labeling with the long-lived positron emitter (89)Zr (t1/2: 78.4 h). In vitro assays, including flow cytometry, immunofluorescence microscopy, and Western blot, were performed with two glioblastoma cell lines, U87MG and U251, to determine their CD146 expression levels. Also, YY146 and Df-YY146's CD146-binding affinities were compared using flow cytometry. In vivo CD146-targeting of (89)Zr-Df-YY146 was evaluated by sequential PET imaging, in athymic nude mice bearing subcutaneously implanted U87MG or U251 tumors. CD146 blocking, ex vivo biodistribution, and histological studies were carried out to confirm (89)Zr-Df-YY146 specificity, as well as the accuracy of PET data. In vitro studies exposed elevated CD146 expression levels in U87MG cells, but negligible levels in U251 cells. Flow cytometry revealed no differences in affinity between YY146 and Df-YY146. (89)Zr labeling of Df-YY146 proceeded with excellent yield (∼80%), radiochemical purity (>95%), and specific activity (∼44 GBq/μmol). Longitudinal PET revealed prominent and persistent (89)Zr-Df-YY146 uptake in mice bearing U87MG tumors that peaked at 14.00 ± 3.28%ID/g (n = 4), 48 h post injection of the tracer. Conversely, uptake was significantly lower in CD146-negative U251 tumors (5.15 ± 0.99%ID/g, at 48 h p.i.; n = 4; P < 0.05). Uptake in U87MG tumors was effectively blocked in a competitive inhibition experiment, corroborating the CD146 specificity of (89)Zr-Df-YY146. Finally, ex vivo biodistribution validated the accuracy of PET data

  11. Breast Cancer Imaging with Novel PET Tracers.

    PubMed

    Mankoff, David A; Lee, Jean H; Eubank, William B

    2009-10-01

    Whereas (18)F-fluorodeoxyglucose (FDG)-PET/computed tomography has proven to be valuable for breast cancer diagnosis and response evaluation, it is likely that PET radiopharmaceuticals beyond FDG will contribute further to the understanding of breast cancer and thereby further direct breast cancer care. Increasingly specific and quantitative approaches will help direct treatment selection from an ever-expanding and increasing array of targeted breast cancer therapies. This article highlights 4 areas of ongoing research where preliminary patient results look promising: (1) tumor perfusion and angiogenesis, (2) drug delivery and transport, (3) tumor receptor imaging, and (4) early response evaluation. For each area, the biologic background is reviewed and early results are highlighted. PMID:27157306

  12. 24 CFR 960.707 - Pet ownership.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 24 Housing and Urban Development 4 2011-04-01 2011-04-01 false Pet ownership. 960.707 Section 960... ADMISSION TO, AND OCCUPANCY OF, PUBLIC HOUSING Pet Ownership in Public Housing § 960.707 Pet ownership. (a..., may own one or more common household pets or have one or more common household pets present in...

  13. 24 CFR 960.707 - Pet ownership.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 24 Housing and Urban Development 4 2014-04-01 2014-04-01 false Pet ownership. 960.707 Section 960... ADMISSION TO, AND OCCUPANCY OF, PUBLIC HOUSING Pet Ownership in Public Housing § 960.707 Pet ownership. (a..., may own one or more common household pets or have one or more common household pets present in...

  14. Talking with Children about Furry Classroom Pets.

    ERIC Educational Resources Information Center

    Texas Child Care, 1994

    1994-01-01

    Notes that rodents and rabbits share many characteristics that make them suitable classroom pets and gives background information on rabbits, guinea pigs, hamsters, and gerbils. Offers advice on buying a classroom pet, the pet's home, feeding, helping the children handle the pet, and pet health and family planning. (TJQ)

  15. Imaging of Lymph Node Micrometastases Using an Oncolytic Herpes Virus and [18F]FEAU PET

    PubMed Central

    Brader, Peter; Kelly, Kaitlyn; Gang, Sheng; Shah, Jatin P.; Wong, Richard J.; Hricak, Hedvig; Blasberg, Ronald G.; Fong, Yuman; Gil, Ziv

    2009-01-01

    Background In patients with melanoma, knowledge of regional lymph node status provides important information on outlook. Since lymph node status can influence treatment, surgery for sentinel lymph node (SLN) biopsy became a standard staging procedure for these patients. Current imaging modalities have a limited sensitivity for detection of micrometastases in lymph nodes and, therefore, there is a need for a better technique that can accurately identify occult SLN metastases. Methodology/Principal Findings B16-F10 murine melanoma cells were infected with replication-competent herpes simplex virus (HSV) NV1023. The presence of tumor-targeting and reporter-expressing virus was assessed by [18F]-2′-fluoro-2′-deoxy-1-β-D-β-arabinofuranosyl-5-ethyluracil ([18F]FEAU) positron emission tomography (PET) and confirmed by histochemical assays. An animal foot pad model of melanoma lymph node metastasis was established. Mice received intratumoral injections of NV1023, and 48 hours later were imaged after i.v. injection of [18F]FEAU. NV1023 successfully infected and provided high levels of lacZ transgene expression in melanoma cells. Intratumoral injection of NV1023 resulted in viral trafficking to melanoma cells that had metastasized to popliteal and inguinal lymph nodes. Presence of virus-infected tumor cells was successfully imaged with [18F]FEAU-PET, that identified 8 out of 8 tumor-positive nodes. There was no overlap between radioactivity levels (lymph node to surrounding tissue ratio) of tumor-positive and tumor-negative lymph nodes. Conclusion/Significance A new approach for imaging SLN metastases using NV1023 and [18F]FEAU-PET was successful in a murine model. Similar studies could be translated to the clinic and improve the staging and management of patients with melanoma. PMID:19274083

  16. [The PET, Past and Future].

    PubMed

    Fujii, Hirofumi

    2015-01-01

    Positron emission tomography (PET) is a unique nuclear medicine test using positron emitters such as 18F and 11C. In PET tests, various kinds of functional aspects of human bodies can be evaluated by using compounds labeled by these positron emitters. Recently, combined scanners of PET and anatomical imaging modalities such as CT and MRI have been developed and functional information with anatomical location can be easily obtained, increasing the usefulness of PET tests. PET tests are now essential imaging tools to diagnose various kinds of disease with functional abnormalities. In the field of oncology, 18F-fluorodeoxy glucose PET tests are routinely used in clinical practice under health insurance. In the field of neurology, PET tests are actively used to investigate cerebral function by labeled neurotransmitters and so on. Currently, brain PET tests to detect beta-amyloid are applied to the diagnosis of dementia. In the field of cardiology, cardiac perfusion and myocardial metabolism are quantitatively measured by using PET and obtained results have successfully revealed the pathogenesis of intractable cardiac diseases. Future technical advances will enhance the usefulness of PET tests more and more. PMID:26753390

  17. Clinical evaluation of 4D PET motion compensation strategies for treatment verification in ion beam therapy.

    PubMed

    Gianoli, Chiara; Kurz, Christopher; Riboldi, Marco; Bauer, Julia; Fontana, Giulia; Baroni, Guido; Debus, Jürgen; Parodi, Katia

    2016-06-01

    A clinical trial named PROMETHEUS is currently ongoing for inoperable hepatocellular carcinoma (HCC) at the Heidelberg Ion Beam Therapy Center (HIT, Germany). In this framework, 4D PET-CT datasets are acquired shortly after the therapeutic treatment to compare the irradiation induced PET image with a Monte Carlo PET prediction resulting from the simulation of treatment delivery. The extremely low count statistics of this measured PET image represents a major limitation of this technique, especially in presence of target motion. The purpose of the study is to investigate two different 4D PET motion compensation strategies towards the recovery of the whole count statistics for improved image quality of the 4D PET-CT datasets for PET-based treatment verification. The well-known 4D-MLEM reconstruction algorithm, embedding the motion compensation in the reconstruction process of 4D PET sinograms, was compared to a recently proposed pre-reconstruction motion compensation strategy, which operates in sinogram domain by applying the motion compensation to the 4D PET sinograms. With reference to phantom and patient datasets, advantages and drawbacks of the two 4D PET motion compensation strategies were identified. The 4D-MLEM algorithm was strongly affected by inverse inconsistency of the motion model but demonstrated the capability to mitigate the noise-break-up effects. Conversely, the pre-reconstruction warping showed less sensitivity to inverse inconsistency but also more noise in the reconstructed images. The comparison was performed by relying on quantification of PET activity and ion range difference, typically yielding similar results. The study demonstrated that treatment verification of moving targets could be accomplished by relying on the whole count statistics image quality, as obtained from the application of 4D PET motion compensation strategies. In particular, the pre-reconstruction warping was shown to represent a promising choice when combined with intra

  18. Clinical evaluation of 4D PET motion compensation strategies for treatment verification in ion beam therapy

    NASA Astrophysics Data System (ADS)

    Gianoli, Chiara; Kurz, Christopher; Riboldi, Marco; Bauer, Julia; Fontana, Giulia; Baroni, Guido; Debus, Jürgen; Parodi, Katia

    2016-06-01

    A clinical trial named PROMETHEUS is currently ongoing for inoperable hepatocellular carcinoma (HCC) at the Heidelberg Ion Beam Therapy Center (HIT, Germany). In this framework, 4D PET-CT datasets are acquired shortly after the therapeutic treatment to compare the irradiation induced PET image with a Monte Carlo PET prediction resulting from the simulation of treatment delivery. The extremely low count statistics of this measured PET image represents a major limitation of this technique, especially in presence of target motion. The purpose of the study is to investigate two different 4D PET motion compensation strategies towards the recovery of the whole count statistics for improved image quality of the 4D PET-CT datasets for PET-based treatment verification. The well-known 4D-MLEM reconstruction algorithm, embedding the motion compensation in the reconstruction process of 4D PET sinograms, was compared to a recently proposed pre-reconstruction motion compensation strategy, which operates in sinogram domain by applying the motion compensation to the 4D PET sinograms. With reference to phantom and patient datasets, advantages and drawbacks of the two 4D PET motion compensation strategies were identified. The 4D-MLEM algorithm was strongly affected by inverse inconsistency of the motion model but demonstrated the capability to mitigate the noise-break-up effects. Conversely, the pre-reconstruction warping showed less sensitivity to inverse inconsistency but also more noise in the reconstructed images. The comparison was performed by relying on quantification of PET activity and ion range difference, typically yielding similar results. The study demonstrated that treatment verification of moving targets could be accomplished by relying on the whole count statistics image quality, as obtained from the application of 4D PET motion compensation strategies. In particular, the pre-reconstruction warping was shown to represent a promising choice when combined with intra

  19. Human Immunodeficiency Virus-Specific Gamma Interferon Enzyme-Linked Immunospot Assay Responses Targeting Specific Regions of the Proteome during Primary Subtype C Infection Are Poor Predictors of the Course of Viremia and Set Point▿

    PubMed Central

    Gray, Clive M.; Mlotshwa, Mandla; Riou, Catherine; Mathebula, Tiyani; de Assis Rosa, Debra; Mashishi, Tumelo; Seoighe, Cathal; Ngandu, Nobubelo; van Loggerenberg, Francois; Morris, Lynn; Mlisana, Koleka; Williamson, Carolyn; Karim, Salim Abdool

    2009-01-01

    It is unknown whether patterns of human immunodeficiency virus (HIV)-specific T-cell responses during acute infection may influence the viral set point and the course of disease. We wished to establish whether the magnitude and breadth of HIV type 1 (HIV-1)-specific T-cell responses at 3 months postinfection were correlated with the viral-load set point at 12 months and hypothesized that the magnitude and breadth of HIV-specific T-cell responses during primary infection would predict the set point. Gamma interferon (IFN-γ) enzyme-linked immunospot (ELISPOT) assay responses across the complete proteome were measured in 47 subtype C HIV-1-infected participants at a median of 12 weeks postinfection. When corrected for amino acid length and individuals responding to each region, the order of recognition was as follows: Nef > Gag > Pol > Rev > Vpr > Env > Vpu > Vif > Tat. Nef responses were significantly (P < 0.05) dominant, targeted six epitopic regions, and were unrelated to the course of viremia. There was no significant difference in the magnitude and breadth of responses for each protein region with disease progression, although there was a trend of increased breadth (mean, four to seven pools) in rapid progressors. Correlation of the magnitude and breadth of IFN-γ responses with the viral set point at 12 months revealed almost zero association for each protein region. Taken together, these data demonstrate that the magnitude and breadth of IFN-γ ELISPOT assay responses at 3 months postinfection are unrelated to the course of disease in the first year of infection and are not associated with, and have low predictive power for, the viral set point at 12 months. PMID:18945774

  20. Real-time qPCR is a powerful assay to estimate the 171 R/Q alleles at the PrP locus directly in a flock's raw milk: a comparison with the targeted next-generation sequencing.

    PubMed

    Feligini, Maria; Bongioni, Graziella; Brambati, Eva; Amadesi, Alessandra; Cambuli, Caterina; Panelli, Simona; Bonacina, Cesare; Galli, Andrea

    2014-10-01

    The hazard to human health represented by transmissible spongiform encephalopathies in sheep is one of the major reasons for implementing the genetic selection plan to break down prion diseases. The problem is particularly important because of the risk of disease transmission from ewe to lamb via milk or colostrum. In order to establish an active and convenient monitoring of the flocks already undergone genetic selection and thus, indirectly increase consumers' security, the challenge of the work was quantifying the classical scrapie risk in bulk milk. A new quantitative real-time PCR assay for the estimation of the 171 R and Q allelic frequencies in a DNA pool representative of all the lactating ewes present in a flock was optimized and validated "in field". The repeatability range was 3.69-5.27 for R and 4.20-5.75 for Q. The ruggedness of the allele frequencies resulted 4.26 for R and 4.77 for Q. Regarding the validation "in field", none of the considered sources of variability (flock, month, number of genotyped animals and somatic cell count) showed a significant effect on flock and milk at the linear model. The targeted next-generation sequencing was also tested to evaluate its applicability in this context. Results show that the real-time PCR assay could represent a valid tool for the determination of 171 R/Q allele frequencies in bulk milk. The implementation of a service for breeder self-control along the production chain would aim to increase the production of high-security dairy products, while monitoring over time of the effects of genetic selection in the flocks. PMID:25066278

  1. HIV-1 Fusion Assay

    PubMed Central

    Cavrois, Marielle; Neidleman, Jason; Greene, Warner C.

    2016-01-01

    The HIV-1 fusion assay measures all steps in the HIV-1 life cycle up to and including viral fusion. It relies on the incorporation of a β-lactamase Vpr (BlaM-Vpr) protein chimera into the virion and the subsequent transfer of this chimera into the target cell by fusion (Figure 1). The transfer is monitored by the enzymatic cleavage of CCF2, a fluorescent dye substrate of β-lactamase, loaded into the target cells. Cleavage of the β-lactam ring in CCF2 by β-lactamase changes the fluorescence emission spectrum of the dye from green (520 nm) to blue (447 nm). This change reflects virion fusion and can be detected by flow cytometry (Figure 2).

