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At least 19 records

Neurotrophic factor - Characterization and partial purification

Recent evidence suggests that neurotrophic activity is required for the normal proliferation and development of muscle cells. The present paper reports a study of the purification and characterization of a neurotrophic factor (NTF) from adult chicken ischiatic-peroneal nerves using two independent quantitative in vitro assay systems. The assays were performed by the measurement of the incorporation of tritiated thymidine or the sizes of single-cell clones by chick muscle cells grown in culture. The greatest amount of neutrotrophic activity is found to be extracted at a pH of 8; aqueous suspensions of the activity are stable to long-term storage at room temperature. The specific activity of the substance is doubled upon precipitation with ammonium sulfate or after gel filtration, and increase 4 to 5 fold after salt gradient elution from DEAE cellulose columns. The active fraction obtained after gel filtration and rechromatography on DEAE cellulose exhibits a 7 to 10-fold increase in specific activity. Electrophoresis of the most highly purified material yields a greatly concentrated band at around 80,000 daltons. Although NTF is purified almost 10-fold as indicated by the increase in specific activity, the maximum activity of the partially purified material is greatly reduced, possibly due to a requirement for a cofactor for the expression of maximum activity.

Popiela, H.↗

A Novel and Secure Pseudovirus Reporter System Based Assay for Neutralizing and Enhancing Antibody Assay Against Marburg Virus

Marburg virus (MARV) is one of the principal members of the filovirus family, which can cause fatal hemorrhagic fever in humans. There are currently no prophylactic and therapeutic drugs on the market, and the high pathogenicity and infectivity of MARV make its research highly dependent on biosafety level 4 conditions, severely hindering the development of vaccines and therapies. Therefore, the development of medicines, such as MARV serological diagnosis, vaccines, and therapeutic antibody drugs, urgently needs a safe, convenient, and biosafety level 2 detection method to measure the neutralizing activity of MARV antibodies. To this end, we report a neutralization assay relying on a Rabies virus (RABV) reverse genetic operating system. We constructed infectious clones carrying the eGFP reporter gene and the full length of the original unmodified MARV GP gene. Based on the critical parameters of phylogenetic analysis, recombinant viruses targeting representative strains in the two major MARV lineages were successfully rescued. These pseudoviruses are safe in mice, and their inability to infect cells after being neutralized by antibodies can be visualized under a fluorescence microscope. We tested the system using the neutralizing antibody MR191. MR191 can significantly block the infection of BSR cells with pseudovirus. We compared it with the traditional lentivirus-type pseudovirus system to verify the system’s credibility and obtained the same results as reported in the literature. In general, we have established a safe and visualized method for evaluating the neutralizing activity of MARV antibodies. Compared with traditional methods, it has the advantages of convenient operation, short cycle, and low cost. It is a candidate method that can replace actual viruses for a neutralization assay.

Bi, Jinhao↗

Bldg. 7930 Cell G Neutron Detector New Electronics Testing

The Cell G neutron assay system within 7930 provides an important function in the production of Californium-252 in the quantitative determination of the amount of Cf-252 present in a sample. The assay system consists of a Uranium-238 fission chamber coupled to the necessary counting electronics including preamplifier, amplifier, discriminator and counter/timer. The original electronics date back to the 1970s but were replaced back in 2011 with little documentation detailing the testing and replacement efforts. This work focused on procuring a new spare set of electronics, their subsequent testing as a backup in case any of the currently in use electronics were to fail and documentation of the existing and new setups.

