Search NASA⌕ Search

SEARCH · Search NASA

Results for “Assay systems”

Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 523 records · Page 29
All sourcesNASA NTRSDOE OSTI
DOE OSTI2020

Predicting Biological Cleanliness: An Empirical Bayes Approach for Spacecraft Bioburden Accounting

To comply with the international planetary protection policy set forth by the Committee on Space Research and NASA Agency level requirements, spacecraft destined to biologically sensitive planetary bodies have to minimize terrestrial biological contamination. Analysis, testing and inspection are the standard forward verification activities that are used to demonstrate compliance with the biological contamination requirements. For testing of spacecraft surface areas, a swab or wipe sample is collected from surfaces prior to last access and subsequently processed in the lab using NASA Approved Planetary Protection Methods for Culture Based Assays. Raw data resulting from this assay is then statistically treated employing a mathematical paradigm stemming from the 1970’s Viking Lander Project to generate the bioburden density and total microbial bioburden present. This standard approach arbitrarily accounts for error and provides an upper conservative bound as it reports the maximum number of spores estimated to be present on flight hardware surfaces. A bioburden density estimate factors in the following variables: the observed bioburden count, representative volume processed, sampling efficiencies. Notably, to account for error in the approach, a 0 observed count is arbitrarily changed to a count of 1 for each hardware grouping. The data generated by spacecraft bioburden verification campaigns in the past have resulted in <80% of wipes and <90% of swabs containing a bioburden count of 0. As such, having a robust and well documented statistical approach for dealing with the probability of low incident rates is necessary to be able to estimate spacecraft bioburden. Being able to statistically describe the bioburden distribution and associated confidence level is a gamechanger for the development of bioburden allocations during mission design and will allow for tighter management of risk throughout spacecraft build. Thus, Empirical Bayes statistical approach was evaluated to estimate the microbial bioburden on spacecraft to mitigate the aforementioned mathematical concerns and provide a probabilistic bioburden distribution of the flight hardware surface. For application of this approach to performing bioburden calculations, a range of non-informative prior assumptions on hardware surfaces are explored for Bayesian analyses while informative priors using posterior distributions from prior assays are utilized for Empirical Bayes analyses. Several non-informative priors are currently under investigation to assess fitness including use of these priors to serve as a foundation to build off of NASA specification values or a basis of risk to account for unknowns during the integration and testing process. Informative priors under consideration are generated using sampled bioburden values from hardware originating within like processing environments (e.g. vendor cleaning process or similar assembly process), temporal spacecraft status events as a prediction for hardware cleanliness of future samples, and heritage system bioburden actuals to predict allocation for subsequent missions. Informative priors and probabilistic bioburden distributions are then validated using data sets from the Mars Exploration Rover, Mars Science Laboratory, and InSight missions. Using Empirical Bayes approach to generate a probabilistic bioburden distribution as demonstrated through mission use cases provides a valid approach for use in the end-to-end requirements verification process.