  2. Molecular inversion probe assay.

    PubMed

    Absalan, Farnaz; Ronaghi, Mostafa

    2007-01-01

    We have described molecular inversion probe technologies for large-scale genetic analyses. This technique provides a comprehensive and powerful tool for the analysis of genetic variation and enables affordable, large-scale studies that will help uncover the genetic basis of complex disease and explain the individual variation in response to therapeutics. Major applications of the molecular inversion probes (MIP) technologies include targeted genotyping from focused regions to whole-genome studies, and allele quantification of genomic rearrangements. The MIP technology (used in the HapMap project) provides an efficient, scalable, and affordable way to score polymorphisms in case/control populations for genetic studies. The MIP technology provides the highest commercially available multiplexing levels and assay conversion rates for targeted genotyping. This enables more informative, genome-wide studies with either the functional (direct detection) approach or the indirect detection approach. PMID:18025701

  3. Progress reported in PET recycling

    SciTech Connect

    Not Available

    1989-06-01

    The Goodyear Polyester Division has demonstrated its ability to break down polyethylene terephthalate (PET) from recycled plastic soft drink bottles and remanufacture the material into PET suitable for containers. Most people are familiar with PET in the form of lightweight, shatter resistant beverage bottles. About 20 percent of these beverage containers currently are being recycled. The recycled PET is currently used in many applications such as carpeting, pillow stuffing, sleeping bag filling, insulation for water heaters and non-food containers. This is the first step of Goodyear's increased efforts to recycle PET from containers into a material suitable for food packing. The project is extremely complex, involving sophisticated understanding of the chemical reactions involved, PET production and the technology testing protocols necessary to design a process that addresses all the technical, safety, and regulatory concerns. The research conducted so far indicated that additional processing beyond simply cleaning the shredded material, called flake, will be required to assure a quality polymer.

  4. RPC PET: Status and perspectives

    NASA Astrophysics Data System (ADS)

    Couceiro, M.; Blanco, A.; Ferreira, Nuno C.; Ferreira Marques, R.; Fonte, P.; Lopes, L.

    2007-10-01

    The status of the resistive plate chamber (RPC)-PET technology for small animals is briefly reviewed and its sensitivity performance for human PET studied through Monte-Carlo simulations. The cost-effectiveness of these detectors and their very good timing characteristics open the possibility to build affordable Time of Flight (TOF)-PET systems with very large fields of view. Simulations suggest that the sensitivity of such systems for human whole-body screening, under reasonable assumptions, may exceed the present crystal-based PET technology by a factor up to 20.

  5. Extended suicide with a pet.

    PubMed

    Cooke, Brian K

    2013-01-01

    The combination of the killing of a pet and a suicide is a perplexing scenario that is largely unexplored in the literature. Many forensic psychiatrists and psychologists may be unaccustomed to considering the significance of the killing of a pet. The subject is important, however, because many people regard their pets as members of their family. A case is presented of a woman who killed her pet dog and herself by carbon monoxide poisoning. The purpose of this article is to provide an initial exploration of the topic of extended suicide with a pet. Forensic mental health evaluations may have a role in understanding the etiology of this event and in opining as to the culpability of individuals who attempt to or successfully kill a pet and then commit suicide. Because the scientific literature is lacking, there is a need to understand this act from a variety of perspectives. First, a social and anthropological perspective will be presented that summarizes the history of the practice of killing of one's pet, with a focus on the ancient Egyptians. A clinical context will examine what relationship animals have to mental illness. A vast body of existing scientific data showing the relevance of human attachment to pets suggests that conclusions from the phenomena of homicide-suicide and filicide-suicide are applicable to extended suicide with a pet. Finally, recommendations will be proposed for both clinical and forensic psychiatrists faced with similar cases. PMID:24051598

  6. Post-PET ultrasound improves specificity of 18F-FDG-PET for recurrent differentiated thyroid cancer while maintaining sensitivity

    PubMed Central

    Kråkenes, Jostein; Brauckhoff, Katrin; Haugland, Hans Kristian; Heinecke, Achim; Akslen, Lars A; Varhaug, Jan Erik; Brauckhoff, Michael

    2015-01-01

    Background Positron emission tomography (PET) using fluor-18-deoxyglucose (18F-FDG) with or without computed tomography (CT) is generally accepted as the most sensitive imaging modality for diagnosing recurrent differentiated thyroid cancer (DTC) in patients with negative whole body scintigraphy with iodine-131 (I-131). Purpose To assess the potential incremental value of ultrasound (US) over 18F-FDG-PET-CT. Material and Methods Fifty-one consecutive patients with suspected recurrent DTC were prospectively evaluated using the following multimodal imaging protocol: (i) US before PET (pre-US) with or without fine needle biopsy (FNB) of suspicious lesions; (ii) single photon emission computed tomography (≥3 GBq I-131) with co-registered CT (SPECT-CT); (iii) 18F-FDG-PET with co-registered contrast-enhanced CT of the neck; (iv) US in correlation with the other imaging modalities (post-US). Postoperative histology, FNB, and long-term follow-up (median, 2.8 years) were taken as composite gold standard. Results Fifty-eight malignant lesions were identified in 34 patients. Forty lesions were located in the neck or upper mediastinum. On receiver operating characteristics (ROC) analysis, 18F-FDG-PET had a limited lesion-based specificity of 59% at a set sensitivity of 90%. Pre-US had poor sensitivity and specificity of 52% and 53%, respectively, increasing to 85% and 94% on post-US, with knowledge of the PET/CT findings (P < 0.05 vs. PET and pre-US). Multimodal imaging changed therapy in 15 out of 51 patients (30%). Conclusion In patients with suspected recurrent DTC, supplemental targeted US in addition to 18F-FDG-PET-CT increases specificity while maintainin sensitivity, as non-malignant FDG uptake in cervical lesions can be confirmed. PMID:25770086

  7. PET/CT imaging in neuroblastoma.

    PubMed

    Piccardo, A; Lopci, E; Conte, M; Foppiani, L; Garaventa, A; Cabria, M; Villavecchia, G; Fanti, S; Cistaro, A

    2013-03-01

    123Iodine-metaiodobenzylguanidine (123I-MIBG) scintigraphy is currently the tracer of choice for neuroblastoma (NB). It has high diagnostic accuracy and prognostic value for the assessment of patients after chemotherapy. A positive 123I-MIBG scan is also used for the basis of targeted radionuclide therapy with 131I-MIBG. I-123 MIBG scan however has some limitations which should be taken into account. Moreover the reasons for false negative MIBG results have not been entirely elucidated. Meticulous correlation with radiological examinations and recognition of the normal distribution pattern of 123I-MIBG in children is vital to obtain optimal results. With its technical superiorities, positron emission tomography/computed tomography (PET/CT) can be successfully introduced into the diagnostic workup of NB. Different PET tracers have been offered for imaging in patients with NB, and the efficacy of this modality has been compared with that of 123I-MIBG scan. Our review aims to analyze the present role of PET/CT imaging and radiopharmaceuticals in NB. PMID:23474633

  8. Predicting standard-dose PET image from low-dose PET and multimodal MR images using mapping-based sparse representation

    NASA Astrophysics Data System (ADS)

    Wang, Yan; Zhang, Pei; An, Le; Ma, Guangkai; Kang, Jiayin; Shi, Feng; Wu, Xi; Zhou, Jiliu; Lalush, David S.; Lin, Weili; Shen, Dinggang

    2016-01-01

    Positron emission tomography (PET) has been widely used in clinical diagnosis for diseases and disorders. To obtain high-quality PET images requires a standard-dose radionuclide (tracer) injection into the human body, which inevitably increases risk of radiation exposure. One possible solution to this problem is to predict the standard-dose PET image from its low-dose counterpart and its corresponding multimodal magnetic resonance (MR) images. Inspired by the success of patch-based sparse representation (SR) in super-resolution image reconstruction, we propose a mapping-based SR (m-SR) framework for standard-dose PET image prediction. Compared with the conventional patch-based SR, our method uses a mapping strategy to ensure that the sparse coefficients, estimated from the multimodal MR images and low-dose PET image, can be applied directly to the prediction of standard-dose PET image. As the mapping between multimodal MR images (or low-dose PET image) and standard-dose PET images can be particularly complex, one step of mapping is often insufficient. To this end, an incremental refinement framework is therefore proposed. Specifically, the predicted standard-dose PET image is further mapped to the target standard-dose PET image, and then the SR is performed again to predict a new standard-dose PET image. This procedure can be repeated for prediction refinement of the iterations. Also, a patch selection based dictionary construction method is further used to speed up the prediction process. The proposed method is validated on a human brain dataset. The experimental results show that our method can outperform benchmark methods in both qualitative and quantitative measures.

  9. Predicting standard-dose PET image from low-dose PET and multimodal MR images using mapping-based sparse representation.

    PubMed

    Wang, Yan; Zhang, Pei; An, Le; Ma, Guangkai; Kang, Jiayin; Shi, Feng; Wu, Xi; Zhou, Jiliu; Lalush, David S; Lin, Weili; Shen, Dinggang

    2016-01-21

    Positron emission tomography (PET) has been widely used in clinical diagnosis for diseases and disorders. To obtain high-quality PET images requires a standard-dose radionuclide (tracer) injection into the human body, which inevitably increases risk of radiation exposure. One possible solution to this problem is to predict the standard-dose PET image from its low-dose counterpart and its corresponding multimodal magnetic resonance (MR) images. Inspired by the success of patch-based sparse representation (SR) in super-resolution image reconstruction, we propose a mapping-based SR (m-SR) framework for standard-dose PET image prediction. Compared with the conventional patch-based SR, our method uses a mapping strategy to ensure that the sparse coefficients, estimated from the multimodal MR images and low-dose PET image, can be applied directly to the prediction of standard-dose PET image. As the mapping between multimodal MR images (or low-dose PET image) and standard-dose PET images can be particularly complex, one step of mapping is often insufficient. To this end, an incremental refinement framework is therefore proposed. Specifically, the predicted standard-dose PET image is further mapped to the target standard-dose PET image, and then the SR is performed again to predict a new standard-dose PET image. This procedure can be repeated for prediction refinement of the iterations. Also, a patch selection based dictionary construction method is further used to speed up the prediction process. The proposed method is validated on a human brain dataset. The experimental results show that our method can outperform benchmark methods in both qualitative and quantitative measures. PMID:26732849

  10. SU-E-J-222: Evaluation of Deformable Registration of PET/CT Images for Cervical Cancer Brachytherapy

    SciTech Connect

    Liao, Y; Turian, J; Templeton, A; Kiel, K; Chu, J; Kadir, T

    2014-06-01

    Purpose: PET/CT provides important functional information for radiotherapy targeting of cervical cancer. However, repeated PET/CT procedures for external beam and subsequent brachytherapy expose patients to additional radiation and are not cost effective. Our goal is to investigate the possibility of propagating PET-active volumes for brachytherapy procedures through deformable image registration (DIR) of earlier PET/CT and ultimately to minimize the number of PET/CT image sessions required. Methods: Nine cervical cancer patients each received their brachytherapy preplanning PET/CT at the end of EBRT with a Syed template in place. The planning PET/CT was acquired on the day of brachytherapy treatment with the actual applicator (Syed or Tandem and Ring) and rigidly registered. The PET/CT images were then deformably registered creating a third (deformed) image set for target prediction. Regions of interest with standardized uptake values (SUV) greater than 65% of maximum SUV were contoured as target volumes in all three sets of PET images. The predictive value of the registered images was evaluated by comparing the preplanning and deformed PET volumes with the planning PET volume using Dice's coefficient (DC) and center-of-mass (COM) displacement. Results: The average DCs were 0.12±0.14 and 0.19±0.16 for rigid and deformable predicted target volumes, respectively. The average COM displacements were 1.9±0.9 cm and 1.7±0.7 cm for rigid and deformable registration, respectively. The DCs were improved by deformable registration, however, both were lower than published data for DIR in other modalities and clinical sites. Anatomical changes caused by different brachytherapy applicators could have posed a challenge to the DIR algorithm. The physiological change from interstitial needle placement may also contribute to lower DC. Conclusion: The clinical use of DIR in PET/CT for cervical cancer brachytherapy appears to be limited by applicator choice and requires further

  11. PET/CT: fundamental principles.

    PubMed

    Seemann, Marcus D

    2004-05-28

    Positron emission tomography (PET) facilitates the evaluation of metabolic and molecular characteristics of a wide variety of cancers, but is limited in its ability to visualize anatomical structures. Computed tomography (CT) facilitates the evaluation of anatomical structures of cancers, but can not visualize their metabolic and molecular aspects. Therefore, the combination of PET and CT provides the ability to accurately register metabolic and molecular aspects of disease with anatomical findings, adding further information to the diagnosis and staging of tumors. The recent generation of high performance PET/CT scanners combines a state of the art full-ring 3D PET scanner and a high-end 16-slice CT scanner. In PET/CT scanners, a CT examination is used for attenuation correction of PET images rather than standard transmission scanning using superset 68 Ge sources. This reduces the examination time, but metallic objects and contrast agents that alter the CT image quality and quantitative measurements of standardized uptake values (SUV) may lead to artifacts in the PET images. Hybrid PET/CT imaging will be very important in oncological applications in the decades to come, and possibly for use in cancer screening and cardiac imaging. PMID:15257877

  12. Get Set for a Pet.

    ERIC Educational Resources Information Center

    DeRosa, Bill

    1987-01-01

    Describes a game in which students deal with some of the factors involved in being a responsible pet owner. Includes a list of the materials needed for the game and provides the game board and the game pieces, along with a fold-out poster about neutering and spaying pets. (TW)

  13. Meet the Alpha-Pets.

    ERIC Educational Resources Information Center

    Zitlaw, Jo Ann Bruce; Frank, Cheryl Standish

    1985-01-01

    "Alpha-Pets" are the focal point of an integrated, multidisciplinary curriculum. Each pet is featured for a week in a vocabulary-rich story and introduces related activities beginning with the featured letter, such as the four food groups during Freddie Fish's week or universe during Ulysses Unicorn's week. (MT)

  14. Sequencing of the Escherichia coli O22 O Antigen Gene Cluster and Detection of E. coli Serogroups O22 and O91 by Multiplex PCR Assays Targeting Virulence Genes and the wzy Gene in the Respective O-Antigen Gene Clusters

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The O antigen gene cluster of Escherichia coli O22, an extraintestinal pathogenic E. coli (ExPEC) serogroup was sequenced, and eight open reading frames encoding O antigen sugar biosynthesis, transfer, and processing were identified. Multiplex PCR assays were developed targeting the wzx and wzy gen...