Taylor, Neil [Oak Ridge National Laboratory (ORNL)↗

Development of a Neutron List Mode Collar (LMCL) and a List Mode Response Matrix Analysis Concept

This report was prepared for the Safeguards Program of the US Department of Energy’s (DOE’s) National Nuclear Security Administration (NNSA), Office of Defense Nuclear Nonproliferation Research and Development (DNN R&D). The report presents the development of the neutron nondestructive assay system, the List Mode Collar (LMCL) for the project OR16-List Mode for Collar-PD1La “List Mode Response Matrix for Advanced Correlated Neutron Analysis for Nuclear Safeguards.” The new list mode electronics developed under this project, and a spatial analysis concept called the List Mode Response Matrix are also described in this report. Analysis algorithms based on classification methods are published in a separate report. This research addresses the need to expand the capabilities of current nondestructive assay systems used for nuclear safeguards applications and considers the sustainability of safeguards technologies by the development of a “retrofit” concept using electronics based on modern standards. Furthermore, employing list mode data acquisition enabled the development of a spatial analysis concept and empirical measurement of a spatial response not previously used for safeguards neutron counting applications or measured in a traditional neutron collar detector and, therefore, provides new capability.

73 NUCLEAR PHYSICS AND RADIATION PHYSICS↗

Real-time biochemical assay telemetering system

The present invention is an apparatus and a method of detecting a chemical released by perspiration, typically through sweat and broadcasting the detection to a receiver. The chemical may be a drug of abuse. The device which is attached to the skin of a subject contains labeled antibodies or label containing microspheres attached to antibodies. The labeled antibodies are bound to solid phase drug via antigen-antibody interaction. These labeled antibodies are displaced from the solid phase support to which they are bound by free drug molecules in the perspiration. These labeled antibodies then migrate through a spacer layer and are trapped by a layer containing a suitable selective binding material. The label is illuminated or excited by a light source and detected by a photodetector. The signal can be recorded, or transmitted to a remote radio monitor.

Kidwell, David A.↗

Development of MCNP Training Modules for Safeguards Practitioners [Abstract]

The Monte-Carlo N-Particle (MCNP) software developed at LANL is the most widely used neutron transport code in the world. It is an essential tool for a variety of applications including detector development and design, nuclear fuel burnup simulation, criticality safety, and nondestructive assay system optimization. For this reason, it is indispensable within the safeguards and materials control & accountability (MC&A) communities. Multiple MCNP training courses have been created and taught over the last several decades by the MCNP development team at LANL, however there are no existing courses that cover specialized topics considered fundamental to NDA and safeguards models. To fill this gap, the MCNP team and Safeguards Science and Technology group at LANL have co-created a set of training modules customized to meet the specialized needs of the safeguards and MC&A communities. The basic modules cover concepts such as NDA system optimization, He-specific and other capture tallies, and tools for improved theoretical understanding. An advanced module was also created to cover topics including variance reduction for active interrogation simulations, use of the LANL MCNPTools post-processor, PTRAC (particle tracking) and list-mode data simulations, and fuel burnup simulations. The training modules teach to the latest and most state-of-the-art MCNP features and tools released by the development team at LANL and are intended to be taught jointly by the developers and safeguards experts. Ultimately, we hope that creation of these modules will serve to capture and convey the safeguards modeling and MCNP expertise at LANL, and that we will be able to share the modules more broadly with the MC&A and safeguards communities.

73 NUCLEAR PHYSICS AND RADIATION PHYSICS↗

Development of a novel minigenome and recombinant VSV expressing Seoul hantavirus glycoprotein-based assays to identify anti-hantavirus therapeutics

Seoul virus (SEOV) is an emerging global health threat that can cause hemorrhagic fever with renal syndrome (HFRS), which results in case fatality rates of ~2%. There are no approved treatments for SEOV infections. We developed a cell-based assay system to identify potential antiviral compounds for SEOV and generated additional assays to characterize the mode of action of any promising antivirals. Here, to test if candidate antivirals targeted SEOV glycoprotein-mediated entry, we developed a recombinant reporter vesicular stomatitis virus expressing SEOV glycoproteins. To facilitate the identification of candidate antiviral compounds targeting viral transcription/replication, we successfully generated the first reported minigenome system for SEOV. This SEOV minigenome (SEOV-MG) screening assay will also serve as a prototype assay for discovery of small molecules inhibiting replication of other hantaviruses, including Andes and Sin Nombre viruses. Ours is a proof-of-concept study in which we tested several compounds previously reported to have activity against other negative-strand RNA viruses using our newly developed hantavirus antiviral screening systems. These systems can be used under lower biocontainment conditions than those needed for infectious viruses, and identified several compounds with robust anti-SEOV activity. Our findings have important implications for the development of anti-hantavirus therapeutics.