97 MATHEMATICS AND COMPUTING↗
DOE OSTI2024

Bayesian Framework for Bioburden Density Estimation in Planetary Protection

To comply with the international planetary protection policy set forth by the Committee on Space Research and NASA Agency level requirements, spacecraft destined to biologically sensitive planetary bodies have to minimize terrestrial biological contamination. Analysis, testing and inspection are the standard forward verification activities that are used to demonstrate compliance with the biological contamination requirements. For testing of spacecraft surface areas, a swab or wipe sample is collected from surfaces prior to last access and subsequently processed in the lab using NASA Approved Planetary Protection Methods for Culture Based Assays. Raw data resulting from this assay is then statistically treated employing a mathematical paradigm stemming from the 1970’s Viking Lander Project to generate the bioburden density and total microbial bioburden present. This standard approach arbitrarily accounts for error and provides an upper conservative bound as it reports the maximum number of spores estimated to be present on flight hardware surfaces. A bioburden density estimate factors in the following variables: the observed bioburden count, representative volume processed, sampling efficiencies. Notably, to account for error in the approach, a 0 observed count is arbitrarily changed to a count of 1 for each hardware grouping. The data generated by spacecraft bioburden verification campaigns in the past have resulted in <80% of wipes and <90% of swabs containing a bioburden count of 0. As such, having a robust and well documented statistical approach for dealing with the probability of low incident rates is necessary to be able to estimate spacecraft bioburden. Being able to statistically describe the bioburden distribution and associated confidence level is a gamechanger for the development of bioburden allocations during mission design and will allow for tighter management of risk throughout spacecraft build. Thus, Empirical Bayes statistical approach was evaluated to estimate the microbial bioburden on spacecraft to mitigate the aforementioned mathematical concerns and provide a probabilistic bioburden distribution of the flight hardware surface. For application of this approach to performing bioburden calculations, a range of non-informative prior assumptions on hardware surfaces are explored for Bayesian analyses while informative priors using posterior distributions from prior assays are utilized for Empirical Bayes analyses. Several non-informative priors are currently under investigation to assess fitness including use of these priors to serve as a foundation to build off of NASA specification values or a basis of risk to account for unknowns during the integration and testing process. Informative priors under consideration are generated using sampled bioburden values from hardware originating within like processing environments (e.g. vendor cleaning process or similar assembly process), temporal spacecraft status events as a prediction for hardware cleanliness of future samples, and heritage system bioburden actuals to predict allocation for subsequent missions. Informative priors and probabilistic bioburden distributions are then validated using data sets from the Mars Exploration Rover, Mars Science Laboratory, and InSight missions. Using Empirical Bayes approach to generate a probabilistic bioburden distribution as demonstrated through mission use cases provides a valid approach for use in the end-to-end requirements verification process.

97 - MATHEMATICS AND COMPUTING↗
NASA NTRS2004

Molecular Microbial Analyses of the Mars Exploration Rovers Assembly Facility

During space exploration, the control of terrestrial microbes associated with robotic space vehicles intended to land on extraterrestrial solar system bodies is necessary to prevent forward contamination and maintain scientific integrity during the search for life. Microorganisms associated with the spacecraft assembly environment can be a source of contamination for the spacecraft. In this study, we have monitored the microbial burden of air samples of the Mars Exploration Rovers' assembly facility at the Kennedy Space Center utilizing complementary diagnostic tools. To estimate the microbial burden and identify potential contaminants in the assembly facility, several microbiological techniques were used including culturing, cloning and sequencing of 16S rRNA genes, DNA microarray analysis, and ATP assays to assess viable microorganisms. Culturing severely underestimated types and amounts of contamination since many of the microbes implicated by molecular analyses were not cultivable. In addition to the cultivation of Agrobacterium, Burkholderia and Bacillus species, the cloning approach retrieved 16s rDNA sequences of oligotrophs, symbionts, and y-proteobacteria members. DNA microarray analysis based on rational probe design and dissociation curves complemented existing molecular techniques and produced a highly parallel, high resolution analysis of contaminating microbial populations. For instance, strong hybridization signals to probes targeting the Bacillus species indicated that members of this species were present in the assembly area samples; however, differences in dissociation curves between perfect-match and air sample sequences showed that these samples harbored nucleotide polymorphisms. Vegetative cells of several isolates were resistant when subjected to treatments of UVC (254 nm) and vapor H202 (4 mg/L). This study further validates the significance of non-cultivable microbes in association with spacecraft assembly facilities, as our analyses have identified several non-cultivable microbes likely to contaminate the surfaces of spacecraft hardware.

microbial diversity MER 16S rDNA DNA microarray sp↗
DOE OSTI2026

Deimos: HALEU TRISO Heated Critical Experiment Data

Deimos was the first critical experiment using high-assay low-enriched uranium (HALEU) TRistructural ISOtropic (TRISO) fuel in over 40 years. HALEU TRISO is the desired fuel form for many of the advanced reactor designs in development; however, very little experimental data are available for this fuel type. Deimos was designed to utilize existing HALEU TRISO fuel in a large graphite moderator to obtain nuclear and reactor physics data to fill the gaps surrounding this fuel type and enrichment. In addition to cold critical data, three separate heated experiments were conducted to measure the temperature reactivity coefficient for this type of system. These measured coefficients were then compared to simulated coefficients to a first level order of fidelity. This comparison showed very good agreement for the experiment where only the inner core was heated and good agreement for the other two configurations, which included heating portions of the outer core. Less agreement when the outer core was heated is attributed to potential heating in the beryllium reflector, which has a positive temperature reactivity coefficient and was unaccounted for in the first-order models. Future heated experiments with Deimos will include temperature monitoring of the beryllium reflector to account for beryllium heating in the simulations.