  15. 44Sc: An Attractive Isotope for Peptide-Based PET Imaging

    PubMed Central

    2015-01-01

    The overexpression of integrin αvβ3 has been linked to tumor aggressiveness and metastasis in several cancer types. Because of its high affinity, peptides containing the arginine–glycine–aspartic acid (RGD) motif have been proven valuable vectors for noninvasive imaging of integrin αvβ3 expression and for targeted radionuclide therapy. In this study, we aim to develop a 44Sc-labeled RGD-based peptide for in vivo positron emission tomography (PET) imaging of integrin αvβ3 expression in a preclinical cancer model. High quality 44Sc (t1/2, 3.97 h; β+ branching ratio, 94.3%) was produced inexpensively in a cyclotron, via proton irradiation of natural Ca metal targets, and separated by extraction chromatography. A dimeric cyclic-RGD peptide, (cRGD)2, was conjugated to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) and radiolabeled with 44Sc in high yield (>90%) and specific activity (7.4 MBq/nmol). Serial PET imaging of mice bearing U87MG tumor xenografts showed elevated 44Sc-DOTA-(cRGD)2 uptake in the tumor tissue of 3.93 ± 1.19, 3.07 ± 1.17, and 3.00 ± 1.25 %ID/g at 0.5, 2, and 4 h postinjection, respectively (n = 3), which were validated by ex vivo biodistribution experiments. The integrin αvβ3 specificity of the tracer was corroborated, both in vitro and in vivo, by competitive cell binding and receptor blocking assays. These results parallel previously reported studies showing similar tumor targeting and pharmacokinetic profiles for dimeric cRGD peptides labeled with 64Cu or 68Ga. Our findings, together with the advantageous radionuclidic properties of 44Sc, capitalize on the relevance of this isotope as an attractive alternative isotope to more established radiometals for small molecule-based PET imaging, and as imaging surrogate of 47Sc in theranostic applications. PMID:25054618

  16. Recent development in PET instrumentation.

    PubMed

    Peng, By Hao; Levin, Craig S

    2010-09-01

    Positron emission tomography (PET) is used in the clinic and in vivo small animal research to study molecular processes associated with diseases such as cancer, heart disease, and neurological disorders, and to guide the discovery and development of new treatments. This paper reviews current challenges of advancing PET technology and some of newly developed PET detectors and systems. The paper focuses on four aspects of PET instrumentation: high photon detection sensitivity; improved spatial resolution; depth-of-interaction (DOI) resolution and time-of-flight (TOF). Improved system geometry, novel non-scintillator based detectors, and tapered scintillation crystal arrays are able to enhance the photon detection sensitivity of a PET system. Several challenges for achieving high resolution with standard scintillator-based PET detectors are discussed. Novel detectors with 3-D positioning capability have great potential to be deployed in PET for achieving spatial resolution better than 1 mm, such as cadmium-zinc-telluride (CZT) and position-sensitive avalanche photodiodes (PSAPDs). DOI capability enables a PET system to mitigate parallax error and achieve uniform spatial resolution across the field-of-view (FOV). Six common DOI designs, as well as advantages and limitations of each design, are discussed. The availability of fast scintillation crystals such as LaBr(3), and the silicon photomultiplier (SiPM) greatly advances TOF-PET development. Recent instrumentation and initial results of clinical trials are briefly presented. If successful, these technology advances, together with new probe molecules, will substantially enhance the molecular sensitivity of PET and thus increase its role in preclinical and clinical research as well as evaluating and managing disease in the clinic. PMID:20497121

  17. [Interest of FDG-PET for lung cancer radiotherapy].

    PubMed

    Thureau, S; Mezzani-Saillard, S; Modzelewski, R; Edet-Sanson, A; Dubray, B; Vera, P

    2011-10-01

    The recent advances in medical imaging have profoundly altered the radiotherapy of non-small cell lung cancers (NSCLC). A meta-analysis has confirmed the superiority of FDG PET-CT over CT for initial staging. FDG PET-CT improves the reproducibility of target volume delineation, especially close to the mediastinum or in the presence of atelectasia. Although not formally validated by a randomized trial, the reduction of the mediastinal target volume, by restricting the irradiation to FDG-avid nodes, is widely accepted. The optimal method of delineation still remains to be defined. The role of FDG PET-CT in monitoring tumor response during radiotherapy is under investigation, potentially opening the way to adapting the treatment modalities to tumor radiation sensitivity. Other tracers, such as F-miso (hypoxia), are also under clinical investigation. To avoid excessive delays, the integration of PET-CT in routine practice requires quick access to the imaging equipment, technical support (fusion and image processing) and multidisciplinary delineation of target volumes. PMID:21880535

  18. Fluorescence polarization assays in signal transduction discovery.

    PubMed

    Sportsman, J Richard; Daijo, Janet; Gaudet, Elizabeth A

    2003-05-01

    Fluorescence polarization (FP) has become widely employed for high throughput screening used in pharmaceutical drug discovery. Assays of important signal transduction targets are now adapted to FP. In this review we examine assays for cyclic adenosine monophosphate, phosphodiesterases, and protein kinases and phosphatases using FP competitive immunoassays and a direct enzymatic method called IMAP. PMID:12678698

  19. 7 CFR 502.11 - Pets.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 6 2010-01-01 2010-01-01 false Pets. 502.11 Section 502.11 Agriculture Regulations of... CONDUCT ON BELTSVILLE AGRICULTURE RESEARCH CENTER PROPERTY, BELTSVILLE, MARYLAND § 502.11 Pets. Pets... vaccinations. Pets that are the property of employees residing on BARC must be up to date on their...

  20. 36 CFR 1002.15 - Pets.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 3 2011-07-01 2011-07-01 false Pets. 1002.15 Section 1002.15....15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public building, public... possession of pets by the Board. This paragraph shall not apply to guide dogs accompanying visually...

  1. 36 CFR 2.15 - Pets.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Pets. 2.15 Section 2.15 Parks... USE AND RECREATION § 2.15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public... area closed to the possession of pets by the superintendent. This subparagraph shall not apply to...

  2. 7 CFR 502.11 - Pets.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 6 2011-01-01 2011-01-01 false Pets. 502.11 Section 502.11 Agriculture Regulations of... CONDUCT ON BELTSVILLE AGRICULTURE RESEARCH CENTER PROPERTY, BELTSVILLE, MARYLAND § 502.11 Pets. Pets... vaccinations. Pets that are the property of employees residing on BARC must be up to date on their...

  3. 7 CFR 502.11 - Pets.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 6 2012-01-01 2012-01-01 false Pets. 502.11 Section 502.11 Agriculture Regulations of... CONDUCT ON BELTSVILLE AGRICULTURE RESEARCH CENTER PROPERTY, BELTSVILLE, MARYLAND § 502.11 Pets. Pets... vaccinations. Pets that are the property of employees residing on BARC must be up to date on their...

  4. 36 CFR 2.15 - Pets.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Pets. 2.15 Section 2.15 Parks... USE AND RECREATION § 2.15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public... area closed to the possession of pets by the superintendent. This subparagraph shall not apply to...

  5. 36 CFR 2.15 - Pets.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Pets. 2.15 Section 2.15 Parks... USE AND RECREATION § 2.15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public... area closed to the possession of pets by the superintendent. This subparagraph shall not apply to...

  6. 36 CFR 1002.15 - Pets.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 3 2013-07-01 2012-07-01 true Pets. 1002.15 Section 1002.15....15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public building, public... possession of pets by the Board. This paragraph shall not apply to guide dogs accompanying visually...

  7. 36 CFR 1002.15 - Pets.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 3 2012-07-01 2012-07-01 false Pets. 1002.15 Section 1002.15....15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public building, public... possession of pets by the Board. This paragraph shall not apply to guide dogs accompanying visually...

  8. 36 CFR 1002.15 - Pets.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 3 2014-07-01 2014-07-01 false Pets. 1002.15 Section 1002.15....15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public building, public... possession of pets by the Board. This paragraph shall not apply to guide dogs accompanying visually...

  9. 36 CFR 2.15 - Pets.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Pets. 2.15 Section 2.15 Parks... USE AND RECREATION § 2.15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public... area closed to the possession of pets by the superintendent. This subparagraph shall not apply to...

  10. 7 CFR 502.11 - Pets.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 6 2013-01-01 2013-01-01 false Pets. 502.11 Section 502.11 Agriculture Regulations of... CONDUCT ON BELTSVILLE AGRICULTURE RESEARCH CENTER PROPERTY, BELTSVILLE, MARYLAND § 502.11 Pets. Pets... vaccinations. Pets that are the property of employees residing on BARC must be up to date on their...

  11. 7 CFR 502.11 - Pets.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 6 2014-01-01 2014-01-01 false Pets. 502.11 Section 502.11 Agriculture Regulations of... CONDUCT ON BELTSVILLE AGRICULTURE RESEARCH CENTER PROPERTY, BELTSVILLE, MARYLAND § 502.11 Pets. Pets... vaccinations. Pets that are the property of employees residing on BARC must be up to date on their...

  12. Monte Carlo simulation of PET and SPECT imaging of {sup 90}Y

    SciTech Connect

    Takahashi, Akihiko Sasaki, Masayuki; Himuro, Kazuhiko; Yamashita, Yasuo; Komiya, Isao; Baba, Shingo

    2015-04-15

    Purpose: Yittrium-90 ({sup 90}Y) is traditionally thought of as a pure beta emitter, and is used in targeted radionuclide therapy, with imaging performed using bremsstrahlung single-photon emission computed tomography (SPECT). However, because {sup 90}Y also emits positrons through internal pair production with a very small branching ratio, positron emission tomography (PET) imaging is also available. Because of the insufficient image quality of {sup 90}Y bremsstrahlung SPECT, PET imaging has been suggested as an alternative. In this paper, the authors present the Monte Carlo-based simulation–reconstruction framework for {sup 90}Y to comprehensively analyze the PET and SPECT imaging techniques and to quantitatively consider the disadvantages associated with them. Methods: Our PET and SPECT simulation modules were developed using Monte Carlo simulation of Electrons and Photons (MCEP), developed by Dr. S. Uehara. PET code (MCEP-PET) generates a sinogram, and reconstructs the tomography image using a time-of-flight ordered subset expectation maximization (TOF-OSEM) algorithm with attenuation compensation. To evaluate MCEP-PET, simulated results of {sup 18}F PET imaging were compared with the experimental results. The results confirmed that MCEP-PET can simulate the experimental results very well. The SPECT code (MCEP-SPECT) models the collimator and NaI detector system, and generates the projection images and projection data. To save the computational time, the authors adopt the prerecorded {sup 90}Y bremsstrahlung photon data calculated by MCEP. The projection data are also reconstructed using the OSEM algorithm. The authors simulated PET and SPECT images of a water phantom containing six hot spheres filled with different concentrations of {sup 90}Y without background activity. The amount of activity was 163 MBq, with an acquisition time of 40 min. Results: The simulated {sup 90}Y-PET image accurately simulated the experimental results. PET image is visually

  13. Antibody-based PET imaging of amyloid beta in mouse models of Alzheimer's disease

    PubMed Central

    Sehlin, Dag; Fang, Xiaotian T.; Cato, Linda; Antoni, Gunnar; Lannfelt, Lars; Syvänen, Stina

    2016-01-01

    Owing to their specificity and high-affinity binding, monoclonal antibodies have potential as positron emission tomography (PET) radioligands and are currently used to image various targets in peripheral organs. However, in the central nervous system, antibody uptake is limited by the blood–brain barrier (BBB). Here we present a PET ligand to be used for diagnosis and evaluation of treatment effects in Alzheimer's disease. The amyloid β (Aβ) antibody mAb158 is radiolabelled and conjugated to a transferrin receptor antibody to enable receptor-mediated transcytosis across the BBB. PET imaging of two different mouse models with Aβ pathology clearly visualize Aβ in the brain. The PET signal increases with age and correlates closely with brain Aβ levels. Thus, we demonstrate that antibody-based PET ligands can be successfully used for brain imaging. PMID:26892305

  14. WE-G-BRF-06: Positron Emission Tomography (PET)-Guided Dynamic Lung Tumor Tracking for Cancer Radiotherapy: First Patient Simulations

    SciTech Connect

    Yang, J; Loo, B; Graves, E; Yamamoto, T; Keall, P

    2014-06-15

    Purpose: PET-guided dynamic tumor tracking is a novel concept of biologically targeted image guidance for radiotherapy. A dynamic tumor tracking algorithm based on list-mode PET data has been developed and previously tested on dynamic phantom data. In this study, we investigate if dynamic tumor tracking is clinically feasible by applying the method to lung cancer patient PET data. Methods: PET-guided tumor tracking estimates the target position of a segmented volume in PET images reconstructed continuously from accumulated coincidence events correlated with external respiratory motion, simulating real-time applications, i.e., only data up to the current time point is used to estimate the target position. A target volume is segmented with a 50% threshold, consistently, of the maximum intensity in the predetermined volume of interest. Through this algorithm, the PET-estimated trajectories are quantified from four lung cancer patients who have distinct tumor location and size. The accuracy of the PET-estimated trajectories is evaluated by comparing to external respiratory motion because the ground-truth of tumor motion is not known in patients; however, previous phantom studies demonstrated sub-2mm accuracy using clinically derived 3D tumor motion. Results: The overall similarity of motion patterns between the PET-estimated trajectories and the external respiratory traces implies that the PET-guided tracking algorithm can provide an acceptable level of targeting accuracy. However, there are variations in the tracking accuracy between tumors due to the quality of the segmentation which depends on target-to-background ratio, tumor location and size. Conclusion: For the first time, a dynamic tumor tracking algorithm has been applied to lung cancer patient PET data, demonstrating clinical feasibility of real-time tumor tracking for integrated PET-linacs. The target-to-background ratio is a significant factor determining accuracy: screening during treatment planning would

  15. SU-C-9A-06: The Impact of CT Image Used for Attenuation Correction in 4D-PET

    SciTech Connect

    Cui, Y; Bowsher, J; Yan, S; Cai, J; Das, S; Yin, F

    2014-06-01

    Purpose: To evaluate the appropriateness of using 3D non-gated CT image for attenuation correction (AC) in a 4D-PET (gated PET) imaging protocol used in radiotherapy treatment planning simulation. Methods: The 4D-PET imaging protocol in a Siemens PET/CT simulator (Biograph mCT, Siemens Medical Solutions, Hoffman Estates, IL) was evaluated. CIRS Dynamic Thorax Phantom (CIRS Inc., Norfolk, VA) with a moving glass sphere (8 mL) in the middle of its thorax portion was used in the experiments. The glass was filled with {sup 18}F-FDG and was in a longitudinal motion derived from a real patient breathing pattern. Varian RPM system (Varian Medical Systems, Palo Alto, CA) was used for respiratory gating. Both phase-gating and amplitude-gating methods were tested. The clinical imaging protocol was modified to use three different CT images for AC in 4D-PET reconstruction: first is to use a single-phase CT image to mimic actual clinical protocol (single-CT-PET); second is to use the average intensity projection CT (AveIP-CT) derived from 4D-CT scanning (AveIP-CT-PET); third is to use 4D-CT image to do the phase-matched AC (phase-matching- PET). Maximum SUV (SUVmax) and volume of the moving target (glass sphere) with threshold of 40% SUVmax were calculated for comparison between 4D-PET images derived with different AC methods. Results: The SUVmax varied 7.3%±6.9% over the breathing cycle in single-CT-PET, compared to 2.5%±2.8% in AveIP-CT-PET and 1.3%±1.2% in phasematching PET. The SUVmax in single-CT-PET differed by up to 15% from those in phase-matching-PET. The target volumes measured from single- CT-PET images also presented variations up to 10% among different phases of 4D PET in both phase-gating and amplitude-gating experiments. Conclusion: Attenuation correction using non-gated CT in 4D-PET imaging is not optimal process for quantitative analysis. Clinical 4D-PET imaging protocols should consider phase-matched 4D-CT image if available to achieve better accuracy.

  16. Diseases Transmitted by Domestic Livestock: Perils of the Petting Zoo.

    PubMed

    Dunn, John R; Behravesh, Casey Barton; Angulo, Frederick J

    2015-12-01

    Petting zoo venues encourage or permit public contact with animals which provide opportunities for education and entertainment. These venues vary but are common at county or state fairs, zoos, and aquariums. In addition to these common petting zoo settings, animals are present in many other venues where the public is permitted to contact them and their environment. Thus, humans may have contact with animals in a wide range of settings, and transmission of infectious diseases from animals to humans may occur at any of these venues, creating perils associated with petting zoos.There are many considerations when evaluating perils associated with the wide range of venues where animal contact can occur. First, many venues or events draw large numbers of people; some operate during a short time frame, while others, such as zoos and aquariums, operate year round. Second, petting zoos and other animal contact venues are particularly popular with children, who compared with adults, commonly have less stringent hygienic practices and are more susceptible to severe disease outcomes. Finally, there is remarkable variability in the physical layout of venues that permit animal contact and in the types of animals that may be contacted. Animal contact areas range from well-designed permanent exhibits targeting risk reduction to various temporary or seasonal exhibits established without detailed planning. Many petting zoos house only small ruminant species such as sheep and goats, but other venues house a wide variety of mammalian species, exotic animals, poultry and other avian species, reptiles and amphibians, and aquatic animals. PMID:27337283

  17. Understanding regulations affecting pet foods.

    PubMed

    Dzanis, David A

    2008-08-01

    In the United States, pet foods are subject to regulation at both the federal and the state levels. The US Food and Drug Administration has jurisdiction over all animal feeds (including pet foods, treats, chews, supplements, and ingredients) in interstate commerce, which includes imported products. Many states adopt and enforce at least in part the Association of American Feed Control Officials Model Bill and Model Regulations for Pet Food and Specialty Pet Food. Thus, all pet foods in multi-state distribution are subject to a host of labeling requirements covering aspects such as product names, ingredient lists, nutrient content guarantees, and nutritional adequacy statements. Ingredients must be GRAS (generally recognized as safe) substances, approved food additives, or defined by Association of American Feed Control Officials for their intended use. Pet food labels may not bear claims that are false or misleading or that state or imply use for the treatment or prevention of disease. Pet foods that are found to be adulterated or misbranded may be subject to seizure or other enforcement actions. PMID:18656837

  18. PET/CT for Radiotherapy Treatment Planning in Patients With Soft Tissue Sarcomas

    SciTech Connect

    Karam, Irene; Devic, Slobodan; Hickeson, Marc; Roberge, David; Turcotte, Robert E.; Freeman, Carolyn R.