60 APPLIED LIFE SCIENCES↗

HEMODOSE: A Set of Multi-parameter Biodosimetry Tools

After the events of September 11, 2001 and recent events at the Fukushima reactors in Japan, there is an increasing concern of the occurrence of nuclear and radiological terrorism or accidents that may result in large casualty in densely populated areas. To guide medical personnel in their clinical decisions for effective medical management and treatment of the exposed individuals, biological markers are usually applied to examine the radiation induced changes at different biological levels. Among these the peripheral blood cell counts are widely used to assess the extent of radiation induced injury. This is due to the fact that hematopoietic system is the most vulnerable part of the human body to radiation damage. Particularly, the lymphocyte, granulocyte, and platelet cells are the most radiosensitive of the blood elements, and monitoring their changes after exposure is regarded as the most practical and best laboratory test to estimate radiation dose. The HEMODOSE web tools are built upon solid physiological and pathophysiological understanding of mammalian hematopoietic systems, and rigorous coarse-grained biomathematical modeling and validation. Using single or serial granulocyte, lymphocyte, leukocyte, or platelet counts after exposure, these tools can estimate absorbed doses of adult victims very rapidly and accurately. Some patient data in historical accidents are utilized as examples to demonstrate the capabilities of these tools as a rapid point-of-care diagnostic or centralized high-throughput assay system in a large scale radiological disaster scenario. Unlike previous dose prediction algorithms, the HEMODOSE web tools establish robust correlations between the absorbed doses and victim's various types of blood cell counts not only in the early time window (1 or 2 days), but also in very late phase (up to 4 weeks) after exposure

Hu, Shaowen↗

Microbiologic assay of space hardware.

Review of the procedures used in the microbiological examination of space hardware. The general procedure for enumerating aerobic and anaerobic microorganisms and spores is outlined. Culture media and temperature-time cycles used for incubation are reviewed, along with assay systems designed for the enumeration of aerobic and anaerobic spores. The special problems which are discussed are involved in the precise and accurate enumeration of microorganisms on surfaces and in the neutralization of viable organisms buried inside solid materials that could be released to a planet's surface if the solid should be fractured. Special attention is given to sampling procedures including also the indirect techniques of surface assays of space hardware such as those using detachable or fallout strips. Some data on comparative levels of microbial contamination on lunar and planetary spacecraft are presented.

Favero, M. S.↗

HEMODOSE: A Set of Multi-parameter Biodosimetry Tools

There continues to be important concerns of the possibility of the occurrence of acute radiation syndromes following nuclear and radiological terrorism or accidents that may result in mass casualties in densely populated areas. To guide medical personnel in their clinical decisions for effective medical management and treatment of the exposed individuals, biological markers are usually applied to examine radiation induced biological changes to assess the severity of radiation injury to sensitive organ systems. Among these the peripheral blood cell counts are widely used to assess the extent of radiation induced bone marrow (BM) injury. This is due to the fact that hematopoietic system is a vulnerable part of the human body to radiation damage. Particularly, the lymphocyte, granulocyte, and platelet cells are the most radiosensitive of the blood elements, and monitoring their changes after exposure is regarded as a practical and recommended laboratory test to estimate radiation dose and injury. In this work we describe the HEMODOSE web tools, which are built upon solid physiological and pathophysiological understanding of mammalian hematopoietic systems, and rigorous coarse-grained biomathematical modeling and validation. Using single or serial granulocyte, lymphocyte, leukocyte, or platelet counts after exposure, these tools can estimate absorbed doses of adult victims very rapidly and accurately to assess the severity of BM radiation injury. Some patient data from historical accidents are utilized as examples to demonstrate the capabilities of these tools as a rapid point-of-care diagnostic or centralized high-throughput assay system in a large-scale radiological disaster scenario. HEMODOSE web tools establish robust correlations between the absorbed doses and victim's various types of blood cell counts not only in the early time window (1 or 2 days), but also in very late phase (up to 4 weeks) after exposure.