22 GENERAL STUDIES OF NUCLEAR REACTORS↗
DOE OSTI2023

Building a Computational and Experimental Rapid Response Pipeline to Counter the Coronavirus Disease 2019 Outbreak and Emerging Biothreats

The COVID-19 pandemic underscored the promise of monoclonal antibody-based prophylactic and therapeutic drugs, especially where protective candidates can be rapidly identified or developed for emerging biothreats and escape variants. Current cutting-edge technologies for this purpose still rely on pathogen-exposed convalescent volunteers and a large screening effort to find a proverbial needle in a haystack. Computational design of protective antibodies based on pre-existing templates skips those requirements and allows for greater control over the breadth and target epitope, while also co-optimizing for potency and developability or other biophysical characteristics. We approached this problem by building and expanding an in vitro experimental rapid antibody production and characterization pipeline to support development of an autonomous, closed loop, active learning software system based on structural simulation and ground truth experimental data to design and evaluate antibody antigen interactions. Starting from early in the pandemic, we targeted SARS-CoV-2, the causative agent of COVID-19, by re-purposing neutralizing antibodies against SARS-CoV-1 that had been identified in the wake of that outbreak in the early 2000’s. We successfully re-targeted three different anti-SARS-CoV-1 antibodies to neutralize SARS-CoV-2 in vitro, where the antibodies were generated externally and tested through conventional binding and neutralization assays internally or with collaborators. As antibodies were identified from the blood of humans infected with SARS-CoV-2, we shifted to collaborate with academic partners to develop improved versions of their human-derived antibodies. This work reached its most important stage in rapid response to the emergence of the Omicron variant of concern (VOC) in late 2021. In a matter of weeks, enabled by on demand innovation to our screening pipeline, we computationally designed and experimentally characterized derivative antibodies of COV2-2130, one of two antibodies from Vanderbilt that form the basis of the AstraZeneca Evusheld prophylactic drug product. This drug product suffers a serious loss of efficacy against Omicron BA.1 and BA.1.1, the first Omicron strains. Due to tight integration of computational design and experimental evaluation, we were able to identify a pair of designs with potent neutralization of the main targets Omicron BA.1 and BA.1.1; but also the earlier Delta variant, and subsequent Omicron strains including BA.2, BA.4, BA.5, and BA.2.75, demonstrating that our multi-target design process can, by its nature, produce robust antibody designs that strictly improve over the parental antibody. These results, recognized by a 2022 Director’s Science and Technology award, have enabled the follow-on GUIDE program, to commence in FY23. While earlier design campaigns were substantially outsourced, we have engineered better and faster processes internally to better compliment, calibrate, and speed computational designs. As part of the follow-on GUIDE program, we will stand up a rapid and high-throughput antibody production and characterization facility staffed with the expertise and capabilities to foster our current collaboration across PLS and ENG as well as other partnerships toward computational design of biologics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Uranium Oxide Elutriation at Higher Flow Rates and Longer Times

This work supports the recovery of high-assay low-enriched uranium (i.e., U enriched in 235 U to >5% and <20%) from used nuclear fuel. One step includes elutriating triuranium octoxide (U 3 O 8 ) from a fluidized bed of alumina (Al 2 O 3 ) that fills the main reaction chamber. The experiments outlined in this report assessed the effects of the time and flow rate on the elutriation of U 3 O 8 powder from the reaction vessel. Testing at Oak Ridge National Laboratory (ORNL) was performed in a one-fourth scale experimental apparatus based on a pilot-scale system, which is the Material Recovery Pilot Plant (MRPP) at Idaho National Laboratory (INL). The experimental system comprises a supply of pressurized nitrogen (N 2 ), a gas heater, the reaction vessel, a knock-out pot, and a water trap where elutriated particles are collected for sampling. An Al 2 O 3 fluidized bed is contained within the reaction vessel. A bed of Al 2 O 3 with a nominal size of 180 µm and U 3 O 8 with a nominal size of 7 µm was fluidized at 2 different flow rates so the U 3 O 8 physically separated from the bed. The apparatus was simple and had no valves or sharp bends downstream from the reactor. The flow exited the reactor through a side port at the top. The flow passed through a knock-out vessel before the flow was passed through a bubbler to capture the U 3 O 8 . Seven tests were completed with U 3 O 8 using an elutriating gas of N 2 heated to 200°C. Tests were performed with bed loadings that comprise 25.0 g of U 3 O 8 , or 3.6 wt% of the bed mass. Except for one test performed at 12 SLPM, the gas flow rate through the bed for the other experiments was set at 10 SLPM as measured by the mass flow meter. Linear velocities through the bed were calculated as 28 and 34 cm/s, or 0.9 and 1.1 ft·s -1 . The duration of the tests ranged from 20 to 180 min, and one test had a flow cycled between 0 and 5 SLPM every 5 min for 40 min. The testing successfully demonstrated elutriation of U 3 O 8 from the fluidized bed as up to a 100% of the U 3 O 8 was transferred out of the bed. The removal of U 3 O 8 had a logarithmic dependence on flow rate. The design changes made the apparatus easier to clean, and the mass balances were better than those used in previous studies. Some elutriated Al 2 O 3 collected in the tubing that connected the reactor to the knock-out pot, but the amount of Al 2 O 3 was minimal for 10 SLPM. The flow at 12 SLPM showed considerable transport of Al 2 O 3 bed material into the reaction-vessel headspace and the transfer line to the knock-out pot.