    2009-11-01

    Purpose: To study the possibility of incorporating positron emission tomography/computed tomography (PET/CT) information into radiotherapy treatment planning in patients with high-grade soft tissue sarcomas (STS). Methods and Materials: We studied 17 patients treated with preoperative radiotherapy at our institution from 2005 to 2007. All patients had a high-grade STS and had had a staging PET/CT scan. For each patient, an MRI-based gross tumor volume (GTV), considered to be the contemporary standard for radiotherapy treatment planning, was outlined on a T1-gadolinium enhanced axial MRI (GTV{sub MRI}), and a second set of GTVs were outlined using different threshold values on PET images (GTV{sub PET}). PET-based target volumes were compared with the MRI-based GTV. Threshold values for target contouring were determined as a multiple (from 2 to 10 times) of the background soft tissue uptake values (B) sampled over healthy tissue. Results: PET-based GTVs contoured using a threshold value of 2 or 2.5 most closely resembled the GTV{sub MRI} volumes. Higher threshold values lead to PET volumes much smaller than the GTV{sub MRI}. The standard deviations between the average volumes of GTV{sub PET} and GTV{sub MRI} ratios for all thresholds were large, ranging from 36% for 2 xB up to 93% for 10 xB. Maximum uptake-to-background ratio correlated poorly with the maximum standardized uptake values. Conclusions: It is unlikely that PET/CT will make a significant contribution in GTV definition for radiotherapy treatment planning in patients with STS using threshold methods on PET images. Future studies will focus on molecular imaging and tumor physiology.

  19. Preliminary results of a prototype C-shaped PET designed for an in-beam PET system

    NASA Astrophysics Data System (ADS)

    Kim, Hyun-Il; Chung, Yong Hyun; Lee, Kisung; Kim, Kyeong Min; Kim, Yongkwon; Joung, Jinhun

    2016-06-01

    Positron emission tomography (PET) can be utilized in particle beam therapy to verify the dose distribution of the target volume as well as the accuracy of the treatment. We present an in-beam PET scanner that can be integrated into a particle beam therapy system. The proposed PET scanner consisted of 14 detector modules arranged in a C-shape to avoid blockage of the particle beam line by the detector modules. Each detector module was composed of a 9×9 array of 4.0 mm×4.0 mm×20.0 mm LYSO crystals optically coupled to four 29-mm-diameter PMTs using the photomultiplier-quadrant-sharing (PQS) technique. In this study, a Geant4 Application for Tomographic Emission (GATE) simulation study was conducted to design a C-shaped PET scanner and then experimental evaluation of the proposed design was performed. The spatial resolution and sensitivity were measured according to NEMA NU2-2007 standards and were 6.1 mm and 5.61 cps/kBq, respectively, which is in good agreement with our simulation, with an error rate of 12.0%. Taken together, our results demonstrate the feasibility of the proposed C-shaped in-beam PET system, which we expect will be useful for measuring dose distribution in particle therapy.

  20. Longitudinal monitoring adipose-derived stem cell survival by PET imaging hexadecyl-4-{sup 124}I-iodobenzoate in rat myocardial infarction model

    SciTech Connect

    Kim, Min Hwan; Woo, Sang-Keun; Lee, Kyo Chul; An, Gwang Il; Pandya, Darpan; Park, Noh Won; Nahm, Sang-Soep; Eom, Ki Dong; Kim, Kwang Il; Lee, Tae Sup; Kim, Chan Wha; Kang, Joo Hyun; Yoo, Jeongsoo; Lee, Yong Jin

    2015-01-02

    Highlights: • We developed a safe, simple and appropriate stem cell labeling method with {sup 124}I-HIB. • ADSC survival can be monitored with PET in MI model via direct labeling. • Tracking of ADSC labeled with {sup 124}I-HIB was possible for 3 days in MI model using PET. • ADSC viability and differentiation were not affected by {sup 124}I-HIB labeling. • Survival of ADSC in living bodies can be longitudinally tracked with PET imaging. - Abstract: This study aims to monitor how the change of cell survival of transplanted adipose-derived stem cells (ADSCs) responds to myocardial infarction (MI) via the hexadecyl-4-{sup 124}I-iodobenzoate ({sup 124}I-HIB) mediated direct labeling method in vivo. Stem cells have shown the potential to improve cardiac function after MI. However, monitoring of the fate of transplanted stem cells at target sites is still unclear. Rat ADSCs were labeled with {sup 124}I-HIB, and radiolabeled ADSCs were transplanted into the myocardium of normal and MI model. In the group of {sup 124}I-HIB-labeled ADSC transplantation, in vivo imaging was performed using small-animal positron emission tomography (PET)/computed tomography (CT) for 9 days. Twenty-one days post-transplantation, histopathological analysis and apoptosis assay were performed. ADSC viability and differentiation were not affected by {sup 124}I-HIB labeling. In vivo tracking of the {sup 124}I-HIB-labeled ADSCs was possible for 9 and 3 days in normal and MI model, respectively. Apoptosis of transplanted cells increased in the MI model compared than that in normal model. We developed a direct labeling agent, {sup 124}I-HIB, and first tried to longitudinally monitor transplanted stem cell to MI. This approach may provide new insights on the roles of stem cell monitoring in living bodies for stem cell therapy from pre-clinical studies to clinical trials.

  1. Imatinib Analogs as Potential Agents for PET Imaging of Bcr-Abl/c-KIT Expression at a Kinase Level

    PubMed Central

    Peng, Zhenghong; Maxwell, David S.; Sun, Duoli; Bhanu Prasad, Basvoju A.; Pal, Ashutosh; Wang, Shimei; Balatoni, Julius; Ghosh, Pradip; Lim, Seok T.; Volgin, Andrei; Shavrin, Aleksander; Alauddin, Mian M.; Gelovani, Juri G.; Bornmann, William G.

    2014-01-01

    We synthesized two series of imatinib mesylate (STI-571) analogs to develop a Bcr-Abl and c-KIT receptor-specific labeling agent for positron emission tomography (PET) imaging to measure Bcr-Abl and c-KIT expression levels in a mouse model. The methods of molecular modeling, synthesis of STI-571 and its analogs, in vitro kinase assays, and radiolabeling are described. Molecular modeling revealed that these analogs bind the same Bcr-Abl and c-KIT binding sites as those bound by STI-571. The analogs potently inhibit the tyrosine kinase activity of Bcr-Abl and c-KIT, similarly to STI-571. [18F]-labeled STI-571 was prepared with high specific activity (75 GBq/μmol) by nucleophilic displacement and an average radiochemical yield of 12%. [131I]-labeled STI-571 was prepared with high purity (>95%) and an average radiochemical yield of 23%. The uptake rates of [18F]-STI-571 in K562 cells expressing Abl and in U87WT cells overexpressing c-KIT were significantly higher than those in the U87 cell and could be inhibited by STI-71 (confirming the specificity of uptake). PET scans of K562 and U87WT tumor-bearing mice with [18F]-STI-571 as a contrast agent showed visible tumor uptake and tumor-to-non-target contrast. PMID:24280068

  2. Evaluation and use of pet foods: general considerations in using pet foods for adult maintenance.

    PubMed

    Kallfelz, F A

    1989-05-01

    Questions regarding pet animal nutrition are probably among the most frequent queries encountered by companion animal veterinarians. Given the plethora of pet food products available and the amount of advertising used to promote them, it is not surprising that pet owners have concerns as to what they should feed their pets. This "practical" review of pet foods and feeding is designed to assist veterinarians in making nutritional recommendations to their clients, with respect to feeding normal adult pets at maintenance. PMID:2658281

  3. Veterinarians' role for pet owners facing pet loss

    PubMed Central

    Fernandez-Mehler, P.; Gloor, P.; Sager, E.; Lewis, F. I.; Glaus, T. M

    2013-01-01

    Owners' satisfaction with, and expectations from, their veterinarians around euthanasia, including questions on disposal of pet remains subject to animal species, clients' gender, age, family conditions, area of living and type of veterinary clinic visited were evaluated by questionnaire. Questionnaires were to be filled out by clients consecutively visiting the individual practices and hospitals for any kind of consultations. Of 2350 questionnaires distributed, 2008 were returned and available for analysis. Owner satisfaction concerning the procedure of euthanasia was high (92 per cent, 1173/1272). After the event of euthanasia, 14 per cent (170/1250) had changed their veterinarian, even though 75 per cent of these 170 had been satisfied with the procedure. Most owners (88 per cent) expected veterinarians to talk about their pet's final destination, and 38 per cent expected this to happen early in the pet's life. For 81 per cent clients, the veterinarian was the primary informant about the possibilities concerning the disposal of pet remains, and 33 per cent indicated their veterinarian as the contact person to talk about pet loss. Area of living, or veterinary specialisation, only marginally influenced the answers. Veterinarians play an important role to inform their clients concerning questions around euthanasia and the care of pet remains, and to support them during the process of mourning. PMID:23492929

  4. Preclinical Properties of 18F-AV-45: A PET Agent for Aβ Plaques in the Brain

    PubMed Central

    Choi, Seok Rye; Golding, Geoff; Zhuang, Zhiping; Zhang, Wei; Lim, Nathaniel; Hefti, Franz; Benedum, Tyler E.; Kilbourn, Michael R.; Skovronsky, Daniel; Kung, Hank F.

    2011-01-01

    β-amyloid plaques (Aβ plaques) in the brain, containing predominantly fibrillary Aβ peptide aggregates, represent a defining pathologic feature of Alzheimer disease (AD). Imaging agents targeting the Aβ plaques in the living human brain are potentially valuable as biomarkers of pathogenesis processes in AD. (E)-4-(2-(6-(2-(2-(2-18F-fluoroethoxy)ethoxy)ethoxy)pyridin-3-yl)vinyl)-N-methyl benzenamine (18F-AV-45) is such as an agent currently in phase III clinical studies for PET of Aβ plaques in the brain. Methods In vitro binding of 18F-AV-45 to Aβ plaques in the postmortem AD brain tissue was evaluated by in vitro binding assay and autoradiography. In vivo biodistribution of 18F-AV-45 in mice and ex vivo autoradiography of AD transgenic mice (APPswe/PSEN1) with Aβ aggregates in the brain were performed. Small-animal PET of a monkey brain after an intravenous injection of 18F-AV-45 was evaluated. Results 18F-AV-45 displayed a high binding affinity and specificity to Aβ plaques (Kd, 3.72 ± 0.30 nM). In vitro autoradiography of postmortem human brain sections showed substantial plaque labeling in AD brains and not in the control brains. Initial high brain uptake and rapid washout from the brain of healthy mice and monkey were observed. Metabolites produced in the blood of healthy mice after an intravenous injection were identified. 18F-AV-45 displayed excellent binding affinity to Aβ plaques in the AD brain by ex vivo autoradiography in transgenic AD model mice. The results lend support that 18F-AV-45 may be a useful PET agent for detecting Aβ plaques in the living human brain. PMID:19837759

  5. PET Pharmacokinetic Modelling

    NASA Astrophysics Data System (ADS)

    Müller-Schauenburg, Wolfgang; Reimold, Matthias

    Positron Emission Tomography is a well-established technique that allows imaging and quantification of tissue properties in-vivo. The goal of pharmacokinetic modelling is to estimate physiological parameters, e.g. perfusion or receptor density from the measured time course of a radiotracer. After a brief overview of clinical application of PET, we summarize the fundamentals of modelling: distribution volume, Fick's principle of local balancing, extraction and perfusion, and how to calculate equilibrium data from measurements after bolus injection. Three fundamental models are considered: (i) the 1-tissue compartment model, e.g. for regional cerebral blood flow (rCBF) with the short-lived tracer [15O]water, (ii) the 2-tissue compartment model accounting for trapping (one exponential + constant), e.g. for glucose metabolism with [18F]FDG, (iii) the reversible 2-tissue compartment model (two exponentials), e.g. for receptor binding. Arterial blood sampling is required for classical PET modelling, but can often be avoided by comparing regions with specific binding with so called reference regions with negligible specific uptake, e.g. in receptor imaging. To estimate the model parameters, non-linear least square fits are the standard. Various linearizations have been proposed for rapid parameter estimation, e.g. on a pixel-by-pixel basis, for the prize of a bias. Such linear approaches exist for all three models; e.g. the PATLAK-plot for trapping substances like FDG, and the LOGAN-plot to obtain distribution volumes for reversibly binding tracers. The description of receptor modelling is dedicated to the approaches of the subsequent lecture (chapter) of Millet, who works in the tradition of Delforge with multiple-injection investigations.

  6. PET radiopharmaceuticals for probing enzymes in the brain

    PubMed Central

    Holland, Jason P; Cumming, Paul; Vasdev, Neil

    2013-01-01

    Biologically important processes in normal brain function and brain disease involve the action of various protein-based receptors, ion channels, transporters and enzymes. The ability to interrogate the location, abundance and activity of these entities in vivo using non-invasive molecular imaging can provide unprecedented information about the spatio-temporal dynamics of brain function. Indeed, positron emission tomography (PET) imaging is transforming our understanding of the central nervous system and brain disease. Great emphasis has historically been placed on developing radioligands for the non-invasive detection of neuroreceptors. In contrast, relatively few enzymes have been amenable to examination by PET imaging procedures based upon trapping or accumulation of enzymatic products, because only a subset of enzymes have sufficient catalytic rate to produce measureable accumulation within the practical time-limit of PET recordings. However, high affinity inhibitors are now serving as tracers for enzymes, particularly for measuring the abundance of enzymes mediating intracellular signal transduction in the brain, which offer a rich diversity of potential targets for drug discovery. The purpose of this review is to summarize well-known radiotracers for brain enzymes, and draw attention to recent developments in PET radiotracers for imaging signal transduction pathways in the brain. The review is organized by target class and focuses on structural chemistry of the best-established radiotracers identified in each class. PMID:23638333

  7. Should Immunocompromised Patients Have Pets?

    PubMed Central

    Steele, Russell W.