Hu, Shaowen↗

A Multiple Parameters Biodosimetry Tool with Various Blood Cell Counts - the Hemodose Approach

There continue to be important concerns about the possibility of the occurrence of acute radiation syndromes following nuclear and radiological terrorism or accidents that may result in mass casualties in densely populated areas. To guide medical personnel in their clinical decisions for effective medical management and treatment of the exposed individuals, biological markers are usually applied to examine radiation induced biological changes to assess the severity of radiation injury to sensitive organ systems. Among these the peripheral blood cell counts are widely used to assess the extent of radiation induced bone marrow injury. This is due to the fact that the hematopoietic system is the most vulnerable part of the human body to radiation damage. Particularly, the lymphocyte, granulocyte, and platelet cells are the most radiosensitive of the blood elements, and monitoring their changes after exposure is regarded as a practical and recommended laboratory test to estimate radiation dose and injury. Based upon years of physiological and pathophysiological investigation of mammalian hematopoietic systems, and rigorous coarse-grained bio-mathematical modeling and validation on species from mouse, to dog, monkey, and human, we have developed a set of software tools Hemodose, which can use single or serial granulocyte, lymphocyte, leukocyte, or platelet counts after exposure to estimate absorbed doses of adult victims very rapidly and accurately. Some patient data from historical accidents are utilized as examples to demonstrate the capabilities of these tools as a rapid point-of-care diagnostic or centralized high-throughput assay system in a large-scale radiological disaster scenario. Most significant to the improvement of national and local preparedness of a potential nuclear/radiological disaster, this HemoDose approach establishes robust correlations between the absorbed doses and victim's various types of blood cell counts not only in the early time window (1 or 2 days), but also in the very late phase (up to 4 weeks) after exposure.

Hu, Shaowen↗

ISOCS-based Extended In Situ Gamma Spectrometry Services Tool SuperISOCS for Waste Measurements - 20275

Canberra's In Situ Object Counting System (ISOCS) is an established tool used for gamma spectrometry. It allows physical representation of complex geometries and mathematical calculation of the calibration function while avoiding the need for radioisotope standards. ISOCS provides a large range of templates to generate the geometry for most nuclear waste components in nuclear facilities. Nonetheless, there are some complex geometries where the accuracy attainable is limited by the conventional ISOCS templates. An ISOCS-based extended in-situ Gamma Spectrometry service tool has been developed, based on the so-called SuperISOCS (SISOCS) software. This tool was developed to generate the most complex geometries. The principle of geometry construction is based on the composition of 8 different primitive geometries. The primitive object is able to be cut and to change a part of an object. An object is able to overlap to another one to form a more complex object. These functions refine the modelled geometry compared to the real objects and decrease the measurement systematic error because they allow the mathematical model to represent the real-life geometry more faithfully through reducing the approximations and simplifications that need to be made The algorithm of SISOCS is the same as that of standard ISOCS. A set of calculations was done in comparison with MCNP efficiency calculation and SISOCS results. 80 different models were compared between SISOCS and MCNP, with added 30 cases compared between ISOCS and SISOCS. The average differences were less than 2%, for all energy's efficiency calculations. In this paper we illustrate the benefits that this new tool can provide, in terms of improved accuracy for complex realistic geometries, through simulation of some example scenarios. Furthermore, we demonstrate, through SISOCS modelling of the real object data test, the comparison between SISOCS to ISOCS calculations for the same geometry. This paper describes the added functions that are easily employed using SISOCS. Our results show the potential application and benefit of this technique for real waste assay projects and the way to integrate the tool into the waste assay systems. (authors)