11 NUCLEAR FUEL CYCLE AND FUEL MATERIALS↗
DOE OSTI2022

Scale-dependent enhancement of productivity and stability in xenic Nannochloropsis cultures

Although bacterial amendments can enhance algal productivity and stability, the development of algal-bacteria consortia for commercial-scale utilization is limited. Here, for this work, we used an established high-throughput approach to generate algal-bacteria consortia, and tested consortia performance at spatial scales from microplates to 320 L raceway ponds. We used both lab and field-reared strains of Nannochloropsis oceanica to build consortia. In some experiments, we imposed environmental perturbations to test the ability of bacteria to enhance algal culture stability. In repeated assays at the scale of well plates, flasks, and bioreactors, strong effects of bacterial amendments on N. oceanica were observed. These effects were most dramatic when cultures experienced stressors such as temperature perturbations or removal of CO 2 augmentation. Isolates that were advanced for field testing included species in the genera Algoriphagus, Oceanicaulis, and Marinobacter. When consortia were generated in the field, positive effects of bacterial amendments were not observed. The amended bacteria were outcompeted, and bacterial community composition across treatments converged after the first grow out. These results highlight the complexity of using consortia in open systems, where interactions between the existing bacterial community, inoculated bacteria, and changing environmental conditions are layered upon other differences in scale and cultivation regimes. Moreover, functionally redundant bacteria are likely present in the field. Following this work, we hypothesize that tight interactions (e.g., obligate relationships between partners) will scale more predictably to outdoor systems. We suggest relying on true synthetic ecological approaches in which the relationships between bacteria and algae partners are well understood, or synthetic ecological approaches coupled with high throughput approaches to design and test consortia. We also recommend future work to examine the effect of algae-bacteria inoculation ratios on productivity and stability, track dynamics of partners through time, and manage ponds to retain beneficial symbioses.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Polyethylene Glycol Crowder’s Effect on Enzyme Aggregation, Thermal Stability, and Residual Catalytic Activity

Protein stability and performance in various natural and artificial systems incorporating many other macromolecules for therapeutic, diagnostic, sensor, and biotechnological applications attract increasing interest with the expansion of these technologies. Here we address the catalytic activity of lysozyme protein (LYZ) in the presence of a polyethylene glycol (PEG) crowder in a broad range of concentrations and temperatures in aqueous solutions of two different molecular mass PEG samples (Mw = 3350 and 10000 g/mol). The phase behavior of PEG–protein solutions is examined by using dynamic light scattering (DLS) and small-angle X-ray scattering (SAXS), while the enzyme denaturing is monitored by using an activity assay (AS) and circular dichroism (CD) spectroscopy. Molecular dynamic (MD) simulations are used to illustrate the effect of PEG concentration on protein stability at high temperatures. The results demonstrate that LYZ residual activity after 1 h incubation at 80 °C is improved from 15% up to 55% with the addition of PEG. The improvement is attributed to two underlying mechanisms. (i) Primarily, the stabilizing effect is due to the suppression of the enzyme aggregation because of the stronger PEG–protein interactions caused by the increased hydrophobicity of PEG and lysozyme at elevated temperatures. (ii) The MD simulations showed that the addition of PEG to some degree stabilizes the secondary structures of the enzyme by delaying unfolding at elevated temperatures. The more pronounced effect is observed with an increase in PEG concentration. This trend is consistent with CD and AS experimental results, where the thermal stability is strengthened with increasing of PEG concentration and molecular mass. Overall, the results show that the highest stabilizing effect is approached at the critical overlap concentration of PEG.