    2008-01-01

    Purpose: To evaluate the risks and benefits of pet ownership by immunodeficient patients, focusing primarily on organisms that colonize animals and are transmitted to humans. Those diseases that are known to be progressive or more severe in patients with altered immune function are emphasized. Methods: A review of the medical and veterinary literature pertaining to zoonoses transmitted by domestic animals was completed. Information pertaining to issues involving immunosuppressed patients including AIDS was carefully evaluated and summarized for inclusion. Results: There are significant clinical and psychosocial benefits to pet ownership. However, numerous diseases can be acquired from these animals which may be more severe in immunocompromised individuals. Conclusion: Simple guidelines for pet ownership by immunosuppressed patients can be implemented to reduce their risk of disease and allow them to safely interchange with their pets. PMID:21603465

  8. Pets and the immunocompromised person

    MedlinePlus

    ... Do not adopt wild or exotic animals. These animals are more likely to bite. They often carry rare but ... its feces because salmonella is easily passed from animal to human. Wear ... on pet-related infections, contact your veterinarian ...

  9. Take Care with Pet Reptiles

    MedlinePlus

    ... pets in households with young children. [775 KB] Animals and Health Check out two CDC websites with helpful resources. Gastrointestinal (Enteric) Diseases from Animals : Information about zoonotic outbreaks, prevention messages, and helpful ...

  10. 10 "Poison Pills" for Pets

    MedlinePlus

    ... left on the bedside table. Zolpidem may make cats wobbly and sleepy, but most pets become very ... very common pain killer found in most households. Cats are extremely sensitive to acetaminophen, but dogs can ...

  11. Initial report of PET/CT-guided radiofrequency ablation of liver metastases.

    PubMed

    Prior, John O; Kosinski, Marek; Delaloye, Angelika Bischof; Denys, Alban

    2007-06-01

    Computed tomography (CT) and ultrasonography (US) are commonly employed to guide positioning of radiofrequency electrodes within target tumors. However, this technique cannot be used when the tumor is detectable only by positron emission tomography (PET). In such cases, even the use of intraprocedural coregistered PET/CT will not prevent malpositioning of the electrode tip relative to a lesion visualized only on PET as a result of patient breathing and organ shifts during CT-guided electrode placement. The present report describes a single case of successful targeting and complete ablation of a lesion invisible on CT and US with the use of a method to visualize electrode tip positioning by PET. PMID:17538147

  12. 18F-Fluoroestradiol PET: Current Status and Potential Future Clinical Applications.

    PubMed

    Liao, Geraldine J; Clark, Amy S; Schubert, Erin K; Mankoff, David A

    2016-08-01

    Estrogen receptor (ER) expression in breast cancer is associated with a more favorable prognosis and is necessary for a response to endocrine therapies. Traditionally, ER expression is assessed by in vitro assays on biopsied tumor tissue. However, recent advances have allowed in vivo evaluation of ER expression with (18)F-fluoroestradiol ((18)F-FES) PET. Clinical studies have demonstrated the use of (18)F-FES PET as a method for quantifying in vivo ER expression and have explored its potential as a predictive assay and method of assessing in vivo pharmacodynamic response to endocrine therapy. This review outlines the biology and pharmacokinetics of (18)F-FES, highlights the current experience with (18)F-FES in patient studies on breast cancer and other diseases, and discusses potential clinical applications and the possible future clinical use of (18)F-FES PET. PMID:27307345

  13. Should pet owners be regulated?

    PubMed

    Mills, Georgina

    2013-12-21

    To own a television, you have to have a licence, and to drive a car, you have to pass a test. However, there are no such limitations on owning a pet. Should this be changed, and what can be done to encourage more responsible pet ownership? This topic was discussed at the BVA Congress at the London Vet Show on November 21. Georgina Mills reports. PMID:24362802

  14. Advances in Clinical PET/MRI Instrumentation.

    PubMed

    Herzog, Hans; Lerche, Christoph

    2016-04-01

    In 2010, the first whole-body PET/MRI scanners installed for clinical use were the sequential Philips PET/MRI with PMT-based, TOF-capable technology and the integrated simultaneous Siemens PET/MRI. Avalanche photodiodes as non-magneto-sensitive readout electronics allowed PET integrated within the MRI. The experiences with these scanners showed that improvements of software aspects, such as attenuation correction, were necessary and that efficient protocols combining optimally PET and MRI must be still developed. In 2014, General Electric issued an integrated PET/MRI with SiPM-based PET detectors, allowing TOF-PET. Looking at the MRI components of current PET/MR imaging systems, primary improvements come from sequences and new coils. PMID:26952724

  15. Nutritional sustainability of pet foods.

    PubMed

    Swanson, Kelly S; Carter, Rebecca A; Yount, Tracy P; Aretz, Jan; Buff, Preston R

    2013-03-01

    Sustainable practices meet the needs of the present without compromising the ability of future generations to meet their needs. Applying these concepts to food and feed production, nutritional sustainability is the ability of a food system to provide sufficient energy and essential nutrients required to maintain good health in a population without compromising the ability of future generations to meet their nutritional needs. Ecological, social, and economic aspects must be balanced to support the sustainability of the overall food system. The nutritional sustainability of a food system can be influenced by several factors, including the ingredient selection, nutrient composition, digestibility, and consumption rates of a diet. Carbon and water footprints vary greatly among plant- and animal-based ingredients, production strategy, and geographical location. Because the pet food industry is based largely on by-products and is tightly interlinked with livestock production and the human food system, however, it is quite unique with regard to sustainability. Often based on consumer demand rather than nutritional requirements, many commercial pet foods are formulated to provide nutrients in excess of current minimum recommendations, use ingredients that compete directly with the human food system, or are overconsumed by pets, resulting in food wastage and obesity. Pet food professionals have the opportunity to address these challenges and influence the sustainability of pet ownership through product design, manufacturing processes, public education, and policy change. A coordinated effort across the industry that includes ingredient buyers, formulators, and nutritionists may result in a more sustainable pet food system. PMID:23493530

  16. PET and PET/CT with 68Gallium-Labeled Somatostatin Analogues in Non GEP-NETs Tumors

    PubMed Central

    Erba, Paola Anna; Fraternali, Alessandro; Casali, Massimiliano; Di Paolo, Maria Liberata; Froio, Armando; Frasoldati, Andrea; Versari, Annibale

    2014-01-01

    Somatostatin (SST) is a 28-amino-acid cyclic neuropeptide mainly secreted by neurons and endocrine cells. A major interest for SST receptors (SSTR) as target for in vivo diagnostic and therapeutic purposes was born since a series of stable synthetic SST-analouges PET became available, being the native somatostatin non feasible for clinical use due to the very low metabolic stability. The rationale for the employment of SST-analogues to image cancer is both based on the expression of SSTR by tumor and on the high affinity of these compounds for SSTR. The primary indication of SST-analogues imaging is for neuroendocrine tumors (NETs), which usually express a high density of SSTR, so they can be effectively targeted and visualized with radiolabeled SST-analogues in vivo. Particularly, SST-analogues imaging has been widely employed in gastroenteropancreatic (GEP) NETs. Nevertheless, a variety of tumors other than NETs expresses SSTR thus SST-analogues imaging can also be used in these tumors, particularly if treatment with radiolabeled therapeutic SST-analouges PET is being considered. The aim of this paper is to provide a concise overview of the role of positron emission tomography/computed tomography (PET/CT) with 68Ga-radiolabeled SST-analouges PET in tumors other than GEP-NETs. PMID:24693229

  17. Assay development status report for total cyanide

    SciTech Connect

    Simpson, B.C.; Jones, T.E.; Pool, K.H.

    1993-02-01

    A validated cyanide assay that is applicable to a variety of tank waste matrices is necessary to resolve certain waste tank safety issues and for purposes of overall waste characterization. The target for this effort is an assay with an applicable range of greater than 1,000 ppM (0.10 wt%) total cyanide and a confidence level greater than 80%. Figure 1 illustrates the operating regime of the proposed cyanide assay method. The Assay Development Status Report for Total Cyanide will summarize the past experience with cyanide analyses on-tank waste matrices and will rate the status of the analytical methods used to assay total cyanide (CN{sup {minus}} ion) in the tank waste matrices as acceptable or unacceptable. This paper will also briefly describe the current efforts for improving analytical resolution of the assays and the attempts at speciation.

  18. PET/MRI in the infarcted mouse heart with the Cambridge split magnet

    NASA Astrophysics Data System (ADS)

    Buonincontri, Guido; Sawiak, Stephen J.; Methner, Carmen; Krieg, Thomas; Hawkes, Robert C.; Adrian Carpenter, T.

    2013-02-01

    Chronic heart failure, as a result of acute myocardial infarction, is a leading cause of death worldwide. Combining diagnostic imaging modalities may aid the direct assessment of experimental treatments targeting heart failure in vivo. Here we present preliminary data using the Cambridge combined PET/MRI imaging system in a mouse model of acute myocardial infarction. The split-magnet design can deliver uncompromised MRI and PET performance, for better assessment of disease and treatment in a preclinical environment.

  19. Impact of MR based attenuation correction on neurological PET studies

    PubMed Central

    Su, Yi; Rubin, Brian B.; McConathy, Jonathan; Laforest, Richard; Qi, Jing; Sharma, Akash; Priatna, Agus; Benzinger, Tammie L.S.

    2016-01-01

    Hybrid positron emission tomography (PET) and magnetic resonance (MR) scanners have become a reality in recent years with the benefits of reduced radiation exposure, reduction of imaging time, and potential advantages in quantification. Appropriate attenuation correction remains a challenge. Biases in PET activity measurements were demonstrated using the current MR based attenuation correction technique. We aim to investigate the impact of using standard MRAC technique on the clinical and research utility of PET/MR hybrid scanner for amyloid imaging. Methods Florbetapir scans were obtained on 40 participants on a Biograph mMR hybrid scanner with simultaneous MR acquisition. PET images were reconstructed using both MR and CT derived attenuation map. Quantitative analysis was performed for both datasets to assess the impact of MR based attenuation correction to absolute PET activity measurements as well as target to reference ratio (SUVR). Clinical assessment was also performed by a nuclear medicine physician to determine amyloid status based on the criteria in the FDA prescribing information for florbetapir. Results MR based attenuation correction led to underestimation of PET activity for most part of the brain with a small overestimation for deep brain regions. There is also an overestimation of SUVR values with cerebellar reference. SUVR measurements obtained from the two attenuation correction methods were strongly correlated. Clinical assessment of amyloid status resulted in identical classification as positive or negative regardless of the attenuation correction methods. Conclusions MR based attenuation correction cause biases in quantitative measurements. The biases may be accounted for by a linear model, although the spatial variation cannot be easily modelled. The quantitative differences however did not affect clinical assessment as positive or negative. PMID:26823562

  20. Fluorine-18 labeled tracers for PET studies in the neurosciences

    SciTech Connect

    Ding, Yu-Shin; Fowler, J.S.

    1995-12-31

    This chapter focuses on fluorine-18, the positron emitter with the longest half-life, the lowest positron energy and probably, the most challenging chemistry. The incorporation of F-18 into organic compounds presents many challenges, including: the need to synthesize and purify the compound within a 2--3 hour time frame; the limited number of labeled precursor molecules; the need to work on a microscale; and the need to produce radiotracers which are chemically and radiochemically pure, sterile and pyrogen-free, and suitable for intravenous injection. The PET method and F-18 labeling of organic molecules are described followed by highlights of the applications of F-18 labeled compounds in the neurosciences and neuropharmacology. It is important to emphasize the essential and pivotal role that organic synthesis has played in the progression of the PET field over the past twenty years from one in which only a handful of institutions possessed the instrumentation and staff to carry out research to the present-day situation where there are more than 200 PET centers worldwide. During this period PET has become an important scientific tool in the neurosciences, cardiology and oncology. It is important to point out that PET is by no means a mature field. The fact that a hundreds of different F-18 labeled compounds have been developed but only a few possess the necessary selectivity and sensitivity in vivo to track a specific biochemical process illustrates this and underscores a major difficulty in radiotracer development, namely the selection of priority structures for synthesis and the complexities of the interactions between chemical compounds and living systems. New developments in rapid organic synthesis are needed in order to investigate new molecular targets and to improve the quantitative nature of PET experiments.

  1. Respiratory trace feature analysis for the prediction of respiratory-gated PET quantification

    NASA Astrophysics Data System (ADS)

    Wang, Shouyi; Bowen, Stephen R.; Chaovalitwongse, W. Art; Sandison, George A.; Grabowski, Thomas J.; Kinahan, Paul E.

    2014-02-01

    when clinicians quantitatively assess PET/CT for therapy target definition and response assessment.

  2. Respiratory trace feature analysis for the prediction of respiratory-gated PET quantification.

    PubMed

    Wang, Shouyi; Bowen, Stephen R; Chaovalitwongse, W Art; Sandison, George A; Grabowski, Thomas J; Kinahan, Paul E

    2014-02-21

    compensation when clinicians quantitatively assess PET/CT for therapy target definition and response assessment. PMID:24504153

  3. Implementation of a solid target production facility

    NASA Astrophysics Data System (ADS)

    Tochon-Danguy, H. J.; Poniger, S. S.; Sachinidis, J. I.; Panopoulos, H. P.; Scott, A. M.

    2012-12-01

    The desire to utilize long-lived PET isotopes in Australia has significantly increased over the years and several research projects for labelling of peptides, proteins and biomolecules, including labelling of recombinant antibodies has been restricted due to the limited availability of suitable isotopes. This need has led to the recent installation and commissioning of a new facility dedicated to fully automated solid target isotope production, including 24I, 64Cu, 89Zr and 86Y at the Austin Health Centre for PET.

  4. Parasites in pet reptiles

    PubMed Central

    2011-01-01

    Exotic reptiles originating from the wild can be carriers of many different pathogens and some of them can infect humans. Reptiles imported into Slovenia from 2000 to 2005, specimens of native species taken from the wild and captive bred species were investigated. A total of 949 reptiles (55 snakes, 331 lizards and 563 turtles), belonging to 68 different species, were examined for the presence of endoparasites and ectoparasites. Twelve different groups (Nematoda (5), Trematoda (1), Acanthocephala (1), Pentastomida (1) and Protozoa (4)) of endoparasites were determined in 26 (47.3%) of 55 examined snakes. In snakes two different species of ectoparasites were also found. Among the tested lizards eighteen different groups (Nematoda (8), Cestoda (1), Trematoda (1), Acanthocephala (1), Pentastomida (1) and Protozoa (6)) of endoparasites in 252 (76.1%) of 331 examined animals were found. One Trombiculid ectoparasite was determined. In 563 of examined turtles eight different groups (Nematoda (4), Cestoda (1), Trematoda (1) and Protozoa (2)) of endoparasites were determined in 498 (88.5%) animals. In examined turtles three different species of ectoparasites were seen. The established prevalence of various parasites in reptiles used as pet animals indicates the need for examination on specific pathogens prior to introduction to owners. PMID:21624124

  5. Imaging Caspase-3 Activation as a Marker of Apoptosis-Targeted Treatment Response in Cancer

    PubMed Central

    Chen, Delphine L.; Engle, Jacquelyn T.; Griffin, Elizabeth A.; Miller, J. Philip; Chu, Wenhua; Zhou, Dong; Mach, Robert H.