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗

Detector Characterization for Quantitative Spectral Radiography of Uranium Powder Samples

Pacific Northwest National Laboratory (PNNL) is developing a prototype spectral x-ray radiography (SpecX) non-destructive assay system for noninvasive material mass quantification of uranium-bearing powders. The SpecX system uses a custom material estimation algorithm, which requires a high-fidelity pixel-by-pixel detector response model for accurate quantification. We performed a detailed characterization and built a model of a High-Energy X-ray Imaging Technology (HEXITEC) CdTe pixelated spectral detector. Here we present the detector characterization which led to that model, including a determination of the pixel-wise energy calibration, the detection efficiency, the detector uniformity, the detector stability, and a comparison of methods to remove charge-shared events.

spectral X-ray radiography, pixelated detector, ca↗

Human in vitro metabolism of an environmental mixture of polycyclic aromatic hydrocarbons (PAH) found at the Portland Harbor Superfund Site

Polycyclic aromatic hydrocarbons (PAHs) are widespread environmental contaminants that pose health risks to humans. Toxicity testing approaches of PAHs have evolved from traditional rodent models to New Approach Methodologies (NAMs), such as high-throughput screening in zebrafish, enabling rapid evaluation of chemical hazards. However, translating toxicity findings from laboratory systems to humans remains difficult due to complexity and species-specific differences. Chemical dosimetry modeling offers a quantitative framework to bridge this gap, but its accuracy depends on robust knowledge of PAH metabolism. The objective of this study was to measure human metabolism rates of Supermix-10, the ten most abundant PAHs found at the Portland Harbor Superfund Site, to support development of human pharmacokinetic models. We incubated individual PAHs from Supermix-10 in pooled human liver microsomes and quantified parent PAH disappearance using high-performance liquid chromatography (HPLC) with UV and florescent detection. To assess the potential of mixture interactions, we also measured metabolism of all 10 compounds in an equimolar mixture and compared rates of parent disappearance to those observed for individual PAHs. All Supermix-10 PAHs demonstrated rapid parent compound disappearance in human hepatic microsomes. PAHs grouped into three metabolism patterns: high metabolism rates and capacity (2-methylnaphthalene, acenaphthylene, fluorene, naphthalene), high affinity metabolism that rapidly achieves low-level saturation (benzo[a]anthracene, chrysene), and moderate metabolism rates and capacity (fluoranthene, pyrene, retene, phenanthrene). Smaller PAHs exhibited faster metabolism, and higher metabolism rates correlated inversely with molecular weight. When incubated in an equimolar mixture, Supermix-10 demonstrated significantly slower metabolism (47–89 %) compared to metabolism of individual PAHs at the same concentration. These findings enhance our understanding of PAH metabolism in humans and demonstrate significant mixture interactions under the conditions tested. Furthermore, our findings offer insights into the metabolic behavior of Supermix-10 and provide critical metabolism rate data to support the development of physiological based pharmacokinetic (PBPK) models. Dosimetry models can translate PAH chemical dosimetry from high-throughput testing platforms, like zebrafish and cellular system assays, to human exposures enhancing the accuracy and reliability of PAH risk assessments.