77 NANOSCIENCE AND NANOTECHNOLOGY↗
DOE OSTI2022

Altered properties and structures of root exudate polysaccharides in a root hairless mutant of barley

Root exudates and rhizosheaths of attached soil are important features of growing roots. To elucidate factors involved in rhizosheath formation, wild-type (WT) barley (Hordeum vulgare L. cv. Pallas) and a root hairless mutant, bald root barley (brb), were investigated with a combination of physiological, biochemical, and immunochemical assays. When grown in soil, WT barley roots bound ~5-fold more soil than brb per unit root length. High molecular weight (HMW) polysaccharide exudates of brb roots had less soil-binding capacity than those of WT root exudates. Carbohydrate and glycan monoclonal antibody analyses of HMW polysaccharide exudates indicated differing glycan profiles. Relative to WT plants, root exudates of brb had reduced signals for arabinogalactan-protein (AGP), extensin, and heteroxylan epitopes. In contrast, the root exudate of 2-week-old brb plants contained ~25-fold more detectable xyloglucan epitope relative to WT. Root system immunoprints confirmed the higher levels of release of the xyloglucan epitope from brb root apices and root axes relative to WT. Epitope detection with anion-exchange chromatography indicated that the increased detection of xyloglucan in brb exudates was due to enhanced abundance of a neutral polymer. Conversely, brb root exudates contained decreased amounts of an acidic polymer, with soil-binding properties, containing the xyloglucan epitope and glycoprotein and heteroxylan epitopes relative to WT. We, therefore, propose that, in addition to physically structuring soil particles, root hairs facilitate rhizosheath formation by releasing a soil-binding polysaccharide complex.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Comparative analysis of the functional properties of human and mouse ferroportin

Ferroportin (Fpn)—expressed at the plasma membrane of macrophages, enterocytes, and hepatocytes—mediates the transfer of cellular iron into the blood plasma. Under the control of the iron-regulatory hormone hepcidin, Fpn serves a critical role in systemic iron homeostasis. Although we have previously characterized human Fpn, a great deal of research in iron homeostasis and disorders uses mouse models. By way of example, the flatiron mouse, a model of classical ferroportin disease, bears the mutation H32R in Fpn and is characterized by systemic iron deficiency and macrophage iron retention. The flatiron mouse also appears to exhibit a manganese phenotype, raising the possibility that mouse Fpn serves a role in manganese metabolism. At odds with this observation, we have found that human Fpn does not transport manganese, so we considered the possibility that a species difference could explain this discrepancy. We tested the hypothesis that mouse but not human Fpn can transport manganese and performed a comparative analysis of mouse and human Fpn. We examined the functional properties of human Fpn, mouse Fpn, and mutant mouse Fpn by using radiotracer assays in RNA-injected Xenopus oocytes. We found that neither mouse nor human Fpn transports manganese. Mouse and human Fpn share identical properties with respect to substrate profile, calcium dependence, optimal pH, and hepcidin sensitivity. We have also demonstrated that Fpn is not an ATPase pump. Our findings validate the use of mouse models of ferroportin function in iron homeostasis and disease.

Cell Biology↗
DOE OSTI2023

Expression quantitative trait loci mapping identified PtrXB38 as a key hub gene in adventitious root development in Populus

Summary Plant establishment requires the formation and development of an extensive root system with architecture modulated by complex genetic networks. Here, we report the identification of the PtrXB38 gene as an expression quantitative trait loci (eQTL) hotspot, mapped using 390 leaf and 444 xylem Populus trichocarpa transcriptomes. Among predicted targets of this trans ‐eQTL were genes involved in plant hormone responses and root development. Overexpression of PtrXB38 in Populus led to significant increases in callusing and formation of both stem‐born roots and base‐born adventitious roots. Omics studies revealed that genes and proteins controlling auxin transport and signaling were involved in PtrXB38‐mediated adventitious root formation. Protein–protein interaction assays indicated that PtrXB38 interacts with components of endosomal sorting complexes required for transport machinery, implying that PtrXB38‐regulated root development may be mediated by regulating endocytosis pathway. Taken together, this work identified a crucial root development regulator and sheds light on the discovery of other plant developmental regulators through combining eQTL mapping and omics approaches.