    2016-01-01

    Purpose We tested whether positron emission tomography (PET) with the caspase-3 targeted isatin analog [18F]WC-4-116 could image caspase-3 activation in response to an apoptosis-inducing anticancer therapy. Procedures [18F]WC-4-116 uptake was determined in etoposide-treated EL4 cells. Biodistribution studies with [18F]WC-4-116 and [18F]ICMT-18, a non-caspase-3-targeted tracer, as well as [18F]WC-4-116 microPET imaging assessed responses in Colo205 tumor bearing mice treated with death receptor 5 (DR5) targeted agonist antibodies. Immunohistochemical staining and enzyme assays confirmed caspase-3 activation. Two-way analysis of variance or Student’s t-test assessed for treatment-related changes in tracer uptake. Results [18F]WC-4-116 increased 8 ± 2-fold in etoposide-treated cells. The [18F]WC-4-116 %ID/g also increased significantly in tumors with high caspase-3 enzyme activity (p < 0.05). [18F]ICMT-18 tumor uptake did not differ in tumors with high or low caspase-3 enzyme activity. Conclusions [18F]WC-4-116 uptake in vivo reflects increased caspase-3 activation and may be useful for detecting caspase-3 mediated apoptosis treatment responses in cancer. PMID:25344147

  6. Quantitative PET imaging with the 3T MR-BrainPET

    NASA Astrophysics Data System (ADS)

    Weirich, C.; Scheins, J.; Lohmann, P.; Tellmann, L.; Byars, L.; Michel, C.; Rota Kops, E.; Brenner, D.; Herzog, H.; Shah, N. J.

    2013-02-01

    The new hybrid imaging technology of MR-PET allows for simultaneous acquisition of versatile MRI contrasts and the quantitative metabolic imaging with PET. In order to achieve the quantification of PET images with minimal residual error the application of several corrections is crucial. In this work we present our results on quantification with the 3T MR BrainPET scanner.

  7. Children's drawings and attachment to pets.

    PubMed

    Kidd, A H; Kidd, R M

    1995-08-01

    To help confirm the concept that distances placed between the self and other figures in children's drawings represent emotional distances, 242 pet-owning and 35 nonpet-owning kindergartners through eighth graders drew pictures of themselves, a pet, and/or a family member. Owners drew pets significantly closer than family-figures although the younger the child, the greater the distance between self and pet. Older children drew themselves holding pets significantly more often, but younger children placed the family-figure between the self and the pet significantly more often. There were no significant gender differences in self-figure/pet-figure distances, but cats, dogs, caged animals, and farm animals were placed significantly closer to self-figures than were fish. Over-all, owners were clearly emotionally closer to pets than to family members, but nonowners were as close emotionally to family members as were owners. PMID:7501763

  8. F-18 Labeled Diabody-Luciferase Fusion Proteins for Optical-ImmunoPET

    SciTech Connect

    Wu, Anna M

    2013-01-18

    The goal of the proposed work is to develop novel dual-labeled molecular imaging probes for multimodality imaging. Based on small, engineered antibodies called diabodies, these probes will be radioactively tagged with Fluorine-18 for PET imaging, and fused to luciferases for optical (bioluminescence) detection. Performance will be evaluated and validated using a prototype integrated optical-PET imaging system, OPET. Multimodality probes for optical-PET imaging will be based on diabodies that are dually labeled with 18F for PET detection and fused to luciferases for optical imaging. 1) Two sets of fusion proteins will be built, targeting the cell surface markers CEA or HER2. Coelenterazine-based luciferases and variant forms will be evaluated in combination with native substrate and analogs, in order to obtain two distinct probes recognizing different targets with different spectral signatures. 2) Diabody-luciferase fusion proteins will be labeled with 18F using amine reactive [18F]-SFB produced using a novel microwave-assisted, one-pot method. 3) Sitespecific, chemoselective radiolabeling methods will be devised, to reduce the chance that radiolabeling will inactivate either the target-binding properties or the bioluminescence properties of the diabody-luciferase fusion proteins. 4) Combined optical and PET imaging of these dual modality probes will be evaluated and validated in vitro and in vivo using a prototype integrated optical-PET imaging system, OPET. Each imaging modality has its strengths and weaknesses. Development and use of dual modality probes allows optical imaging to benefit from the localization and quantitation offered by the PET mode, and enhances the PET imaging by enabling simultaneous detection of more than one probe.

  9. Recent Understandings of Pet Allergies

    PubMed Central

    Ownby, Dennis; Johnson, Christine Cole

    2016-01-01

    Allergic reactions to pets have been recognized for at least a hundred years. Yet our understanding of the effects of all of the interactions between pet exposures and human immune responses continues to grow. Allergists, epidemiologists, and immunologists have spent years trying to better understand how exposures to pet allergens lead to allergic sensitization (the production of allergen-specific immunoglobulin class E [IgE] antibodies) and subsequent allergic disease. A major new development in this understanding is the recognition that pet exposures consist of not only allergen exposures but also changes in microbial exposures. Exposures to certain pet-associated microbes, especially in the neonatal period, appear to be able to dramatically alter how a child’s immune system develops and this in turn reduces the risk of allergic sensitization and disease. An exciting challenge in the next few years will be to see whether these changes can be developed into a realistic preventative strategy with the expectation of significantly reducing allergic disease, especially asthma. PMID:26918180

  10. [PET and SPECT in epilepsy].

    PubMed

    Setoain, X; Carreño, M; Pavía, J; Martí-Fuster, B; Campos, F; Lomeña, F

    2014-01-01

    Epilepsy is one of the most frequent chronic neurological disorders, affecting 1-2% of the population. Patients with complex partial drug resistant episodes may benefit from a surgical treatment consisting in the excision of the epileptogenic area. Localization of the epileptogenic area was classically performed with video-EEG and magnetic resonance (MR). Recently, functional neuroimaging studies of Nuclear Medicine, positron emission tomography (PET) and single photon emission tomography (SPECT) have demonstrated their utility in the localization of the epileptogenic area prior to surgery. Ictal SPECT with brain perfusion tracers show an increase in blood flow in the initial ictal focus, while PET with (18)FDG demonstrates a decrease of glucose metabolism in the interictal functional deficit zone. In this review, the basic principles and methodological characteristics of the SPECT and PET in epilepsy are described. The ictal SPECT injection mechanism, different patterns of perfusion based on the time of ictal, postictal or interictal injection are detailed and the different diagnostic sensitivities of each one of these SPECT are reviewed. Different methods of analysis of the images with substraction and fusion systems with the MR are described. Similarly, the injection methodology, quantification and evaluation of the images of the PET in epilepsy are described. Finally, the main clinical indications of SPECT and PET in temporal and extratemporal epilepsy are detailed. PMID:24565567

  11. F-18-FDG-PET Confined Radiotherapy of Locally Advanced NSCLC With Concomitant Chemotherapy: Results of the PET-PLAN Pilot Trial

    SciTech Connect

    Fleckenstein, Jochen; Hellwig, Dirk; Kremp, Stephanie; Grgic, Aleksandar; Groeschel, Andreas; Kirsch, Carl-Martin; Nestle, Ursula; Ruebe, Christian

    2011-11-15

    Purpose: The integration of fluoro-deoxy-D-glucose positron emission tomography (FDG-PET) in the process of radiotherapy (RT) planning of locally advanced non-small-cell lung cancer (NSCLC) may improve diagnostic accuracy and minimize interobserver variability compared with target volume definition solely based on computed tomography. Furthermore, irradiating only FDG-PET-positive findings and omitting elective nodal regions may allow dose escalation by treating smaller volumes. The aim of this prospective pilot trial was to evaluate the therapeutic safety of FDG-PET-based RT treatment planning with an autocontour-derived delineation of the primary tumor. Methods and Materials: Eligible patients had Stages II-III inoperable NSCLC, and simultaneous, platinum-based radiochemotherapy was indicated. FDG-PET and computed tomography acquisitions in RT treatment planning position were coregistered. The clinical target volume (CTV) included the FDG-PET-defined primary tumor, which was autodelineated with a source-to-background algorithm, plus FDG-PET-positive lymph node stations. Limited by dose restrictions for normal tissues, prescribed total doses were in the range of 66.6 to 73.8 Gy. The primary endpoint was the rate of out-of-field isolated nodal recurrences (INR). Results: As per intent to treat, 32 patients received radiochemotherapy. In 15 of these patients, dose escalation above 66.6 Gy was achieved. No Grade 4 toxicities occurred. After a median follow-up time of 27.2 months, the estimated median survival time was 19.3 months. During the observation period, one INR was observed in 23 evaluable patients. Conclusions: FDG-PET-confined target volume definition in radiochemotherapy of NSCLC, based on a contrast-oriented source-to-background algorithm, was associated with a low risk of INR. It might provide improved tumor control because of dose escalation.

  12. Current use of PSMA-PET in prostate cancer management.

    PubMed

    Maurer, Tobias; Eiber, Matthias; Schwaiger, Markus; Gschwend, Jürgen E

    2016-04-01

    Currently, the findings of imaging procedures used for detection or staging of prostate cancer depend on morphology of lymph nodes or bone metabolism and do not always meet diagnostic needs. Prostate-specific membrane antigen (PSMA), a transmembrane protein that has considerable overexpression on most prostate cancer cells, has gained increasing interest as a target molecule for imaging. To date, several small compounds for labelling PSMA have been developed and are currently being investigated as imaging probes for PET with the (68)Ga-labelled PSMA inhibitor Glu-NH-CO-NH-Lys(Ahx)-HBED-CC being the most widely studied agent. (68)Ga-PSMA-PET imaging in combination with multiparametric MRI (mpMRI) might provide additional molecular information on cancer localization within the prostate. In patients with primary prostate cancer of intermediate-risk to high-risk, PSMA-based imaging has been reported to improve detection of metastatic disease compared with CT or mpMRI, rendering additional cross-sectional imaging or bone scintigraphy unnecessary. Furthermore, in patients with biochemically recurrent prostate cancer, use of (68)Ga-PSMA-PET imaging has been shown to increase detection of metastatic sites, even at low serum PSA values, compared with conventional imaging or PET examination with different tracers. Thus, although current knowledge is still limited and derived mostly from retrospective series, PSMA-based imaging holds great promise to improve prostate cancer management. PMID:26902337

  13. The evolution of PET-CT.

    PubMed

    Wilson, Bettye G

    2005-01-01

    Positron emission tomography-computed tomography (PET-CT) was the first fused or combined medical imaging technique. Although PET-CT has received widespread acclaim as a major imaging advancement, many questions have surfaced regarding its use. This article answers some of these questions and examines what PET-CT means to medicine and the medical imaging community. PMID:15835615

  14. 36 CFR 13.1234 - Pets.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Pets. 13.1234 Section 13.1234 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... § 13.1234 Pets. Possessing a pet in the BCDA is prohibited....

  15. 36 CFR 13.978 - Pets.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Pets. 13.978 Section 13.978 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... (fda) § 13.978 Pets. Possessing a pet is prohibited— (a) In the FDA, except in public parking areas,...

  16. 36 CFR 13.978 - Pets.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Pets. 13.978 Section 13.978 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... (fda) § 13.978 Pets. Possessing a pet is prohibited— (a) In the FDA, except in public parking areas,...

  17. 36 CFR 13.1106 - Pets.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Pets. 13.1106 Section 13.1106 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... Provisions § 13.1106 Pets. Pets are prohibited except— (a) On the Bartlett Cove Public Use Dock; (b) On...

  18. 36 CFR 13.1234 - Pets.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Pets. 13.1234 Section 13.1234 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... § 13.1234 Pets. Possessing a pet in the BCDA is prohibited....

  19. 36 CFR 13.1106 - Pets.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Pets. 13.1106 Section 13.1106 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... Provisions § 13.1106 Pets. Pets are prohibited except— (a) On the Bartlett Cove Public Use Dock; (b) On...

  20. 7 CFR 500.10 - Pets.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 6 2010-01-01 2010-01-01 false Pets. 500.10 Section 500.10 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE NATIONAL ARBORETUM Conduct on U.S. National Arboreturm Property § 500.10 Pets. Pets brought upon...

  1. 7 CFR 500.10 - Pets.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 6 2011-01-01 2011-01-01 false Pets. 500.10 Section 500.10 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE NATIONAL ARBORETUM Conduct on U.S. National Arboreturm Property § 500.10 Pets. Pets brought upon...

  2. Pet therapy: dogs de-stress students.

    PubMed

    Young, Judith S

    2012-01-01

    Research supports the efficacy of the human-animal bond and pet therapy in a variety of settings. At nursing students' request at one school, the author began offering pet therapy prior to examinations. Anecdotal evidence of a study with the author's Golden Retriever, Goldilocks, demonstrates that pet therapy can reduce test anxiety and improve nursing student performance. PMID:23082615

  3. 7 CFR 500.10 - Pets.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 6 2013-01-01 2013-01-01 false Pets. 500.10 Section 500.10 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE NATIONAL ARBORETUM Conduct on U.S. National Arboreturm Property § 500.10 Pets. Pets brought upon...

  4. 36 CFR 13.1234 - Pets.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Pets. 13.1234 Section 13.1234 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... § 13.1234 Pets. Possessing a pet in the BCDA is prohibited....

  5. 36 CFR 13.1234 - Pets.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Pets. 13.1234 Section 13.1234 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... § 13.1234 Pets. Possessing a pet in the BCDA is prohibited....

  6. 36 CFR 13.1106 - Pets.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Pets. 13.1106 Section 13.1106 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... Provisions § 13.1106 Pets. Pets are prohibited except— (a) On the Bartlett Cove Public Use Dock; (b) On...

  7. 36 CFR 13.1106 - Pets.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Pets. 13.1106 Section 13.1106 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... Provisions § 13.1106 Pets. Pets are prohibited except— (a) On the Bartlett Cove Public Use Dock; (b) On...

  8. 36 CFR 13.978 - Pets.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Pets. 13.978 Section 13.978 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... (fda) § 13.978 Pets. Possessing a pet is prohibited— (a) In the FDA, except in public parking areas,...

  9. 36 CFR 13.1234 - Pets.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Pets. 13.1234 Section 13.1234 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... § 13.1234 Pets. Possessing a pet in the BCDA is prohibited....

  10. 36 CFR 13.978 - Pets.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Pets. 13.978 Section 13.978 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... (fda) § 13.978 Pets. Possessing a pet is prohibited— (a) In the FDA, except in public parking areas,...

  11. 7 CFR 500.10 - Pets.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 6 2014-01-01 2014-01-01 false Pets. 500.10 Section 500.10 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE NATIONAL ARBORETUM Conduct on U.S. National Arboreturm Property § 500.10 Pets. Pets brought upon...

  12. 36 CFR 13.1106 - Pets.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Pets. 13.1106 Section 13.1106 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... Provisions § 13.1106 Pets. Pets are prohibited except— (a) On the Bartlett Cove Public Use Dock; (b) On...

  13. 36 CFR 13.978 - Pets.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Pets. 13.978 Section 13.978 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR NATIONAL PARK... (fda) § 13.978 Pets. Possessing a pet is prohibited— (a) In the FDA, except in public parking areas,...

  14. 7 CFR 500.10 - Pets.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 6 2012-01-01 2012-01-01 false Pets. 500.10 Section 500.10 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE NATIONAL ARBORETUM Conduct on U.S. National Arboreturm Property § 500.10 Pets. Pets brought upon...

  15. 36 CFR 2.15 - Pets.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Pets. 2.15 Section 2.15 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR RESOURCE PROTECTION, PUBLIC USE AND RECREATION § 2.15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public building, public transportation...

  16. 36 CFR 1002.15 - Pets.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 3 2010-07-01 2010-07-01 false Pets. 1002.15 Section 1002.15 Parks, Forests, and Public Property PRESIDIO TRUST RESOURCE PROTECTION, PUBLIC USE AND RECREATION § 1002.15 Pets. (a) The following are prohibited: (1) Possessing a pet in a public building, public transportation vehicle, or location designated as...