2-methylnaphthalene↗

Antibody dynamics to SARS‐CoV‐2 in asymptomatic COVID‐19 infections

Abstract Background The missing asymptomatic COVID‐19 infections have been overlooked because of the imperfect sensitivity of the nucleic acid testing (NAT). Globally understanding the humoral immunity in asymptomatic carriers will provide scientific knowledge for developing serological tests, improving early identification, and implementing more rational control strategies against the pandemic. Measure Utilizing both NAT and commercial kits for serum IgM and IgG antibodies, we extensively screened 11 766 epidemiologically suspected individuals on enrollment and 63 asymptomatic individuals were detected and recruited. Sixty‐three healthy individuals and 51 mild patients without any preexisting conditions were set as controls. Serum IgM and IgG profiles were further probed using a SARS‐CoV‐2 proteome microarray, and neutralizing antibody was detected by a pseudotyped virus neutralization assay system. The dynamics of antibodies were analyzed with exposure time or symptoms onset. Results A combination test of NAT and serological testing for IgM antibody discovered 55.5% of the total of 63 asymptomatic infections, which significantly raises the detection sensitivity when compared with the NAT alone (19%). Serum proteome microarray analysis demonstrated that asymptomatics mainly produced IgM and IgG antibodies against S1 and N proteins out of 20 proteins of SARS‐CoV‐2. Different from strong and persistent N‐specific antibodies, S1‐specific IgM responses, which evolved in asymptomatic individuals as early as the seventh day after exposure, peaked on days from 17 days to 25 days, and then disappeared in two months, might be used as an early diagnostic biomarker. 11.8% (6/51) mild patients and 38.1% (24/63) asymptomatic individuals did not produce neutralizing antibody. In particular, neutralizing antibody in asymptomatics gradually vanished in two months. Conclusion Our findings might have important implications for the definition of asymptomatic COVID‐19 infections, diagnosis, serological survey, public health, and immunization strategies.

Lei, Qing↗

The anaerobic fungus Caecomyces churrovis produces H2 via a non-3 bifurcating NADH-dependent enzyme complex

Anaerobic fungi (AF) decompose lignocellulose-based biomass into fermentable sugars through the production of powerful biomass-degrading enzymes. AF are unusual among fungi in that they generate energy via hydrogenosomes, which are also associated with the release of H2 though yet unknown metabolic mechanisms. In particular, it remains unclear how NAD(P)+ is regenerated within hydrogenosomes and how H2 is formed. Here, we reveal the molecular mechanism for hydrogenosomal H2 production in the AF strain C. churrovis by combining genomic search, proteomic analysis, and enzymology. Our enzyme assays on the large organelle fraction of C. churrovis revealed the activity of H2:NAD+ oxidoreductase but not pyruvate:ferredoxin oxidoreductase activity. We identified genes encoding [FeFe] hydrogenase (Hyd) and NADH dehydrogenase subunits E and F (NuoE, NuoF) in C. churrovis, and confirmed their expression in the isolated hydrogenosomal fractions by proteomic analysis. Combining the individually purified proteins, we found that the assay system consisting of Hyd-Strep and NuoEF-Strep reduced NAD+ with H2. Furthermore, this system formed H2 directly from NADH independent of ferredoxin, functioning as a non-bifurcating NADH-dependent enzyme rather than an electron-bifurcating enzyme. We identified homologs of hydrogenosomal NuoE, NuoF, and Hyd in many other AF, indicating this pathway is widely conserved among the early-branching AF. This work demonstrates the existence of a non-bifurcating NADH-dependent enzyme complex in eukaryotes. Moreover, this complex could be a target for controlling AF H2 production and altering fungal metabolism.

fungi↗

Measurement results for 60Co Assay baskets H5, H9, H14, H15, A4, and B3 (C003, C001, C005, C022, C021, C020, C032, C033, C038, C035, C036, C039, C053, C054, C055, C065, C066, & C067)

Eighteen cobalt targets (C003, C001, C005, C022, C021, C020, C032, C033, C038, C035, C036, C039, C053, C054, C055, C065, C066, & C067) were assayed on July 28-29, 2020, following irradiation in ATR cycle 168A. The average specific activity of each target was determined by using the cobalt assay system located in the ATR canal following procedure ACMM-3610.

07 ISOTOPE AND RADIATION SOURCES↗

TCF Base Technology-Specific Final Report: Engineering Enzymes for Crystalline PET Substrate

The primary objective of this project was to develop a new polyethylene terephthalate (PET) hydrolase enzyme to depolymerize relevant PET substrates for Birch Biosciences, using high-throughput protein expression, purification, and assaying systems and machine learning-guided enzyme design. As a secondary project objective, we also aimed to develop a more energy-efficient ethylene glycol (EG) recovery strategy relative to distillation.

09 BIOMASS FUELS↗