54 ENVIRONMENTAL SCIENCES↗
NASA NTRS2012

Improved Whole-Blood-Staining Device

Dramatic improvements have been made in NASA s Whole Blood Staining Device (WBSD) since it was last described in "Whole-Blood-Staining Device," NASA Tech Briefs, Vol. 23, No. 10 (October 1999), page 64. The new system has a longer shelf life, a simpler and more effective operational procedure, improved interface with instrumentation, and shorter processing time. More specifically, the improvements have targeted bag and locking clip materials, sampling ports, and air pocket prevention. The WBSD stains whole blood collected during spaceflight for subsequent flow cytometric analysis. In short, the main device stains white blood cells by use of monoclonal antibodies conjugated to various fluorochromes, followed by lysing and fixing of the cells by use of a commercial reagent that has been diluted according to NASA safety standards. This system is compact, robust, and does not require electric power, precise mixing, or precise incubation times. Figure 1 depicts the present improved version for staining applications, which is a poly(tetrafluoroethylene) bag with a Luer-lock port and plastic locking clips. An InterLink (or equivalent) intravenous- injection port screws into the Luer-lock port. The inflatable/collapsible nature of the bag facilitates loading and helps to minimize the amount of air trapped in the fully loaded bag. Some additional uses have been identified for the device beyond whole blood staining. The WBSD has been configured for functional assays that require culture of live cells by housing sterile culture media, mitogens, and fixatives prior to use [Figure 2(a)]. Simple injection of whole blood allows cell-stimulation culture to be performed in reduced gravity conditions, and product stabilization prior to storage, while protecting astronauts from liquid biohazardous materials. Also, the improved WBSD has reconstituted powdered injectable antibiotics by mixing them with diluent liquids [Figure 2(b)]. Although such mixing can readily be performed on Earth by shaking in glass vials, it cannot readily be performed this way in outer space without entraining air bubbles. The present device can be preloaded with the powder and diluent(s) in separate compartments. The powder and diluent( s) can be mixed, without introducing air bubbles, by removing the clip(s), then shaking. This use of the device could also be advantageous in terrestrial applications because it maintains the isolation of the constituents until the time of use.

Sams, Clarence F.↗
DOE OSTI2020

Clinically applicable 53-Gene prognostic assay predicts chemotherapy benefit in gastric cancer: A multicenter study

BACKGROUND:We previously established a 53-gene prognostic signature for overall survival (OS) of gastric cancer patients. This retrospective multi-center study aimed to develop a clinically applicable gene expression detection assay and to investigate the prognostic value of this signature. METHODS:A TCGA gastric adenocarcinoma cohort (TCGA-STAD) was used for comparing 53-gene signature with other gene signatures. A high-throughput mRNA hybridization gene expression assay was developed to quantify the expression of 53-genes in formalin-fixed paraffin-embedded tissues of 540 patients enrolled from three hospitals. 180 patents were randomly selected from two hospitals to build a prognostic prediction model based on the 53-gene signature using leave-p-out (one-third out) cross-validation method together with Cox regression and Kaplan-Meier analysis, and the model was assessed on three validation cohorts. FINDINGS:In the evaluation phase, studies based on TCGA-STAD showed that the 53-gene signature was significantly superior to other three prognostic signatures and was independent of TCGA molecular subtypes and clinical factors. For clinical validation and utility, the prognostic scores were generated using the newly developed assay, which was reliable and sensitive, in 100 sampling training sets and were significantly associated with OS in 100 sampling validation sets. The scores were significantly associated with OS in three independent and combined validation cohorts, and in patients with stages II and III/IV. The multivariate Cox regression demonstrated that the prognostic power of the score was independent of clinical factors, consistent with those findings in the TCGA dataset. Finally, patients with good prognostic scores exhibited significantly a better 5-year OS rate from adjuvant FOLFOX chemotherapy after surgery than from other chemotherapies. INTERPRETATION:The 53-gene prognostic score system is clinically applicable for predicting the OS of patients independent of clinical factors in gastric cancers, which could also be a promising predictive biomarker for FOLFOX regimen. FUNDING:Chinese National Science and Technology, National Natural Science Foundation and Natural Science Foundation of Jiangsu Province.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2025

Breaking the reproducibility barrier with standardized protocols for plant–microbiome research