  17. A Guide to Managing Your Classroom Pets.

    ERIC Educational Resources Information Center

    Caras, Robert

    1980-01-01

    The author suggests eight ideal classroom pets: hamsters; turtles; snakes; spiders; frogs and toads; fish; and birds. For each he gives suggestions on selecting the pet and housing and feeding it in the classroom. Desert terrariums and home pet care training are also discussed. (SJL)

  18. Pet care during preadolescence: developmental considerations.

    PubMed

    Davis, J H

    1987-01-01

    This exploratory study investigated pet care in relation to psychosocial development during preadolescence. A group of male and female preadolescents (n = 22) at appropriate grade level for age completed a dog care responsibility inventory. The results revealed that preadolescents in general do not routinely care for pets. Mothers appear to assume most pet care tasks. PMID:3664972

  19. PET/MRI: challenges, solutions and perspectives.

    PubMed

    Herzog, Hans

    2012-12-01

    Already from the start of PET/CT integrating positron emission tomography (PET) and computed tomography (CT) in one instrument, there have been considerations how to combine PET and magnetic resonance imaging (MRI) so that their complementary abilities can be utilized in a single investigation. Since classical PET electronics fail in an even weak magnetic field and PET signal processing might disturb high-frequency signals of MRI, it soon became clear that new solutions had to be found to avoid mutual interferences. During the last fifteen years a number of different approaches towards PET/MRI for small animal imaging have been developed by research groups which together with their specific features are summarized in this review. Recently, PET/MRI for human imaging became available as well - this time by industrial initiatives. First some prototypes of BrainPET/MRI were developed followed by commercial products for simultaneous and non-simultaneous whole-body PET/MRI. Although only PET/MRI integrated in one scanner offers the full diversity of complementary multiparametric imaging, there are also promising applications of non-simultaneous sequential PET/MRI. While describing the present instrumentation for human PET/MRI, this review discusses the challenges and promises related to this new imaging technology. PMID:22925652

  20. PET imaging: An overview and instrumentation

    SciTech Connect

    Daghighian, F.; Sumida, R.; Phelps, M.E. )

    1990-03-01

    This is the first article of a four-part series on positron emission tomography (PET). Upon completing the article, the reader should be able to: (1) comprehend the basic principles of PET; (2) explain various technical aspects; and (3) identify radiopharmaceuticals used in PET imaging.

  1. 7 CFR 503.11 - Pets.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 6 2014-01-01 2014-01-01 false Pets. 503.11 Section 503.11 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE CONDUCT ON PLUM ISLAND ANIMAL DISEASE CENTER § 503.11 Pets. No pets or animals of any kind may be...

  2. 7 CFR 503.11 - Pets.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 6 2011-01-01 2011-01-01 false Pets. 503.11 Section 503.11 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE CONDUCT ON PLUM ISLAND ANIMAL DISEASE CENTER § 503.11 Pets. No pets or animals of any kind may be...

  3. 7 CFR 503.11 - Pets.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 6 2010-01-01 2010-01-01 false Pets. 503.11 Section 503.11 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE CONDUCT ON PLUM ISLAND ANIMAL DISEASE CENTER § 503.11 Pets. No pets or animals of any kind may be...

  4. 7 CFR 503.11 - Pets.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 6 2012-01-01 2012-01-01 false Pets. 503.11 Section 503.11 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE CONDUCT ON PLUM ISLAND ANIMAL DISEASE CENTER § 503.11 Pets. No pets or animals of any kind may be...

  5. 7 CFR 503.11 - Pets.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 6 2013-01-01 2013-01-01 false Pets. 503.11 Section 503.11 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL RESEARCH SERVICE, DEPARTMENT OF AGRICULTURE CONDUCT ON PLUM ISLAND ANIMAL DISEASE CENTER § 503.11 Pets. No pets or animals of any kind may be...

  6. Saying Goodbye: Pet Loss and Its Implications

    ERIC Educational Resources Information Center

    Duffey, Thelma

    2005-01-01

    Pets can be loyal, loving, and entertaining members of a family. Their deaths are generally experienced as painful losses by the people who love them, even though the grief experience is often culturally disenfranchised. In this manuscript, we discuss the role that pets can play in a person's life; the effects that pet loss can have on the people…

  7. Latest achievements in PET techniques

    NASA Astrophysics Data System (ADS)

    Del Guerra, Alberto; Belcari, Nicola; Motta, Alfonso; Di Domenico, Giovanni; Sabba, Nicola; Zavattini, Guido

    2003-11-01

    Positron emission tomography (PET) has moved from a distinguished research tool in physiology, cardiology and neurology to become a major tool for clinical investigation in oncology, in cardiac applications and in neurological disorders. Much of the PET accomplishments is due to the remarkable improvements in the last 10 years both in hardware and software aspects. Nowadays a similar effort is made by many research groups towards the construction of dedicated PET apparatus in new emerging fields such as molecular medicine, gene therapy, breast cancer imaging and combined modalities. This paper reports on some recent results we have obtained in small animal imaging and positron emission mammography, based on the use of advanced technology in the field of scintillators and photodetectors, such as Position-Sensitive Detectors coupled to crystal matrices, combined use of scintillating fibers and Hybrid-Photo-Diodes readout, and Hamamatsu flat panels. New ideas and future developments are discussed.

  8. Promoting the exotic pet practice.

    PubMed

    Harris, Don J

    2005-09-01

    The marketing and promotion of an exotic pet veterinary practice allows the use of strategies that are not necessarily available in other veterinary disciplines. The advantage that an exotics practice enjoys is that it is able to capitalize not only on the unique nature of the species being attended but also on the specialized features of the hospital itself that make it specifically appropriate in caring for exotic pets. Before marketing, however, comes the responsibility that the practice live up to the claims made in promotional materials. A practice cannot ethically be presented as an "exotics" practice if it is nothing more than a dog and cat facility that is willing to attend to exotic pets. It is the competence of the veterinary staff and the appropriateness of the facility that determines the suitability of the practice for exotics management. PMID:16129354

  9. Understanding advertising in pet nutrition.

    PubMed

    Brown, R G

    1994-04-01

    Advertising is part of the effort to attract attention of consumers to products, in this case, pet foods. It is generally benign in its effect, but it can be misleading, although rarely deliberately so. It uses a specialized vocabulary, which must be mastered if one is to understand what is intended. For all of the expense and effort, advertising figures directly in relatively few decisions to purchase. Its main intention is to call our attention to a particular pet food and to give that product an image. If the pet food does not perform in the consumer's hands, then all of the advertising on earth will not be persuasive. On the other hand, if a product performs well, the word-of-mouth will be positive and that mode of advertising is one of the most effective. PMID:8076285

  10. Understanding advertising in pet nutrition.

    PubMed Central

    Brown, R G

    1994-01-01

    Advertising is part of the effort to attract attention of consumers to products, in this case, pet foods. It is generally benign in its effect, but it can be misleading, although rarely deliberately so. It uses a specialized vocabulary, which must be mastered if one is to understand what is intended. For all of the expense and effort, advertising figures directly in relatively few decisions to purchase. Its main intention is to call our attention to a particular pet food and to give that product an image. If the pet food does not perform in the consumer's hands, then all of the advertising on earth will not be persuasive. On the other hand, if a product performs well, the word-of-mouth will be positive and that mode of advertising is one of the most effective. PMID:8076285

  11. ToxCast Assay Network (TCAN) Viewer: A Visualization Tool for High-throughput Assay Chemical Data (SOT)

    EPA Science Inventory

    USEPA’s ToxCast program has generated high-throughput bioactivity screening (HTS) data on thousands of chemicals. The ToxCast program has described and annotated the HTS assay battery with respect to assay design and target information (e.g., gene target). Recent stakeholder and ...

  12. Towards integration of PET/MR hybrid imaging into radiation therapy treatment planning

    SciTech Connect

    Paulus, Daniel H.; Thorwath, Daniela; Schmidt, Holger; Quick, Harald H.

    2014-07-15

    Purpose: Multimodality imaging has become an important adjunct of state-of-the-art radiation therapy (RT) treatment planning. Recently, simultaneous PET/MR hybrid imaging has become clinically available and may also contribute to target volume delineation and biological individualization in RT planning. For integration of PET/MR hybrid imaging into RT treatment planning, compatible dedicated RT devices are required for accurate patient positioning. In this study, prototype RT positioning devices intended for PET/MR hybrid imaging are introduced and tested toward PET/MR compatibility and image quality. Methods: A prototype flat RT table overlay and two radiofrequency (RF) coil holders that each fix one flexible body matrix RF coil for RT head/neck imaging have been evaluated within this study. MR image quality with the RT head setup was compared to the actual PET/MR setup with a dedicated head RF coil. PET photon attenuation and CT-based attenuation correction (AC) of the hardware components has been quantitatively evaluated by phantom scans. Clinical application of the new RT setup in PET/MR imaging was evaluated in anin vivo study. Results: The RT table overlay and RF coil holders are fully PET/MR compatible. MR phantom and volunteer imaging with the RT head setup revealed high image quality, comparable to images acquired with the dedicated PET/MR head RF coil, albeit with 25% reduced SNR. Repositioning accuracy of the RF coil holders was below 1 mm. PET photon attenuation of the RT table overlay was calculated to be 3.8% and 13.8% for the RF coil holders. With CT-based AC of the devices, the underestimation error was reduced to 0.6% and 0.8%, respectively. Comparable results were found within the patient study. Conclusions: The newly designed RT devices for hybrid PET/MR imaging are PET and MR compatible. The mechanically rigid design and the reproducible positioning allow for straightforward CT-based AC. The systematic evaluation within this study provides the

  13. Competitive Advantage of PET/MRI

    PubMed Central

    Jadvar, Hossein; Colletti, Patrick M.

    2013-01-01

    Multimodality imaging has made great strides in the imaging evaluation of patients with a variety of diseases. Positron emission tomography/computed tomography (PET/CT) is now established as the imaging modality of choice in many clinical conditions, particularly in oncology. While the initial development of combined PET/magnetic resonance imaging (PET/MRI) was in the preclinical arena, hybrid PET/MR scanners are now available for clinical use. PET/MRI combines the unique features of MRI including excellent soft tissue contrast, diffusion-weighted imaging, dynamic contrast-enhanced imaging, fMRI and other specialized sequences as well as MR spectroscopy with the quantitative physiologic information that is provided by PET. Most evidence for the potential clinical utility of PET/MRI is based on studies performed with side-by-side comparison or software-fused MRI and PET images. Data on distinctive utility of hybrid PET/MRI are rapidly emerging. There are potential competitive advantages of PET/MRI over PET/CT. In general, PET/MRI may be preferred over PET/CT where the unique features of MRI provide more robust imaging evaluation in certain clinical settings. The exact role and potential utility of simultaneous data acquisition in specific research and clinical settings will need to be defined. It may be that simultaneous PET/MRI will be best suited for clinical situations that are disease-specific, organ-specific, related to diseases of the children or in those patients undergoing repeated imaging for whom cumulative radiation dose must be kept as low as reasonably achievable. PET/MRI also offers interesting opportunities for use of dual modality probes. Upon clear definition of clinical utility, other important and practical issues related to business operational model, clinical workflow and reimbursement will also be resolved. PMID:23791129

  14. Competitive advantage of PET/MRI.

    PubMed

    Jadvar, Hossein; Colletti, Patrick M

    2014-01-01

    Multimodality imaging has made great strides in the imaging evaluation of patients with a variety of diseases. Positron emission tomography/computed tomography (PET/CT) is now established as the imaging modality of choice in many clinical conditions, particularly in oncology. While the initial development of combined PET/magnetic resonance imaging (PET/MRI) was in the preclinical arena, hybrid PET/MR scanners are now available for clinical use. PET/MRI combines the unique features of MRI including excellent soft tissue contrast, diffusion-weighted imaging, dynamic contrast-enhanced imaging, fMRI and other specialized sequences as well as MR spectroscopy with the quantitative physiologic information that is provided by PET. Most evidence for the potential clinical utility of PET/MRI is based on studies performed with side-by-side comparison or software-fused MRI and PET images. Data on distinctive utility of hybrid PET/MRI are rapidly emerging. There are potential competitive advantages of PET/MRI over PET/CT. In general, PET/MRI may be preferred over PET/CT where the unique features of MRI provide more robust imaging evaluation in certain clinical settings. The exact role and potential utility of simultaneous data acquisition in specific research and clinical settings will need to be defined. It may be that simultaneous PET/MRI will be best suited for clinical situations that are disease-specific, organ-specific, related to diseases of the children or in those patients undergoing repeated imaging for whom cumulative radiation dose must be kept as low as reasonably achievable. PET/MRI also offers interesting opportunities for use of dual modality probes. Upon clear definition of clinical utility, other important and practical issues related to business operational model, clinical workflow and reimbursement will also be resolved. PMID:23791129

  15. A High-Throughput Radiometric Kinase Assay.

    PubMed

    Duong-Ly, Krisna C; Peterson, Jeffrey R

    2016-01-01

    Aberrant kinase signaling has been implicated in a number of diseases. While kinases have become attractive drug targets, only a small fraction of human protein kinases have validated inhibitors. Screening of libraries of compounds against a kinase or kinases of interest is routinely performed during kinase inhibitor development to identify promising scaffolds for a particular target and to identify kinase targets for compounds of interest. Screening of more focused compound libraries may also be conducted in the later stages of inhibitor development to improve potency and optimize selectivity. The dot blot kinase assay is a robust, high-throughput kinase assay that can be used to screen a number of small-molecule compounds against one kinase of interest or several kinases. Here, a protocol for a dot blot kinase assay used for measuring insulin receptor kinase activity is presented. This protocol can be readily adapted for use with other protein kinases. PMID:26501904

  16. Development of PhytoPET: A plant imaging PET system

    SciTech Connect

    Dong, H; Lee, S J; McKisson, J; Xi, W; Zorn, C; Howell, C R; Crowell, A S; Cumberbatch, L; Reid, C D; Smith, M F; Stolin, A

    2012-02-01

    The development and initial evaluation of a high-resolution positron emission tomography (PET) system to image the biodistribution of positron emitting tracers in live plants is underway. The positron emitting {sup 11}CO{sub 2} tracer is used in plant biology research investigating carbon sequestration in biomass, optimization of plant productivity and biofuel development. This PhytoPET design allows flexible arrangements of PET detectors based on individual standalone detector modules built from single 5 cm x 5 cm Hamamatsu H8500 position sensitive photomultiplier tubes. Each H8500 is coupled to a LYSO:Ce scintillator array composed of 48 x 48 elements that are 10 mm thick arranged with a 1.0 mm pitch. An Ethernet based 12-bit flash analog to digital data acquisition system with onboard coincident matrix definition is under development to digitize the signals. The detector modules of the PhytoPET system can be arranged and stacked to accommodate various sized plants and plant structures.

  17. Methods for threshold determination in multiplexed assays

    DOEpatents

    Tammero, Lance F. Bentley; Dzenitis, John M; Hindson, Benjamin J

    2014-06-24

    Methods for determination of threshold values of signatures comprised in an assay are described. Each signature enables detection of a target. The methods determine a probability density function of negative samples and a corresponding false positive rate curve. A false positive criterion is established and a threshold for that signature is determined as a point at which the false positive rate curve intersects the false positive criterion. A method for quantitative analysis and interpretation of assay results together with a method for determination of a desired limit of detection of a signature in an assay are also described.

  18. PET-Based Percutaneous Needle Biopsy.

    PubMed

    El-Haddad, Ghassan

    2016-07-01

    PET can be used to guide percutaneous needle biopsy to the most metabolic lesion, improving diagnostic yield. PET biopsy guidance can be performed using visual or software coregistration, electromagnetic needle tracking, cone-beam computed tomography (CT), and intraprocedural PET/CT guidance. PET/CT-guided biopsies allow the sampling of lesions that may not be clearly visible on anatomic imaging, or of lesions that are morphologically normal. PET can identify suspicious locations within complex tumors that are most likely to contain important diagnostic and prognostic information. PMID:27321036

  19. Hybrid MR-PET in Neuroimaging.

    PubMed

    Bisdas, S; Lá Fougere, C; Ernemann, U

    2015-10-01

    Hybrid magnetic resonance (MR)-positron emission tomography (MR-PET) is a novel technology with advantages over sequential MR and PET imaging, allowing maintain full individual diagnostic performance with negligible mutual interference between the two hardware settings. Obvious synergies between MR and PET in acquisition of anatomical, functional, and molecular information for neurological diseases into one single image pave the way for establishing clear clinical indications for hybrid MR-PET as well as addressing unmet neuroimaging needs in future clinics and research. Further developments in attenuation correction, quantification, workflow, and effective MR-PET data management might unfold the full potential of integrated multimodality imaging. PMID:26227618

  20. Development of PET/MRI with insertable PET for simultaneous PET and MR imaging of human brain

    SciTech Connect

    Jung, Jin Ho; Choi, Yong Jung, Jiwoong; Kim, Sangsu; Lim, Hyun Keong; Im, Ki Chun; Oh, Chang Hyun; Park, Hyun-wook; Kim, Kyung Min; Kim, Jong Guk

    2015-05-15

    Purpose: The purpose of this study was to develop a dual-modality positron emission tomography (PET)/magnetic resonance imaging (MRI) with insertable PET for simultaneous PET and MR imaging of the human brain. Methods: The PET detector block was composed of a 4 × 4 matrix of detector modules, each consisting of a 4 × 4 array LYSO coupled to a 4 × 4 Geiger-mode avalanche photodiode (GAPD) array. The PET insert consisted of 18 detector blocks, circularly mounted on a custom-made plastic base to form a ring with an inner diameter of 390 mm and axial length of 60 mm. The PET gantry was shielded with gold-plated conductive fabric tapes with a thickness of 0.1 mm. The charge signals of PET detector transferred via 4 m long flat cables were fed into the position decoder circuit. The flat cables were shielded with a mesh-type aluminum sheet with a thickness of 0.24 mm. The position decoder circuit and field programmable gate array-embedded DAQ modules were enclosed in an aluminum box with a thickness of 10 mm and located at the rear of the MR bore inside the MRI room. A 3-T human MRI system with a Larmor frequency of 123.7 MHz and inner bore diameter of 60 cm was used as the PET/MRI hybrid system. A custom-made radio frequency (RF) coil with an inner diameter of 25 cm was fabricated. The PET was positioned between gradient and the RF coils. PET performance was measured outside and inside the MRI scanner using echo planar imaging, spin echo, turbo spin echo, and gradient echo sequences. MRI performance was also evaluated with and without the PET insert. The stability of the newly developed PET insert was evaluated and simultaneous PET and MR images of a brain phantom were acquired. Results: No significant degradation of the PET performance caused by MR was observed when the PET was operated using various MR imaging sequences. The signal-to-noise ratio of MR images was slightly degraded due to the PET insert installed inside the MR bore while the homogeneity was

  1. PET-Based Personalized Management in Clinical Oncology: An Unavoidable Path for the Foreseeable Future.

    PubMed

    Basu, Sandip; Alavi, Abass

    2016-07-01

    It is imperative that the thrust of clinical practice in the ensuing years would be to develop personalized management model for various disorders. PET-computed tomography (PET-CT) based molecular functional imaging has been increasingly utilized for assessment of tumor and other nonmalignant disorders and has the ability to explore disease phenotype on an individual basis and address critical clinical decision making questions related to practice of personalized medicine. Hence, it is essential to make a concerted systematic effort to explore and define the appropriate place of PET-CT in personalized clinical practice in each of malignancies, which would strengthen the concept further. The potential advantages of PET based disease management can be classified into broad categories: (1) Traditional: which includes assessment of disease extent such as initial disease staging and restaging, treatment response evaluation particularly early in the course and thus PET-CT response adaptive decision for continuing the same regimen or switching to salvage schedules; there has been continuous addition of newer application of PET based disease restaging in oncological parlance (eg, Richter transformation); (2) Recent and emerging developments: this includes exploring tumor biology with FDG and non-FDG PET tracers. The potential of multitracer PET imaging (particularly new and novel tracers, eg, 68Ga-DOTA-TOC/NOC/TATE in NET, 68Ga-PSMA and 18F-fluorocholine in prostate carcinoma, 18F-fluoroestradiol in breast carcinoma) has provided a scientific basis to stratify and select appropriate targeted therapies (both radionuclide and nonradionuclide treatment), a major boost for individualized disease management in clinical oncology. Integrating the molecular level information obtained from PET with structural imaging further individualizing treatment plan in radiation oncology, precision of interventions and biopsies of a particular lesion and forecasting disease prognosis. PMID

  2. Absolute nuclear material assay

    DOEpatents

    Prasad, Manoj K.; Snyderman, Neal J.; Rowland, Mark S.

    2012-05-15

    A method of absolute nuclear material assay of an unknown source comprising counting neutrons from the unknown source and providing an absolute nuclear material assay utilizing a model to optimally compare to the measured count distributions. In one embodiment, the step of providing an absolute nuclear material assay comprises utilizing a random sampling of analytically computed fission chain distributions to generate a continuous time-evolving sequence of event-counts by spreading the fission chain distribution in time.

  3. Absolute nuclear material assay

    DOEpatents

    Prasad, Manoj K.; Snyderman, Neal J.; Rowland, Mark S.

    2010-07-13

    A method of absolute nuclear material assay of an unknown source comprising counting neutrons from the unknown source and providing an absolute nuclear material assay utilizing a model to optimally compare to the measured count distributions. In one embodiment, the step of providing an absolute nuclear material assay comprises utilizing a random sampling of analytically computed fission chain distributions to generate a continuous time-evolving sequence of event-counts by spreading the fission chain distribution in time.

  4. The MiniPET: a didactic PET system

    NASA Astrophysics Data System (ADS)

    Pedro, R.; Silva, J.; Gurriana, L.; Silva, J. M.; Maio, A.; Soares Augusto, J.

    2013-03-01

    The MiniPET project aims to design and build a small PET system. It consists of two 4 × 4 matrices of 16 LYSO scintillator crystals and two PMTs with 16 channels resulting in a low cost system with the essential functionality of a clinical PET instrument. It is designed to illustrate the physics of the PET technique and to provide a didactic platform for the training of students and nuclear imaging professionals as well as for scientific outreach. The PET modules can be configured to test for the coincidence of 511 keV gamma rays. The model has a flexible mechanical setup [1] and can simulate 14 diferent ring geometries, from a configuration with as few as 18 detectors per ring (ring radius phi=51 mm), up to a geometry with 70 detectors per ring (phi=200 mm). A second version of the electronic system [2] allowed measurement and recording of the energy deposited in 4 detector channels by photons from a 137Cs radioactive source and by photons resulting of the annihilation of positrons from a 22Na radioactive source. These energy spectra are used for detector performance studies, as well as angular dependency studies. In this paper, the mechanical setup, the front-end high-speed analog electronics, the digital acquisition and control electronics implemented in a FPGA, as well as the data-transfer interface between the FPGA board and a host PC are described. Recent preliminary results obtained with the 4 active channels in the prototype are also presented.

  5. Principles of PET/MR Imaging.

    PubMed

    Disselhorst, Jonathan A; Bezrukov, Ilja; Kolb, Armin; Parl, Christoph; Pichler, Bernd J

    2014-05-12

    Hybrid PET/MR systems have rapidly progressed from the prototype stage to systems that are increasingly being used in the clinics. This review provides an overview of developments in hybrid PET/MR systems and summarizes the current state of the art in PET/MR instrumentation, correction techniques, and data analysis. The strong magnetic field requires considerable changes in the manner by which PET images are acquired and has led, among others, to the development of new PET detectors, such as silicon photomultipliers. During more than a decade of active PET/MR development, several system designs have been described. The technical background of combined PET/MR systems is explained and related challenges are discussed. The necessity for PET attenuation correction required new methods based on MR data. Therefore, an overview of recent developments in this field is provided. Furthermore, MR-based motion correction techniques for PET are discussed, as integrated PET/MR systems provide a platform for measuring motion with high temporal resolution without additional instrumentation. The MR component in PET/MR systems can provide functional information about disease processes or brain function alongside anatomic images. Against this background, we point out new opportunities for data analysis in this new field of multimodal molecular imaging. PMID:24819419

  6. Two-dimensional electrophoretic mobility shift assay: identification and mapping of transcription factor CTCF target sequences within an FXYD5-COX7A1 region of human chromosome 19.

    PubMed

    Vetchinova, Anna S; Akopov, Sergey B; Chernov, Igor P; Nikolaev, Lev G; Sverdlov, Eugene D

    2006-07-01

    An approach for fast identification and mapping of transcription factor binding sites within long genomic sequences is proposed. Using this approach, 10 CCCTC-binding factor (CTCF) binding sites were identified within a 1-Mb FXYD5-COX7A1 human chromosome 19 region. In vivo binding of CTCF to these sites was verified by chromatin immunoprecipitation assay. CTCF binding sites were mapped within gene introns and intergenic regions, and some of them contained Alu-like repeated elements. PMID:16701069

  7. Detection of Escherichia coli O157:H7 in food using real-time multiplex PCR assays targeting the stx1, stx2, wzyo157, and the fliCh7 or eae genes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Real-time multiplex PCR assays were developed to detect E. coli O157:H7 in different solid and liquid foods. In each of three trials, apple cider and raw milk (25 ml), and ground beef (15% fat) and lettuce (25 g) were inoculated with 2 or 20 CFU of E. coli O157:H7 strain 380-94 (possesses the stx1a...

  8. Diseases Transmitted by Less Common House Pets.

    PubMed

    Chomel, Bruno B

    2015-12-01

    Beside dogs and cats, the most common pets worldwide, an increasing number of pocket pets and exotic pets are making their way to more and more households, especially in North America and Europe. Although many of these animals make appropriate pets, they also can be a source of many zoonotic diseases, especially in young children and immunocompromised individuals. Some of these diseases can be life threatening, such as rabies, rat bite fever, and plague. Some others are quite common, because of the frequency of the pathogens harbored by these species, such as salmonellosis in reptiles and amphibians. Appropriate knowledge of the zoonotic agents carried by these "new" pet species is strongly recommended prior to acquiring pocket or exotic pets. Furthermore, adopting wildlife as pets is strongly discouraged, because it is always a risky action that can lead to major health issues. PMID:27337276

  9. High performance polyester concrete using recycled PET

    SciTech Connect

    Rebeiz, K.S.

    1995-10-01

    Recycled polyethylene terephthalate (PET) plastic wastes could be used in production of unsaturated polyester resins. In turn, these resins could be mixed with inorganic aggregates to produce polymer concrete (PC). Unsaturated polyesters based on recycled PET might be a potentially lower source cost of resins for producing useful PC based-products. The advantage of recycling PET in PC is that the PET materials do not have to be purified, including removal of colors, to the same extent as other PET recycling applications, which should facilitate the recycling operation and minimize its cost. The recycling of PET in PC could also help save energy and allow the long term disposal of the PET waste, an important advantage in recycling applications.

  10. Quantitative simultaneous PET-MR imaging

    NASA Astrophysics Data System (ADS)

    Ouyang, Jinsong; Petibon, Yoann; Huang, Chuan; Reese, Timothy G.; Kolnick, Aleksandra L.; El Fakhri, Georges

    2014-06-01

    Whole-body PET is currently limited by the degradation due to patient motion. Respiratory motion degrades imaging studies of the abdomen. Similarly, both respiratory and cardiac motions significantly hamper the assessment of myocardial ischemia and/or metabolism in perfusion and viability cardiac PET studies. Based on simultaneous PET-MR, we have developed robust and accurate MRI methods allowing the tracking and measurement of both respiratory and cardiac motions during abdominal or cardiac studies. Our list-mode iterative PET reconstruction framework incorporates the measured motion fields into PET emission system matrix as well as the time-dependent PET attenuation map and the position dependent point spread function. Our method significantly enhances the PET image quality as compared to conventional methods.

  11. PET/CT-guided treatment planning for paediatric cancer patients: a simulation study of proton and conventional photon therapy

    PubMed Central

    Brodin, N P; Björk-Eriksson, T; Birk Christensen, C; Kiil-Berthelsen, A; Aznar, M C; Hollensen, C; Markova, E; Munck af Rosenschöld, P

    2015-01-01

    Objective: To investigate the impact of including fluorine-18 fludeoxyglucose (18F-FDG) positron emission tomography (PET) scanning in the planning of paediatric radiotherapy (RT). Methods: Target volumes were first delineated without and subsequently re-delineated with access to 18F-FDG PET scan information, on duplicate CT sets. RT plans were generated for three-dimensional conformal photon RT (3DCRT) and intensity-modulated proton therapy (IMPT). The results were evaluated by comparison of target volumes, target dose coverage parameters, normal tissue complication probability (NTCP) and estimated risk of secondary cancer (SC). Results: Considerable deviations between CT- and PET/CT-guided target volumes were seen in 3 out of the 11 patients studied. However, averaging over the whole cohort, CT or PET/CT guidance introduced no significant difference in the shape or size of the target volumes, target dose coverage, irradiated volumes, estimated NTCP or SC risk, neither for IMPT nor 3DCRT. Conclusion: Our results imply that the inclusion of PET/CT scans in the RT planning process could have considerable impact for individual patients. There were no general trends of increasing or decreasing irradiated volumes, suggesting that the long-term morbidity of RT in childhood would on average remain largely unaffected. Advances in knowledge: 18F-FDG PET-based RT planning does not systematically change NTCP or SC risk for paediatric cancer patients compared with CT only. 3 out of 11 patients had a distinct change of target volumes when PET-guided planning was introduced. Dice and mismatch metrics are not sufficient to assess the consequences of target volume differences in the context of RT. PMID:25494657

  12. PET evaluation of the dopamine system of the human brain

    SciTech Connect

    Volkow, N.D.; Fowler, J.S.; Gatley, S. |

    1996-07-01

    Dopamine plays a pivotal role in the regulation and control of movement, motivation and cognition. It also is closely linked to reward, reinforcement and addiction. Abnormalities in brain dopamine are associated with many neurological and psychiatric disorders including Parkinson`s disease, schizophrenia and substance abuse. This close association between dopamine and neurological and psychiatric diseases and with substance abuse make it an important topic in research in the neurosciences and an important molecular target in drug development. PET enables the direct measurement of components of the dopamine system in the living human brain. It relies on radiotracers which label dopamine receptors, dopamine transporters, precursors of dopamine or compounds which have specificity for the enzymes which degrade dopamine. Additionally, by using tracers that provide information on regional brain metabolism or blood flow as well as neurochemically specific pharmacological interventions, PET can be used to assess the functional consequences of change in brain dopamine activity. PET dopamine measurements have been used to investigate the normal human brain and its involvement in psychiatric and neurological diseases. It has also been used in psychopharmacological research to investigate dopamine drugs used in the treatment of Parkinson`s disease and of schizophrenia as well as to investigate the effects of drugs of abuse on the dopamine system. Since various functional and neurochemical parameters can be studied in the same subject, PET enables investigation of the functional integrity of the dopamine system in the human brain and investigation of the interactions of dopamine with other neurotransmitters. This paper summarizes the different tracers and experimental strategies developed to evaluate the various elements of the dopamine system in the human brain with PET and their applications to clinical research. 254 refs., 7 figs., 3 tabs.

  13. Aging research 2011: exploring the pet dog paradigm.

    PubMed

    Waters, David J

    2011-01-01

    Researchers are counting on comparative biologists to find alternative animal models of human aging that will foster experimental approaches to study disabi