Inter-laboratory replicability is crucial yet challenging in microbiome research. Leveraging microbiomes to promote soil health and plant growth requires understanding underlying molecular mechanisms using reproducible experimental systems. In a global collaborative effort involving five laboratories, we aimed to help advance reproducibility in microbiome studies by testing our ability to replicate synthetic community assembly experiments. Our study compared fabricated ecosystems constructed using two different synthetic bacterial communities, the model grass Brachypodium distachyon, and sterile EcoFAB 2.0 devices. All participating laboratories observed consistent inoculum-dependent changes in plant phenotype, root exudate composition, and final bacterial community structure, where Paraburkholderia sp. OAS925 could dramatically shift microbiome composition. Comparative genomics and exudate utilization linked the pH-dependent colonization ability of Paraburkholderia, which was further confirmed with motility assays. The study provides detailed protocols, benchmarking datasets, and best practices to help advance replicable science and inform future multi-laboratory reproducibility studies.

Novak, Vlastimil↗
DOE OSTI2022

Nuclide inventory validation: radiochemical assay data quality and modeling challenges in benchmark models development

Oak Ridge National Laboratory is conducting radiochemical assay experiments, using high-precision analytical protocols validated with a comprehensive quality assurance plan, to expand the nuclide inventory validation basis for high burnup spent nuclear fuel. Preliminary measurement data for key actinides and fission products in two pressurized water reactor spent fuel samples are being used to investigate the impact of measurement data uncertainty on the sample burnup estimation. These measurement data are also being used to examine the impact of assumptions applied when developing best-estimate computational models to simulate fuel irradiation history. The simulations are being performed using depletion capabilities in the SCALE code system. Comparison of calculated and measured nuclide concentrations shows good agreement for the considered nuclides. (authors)

11 NUCLEAR FUEL CYCLE AND FUEL MATERIALS↗
DOE OSTI2022

Spatiotemporal analysis identifies ABF2 and ABF3 as key hubs of endodermal response to nitrate

Nitrate is a nutrient and a potent signal that impacts global gene expression in plants. However, the regulatory factors controlling temporal and cell type–specific nitrate responses remain largely unknown. We assayed nitrate-responsive transcriptome changes in five major root cell types of the Arabidopsis thaliana root as a function of time. We found that gene-expression response to nitrate is dynamic and highly localized and predicted cell type–specific transcription factor (TF)–target interactions. Among cell types, the endodermis stands out as having the largest and most connected nitrate-regulatory gene network. ABF2 and ABF3 are major hubs for transcriptional responses in the endodermis cell layer. We experimentally validated TF–target interactions for ABF2 and ABF3 by chromatin immunoprecipitation followed by sequencing and a cell-based system to detect TF regulation genome-wide. Validated targets of ABF2 and ABF3 account for more than 50% of the nitrate-responsive transcriptome in the endodermis. Moreover, ABF2 and ABF3 are involved in nitrate-induced lateral root growth. Our approach offers an unprecedented spatiotemporal resolution of the root response to nitrate and identifies important components of cell-specific gene regulatory networks.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

SuFEx-enabled high-throughput medicinal chemistry for developing potent tamoxifen analogs as Ebola virus entry inhibitors

Ebola virus (EBOV) causes severe hemorrhagic fever with a high mortality rate in humans. In acute infection, an abnormal immune response results in excessive inflammatory cytokines and uncontrolled systemic inflammation that can result in organ damage and multi-organ failure. While vaccines and monoclonal antibody therapies are available, there is an urgent need for effective small-molecule antivirals against EBOV. Here, we report on the optimization of tamoxifen, an EBOV-glycoprotein (GP) binder that inhibits viral entry, using our Sulfur-Fluoride Exchange (SuFEx) click chemistry-based high-throughput medicinal chemistry (HTMC) strategy. Using a “Direct-to-Biology” approach, we generated a focused library of 2,496 tamoxifen analogs overnight and screened them in a cell-based pseudo-EBOV infection assay. The HTMC workflow enabled the development of a potent EBOV entry inhibitor with submicromolar EC 50 cellular antiviral activity and more than 50-fold improvement in binding affinity against EBOV-GP compared to the parent compound. Our findings underscore the use of SuFEx-enabled HTMC for rapidly generating and assessing potential therapeutic candidates against viral and immune-mediated diseases in a cell-based assay.

Immunology↗