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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.

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At least 19 records
All sourcesNASA NTRSDOE OSTI
NASA NTRS2004

Novel Sample-handling Approach for XRD Analysis with Minimal Sample Preparation

Sample preparation and sample handling are among the most critical operations associated with X-ray diffraction (XRD) analysis. These operations require attention in a laboratory environment, but they become a major constraint in the deployment of XRD instruments for robotic planetary exploration. We are developing a novel sample handling system that dramatically relaxes the constraints on sample preparation by allowing characterization of coarse-grained material that would normally be impossible to analyze with conventional powder-XRD techniques.

Sarrazin, P.↗
DOE OSTI2019

Portable Sample Preparation For Chemical and Biological Analysis

Preparing samples for chemical or biological analysis is often time consuming and a critical step function to assure an accurate analysis. This invention provides an automated, hands-free, highly accurate device to prepare industrial, chemical, or biological samples for instrumental analysis methods.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2022

Modular automated bottom-up proteomic sample preparation for high-throughput applications

Manual proteomic sample preparation methods limit sample throughput and often lead to poor data quality when thousands of samples must be analyzed. Automated liquid handler systems are increasingly used to overcome these issues for many of the sample preparation steps. Here, we detail a step-by-step protocol to prepare samples for bottom-up proteomic analysis for Gram-negative bacterial and fungal cells. The full modular protocol consists of three optimized protocols to: (A) lyse Gram-negative bacteria and fungal cells; (B) quantify the amount of protein extracted; and (C) normalize the amount of protein and set up tryptic digestion. These protocols have been developed to facilitate rapid, low variance sample preparation of hundreds of samples, be easily implemented on widely-available Beckman-Coulter Biomek automated liquid handlers, and allow flexibility for future protocol development. By using this workflow 50 micrograms of protein from 96 samples can be prepared for tryptic digestion in under an hour. We validate these protocols by analyzing 47 Pseudomonas putida and Rhodosporidium toruloides samples and show that this modular workflow provides robust, reproducible proteomic samples for high-throughput applications. The expected results from these protocols are 94 peptide samples from Gram-negative bacterial and fungal cells prepared for bottom-up quantitative proteomic analysis without the need for desalting column cleanup and with protein relative quantity variance (CV%) below 15%.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Sample Preparation and Experimental Design for In Situ Multi-Beam Transmission Electron Microscopy Irradiation Experiments

We discuss the need to understand materials exposed to overlapping extreme environments such as high temperature, radiation, or mechanical stress. When these stressors are combined there may be synergistic effects that enable unique microstructural evolution mechanisms to activate. Understanding of these mechanisms is necessary for the input and refinement of predictive models and critical for engineering of next generation materials. The basic physics and underlying mechanisms require advanced tools to be investigated. The in situ ion irradiation transmission electron microscope (I 3 TEM) is designed to explore these principles. To quantitatively probe the complex dynamic interactions in materials, careful preparation of samples and consideration of experimental design is required. Particular handling or preparation of samples can easily introduce damage or features that obfuscate the measurements. There is no one correct way to prepare a sample; however, many mistakes can be made. The most common errors and things to consider are highlighted within. The I 3 TEM has many adjustable variables and a large potential experimental space, therefore it is best to design experiments with a specific scientific question or questions in mind. Experiments have been performed on large number of sample geometries, material classes, and with many irradiation conditions. The following are a subset of examples that demonstrate unique in situ capabilities utilizing the I 3 TEM. Au nanoparticles prepared by drop casting have been used to investigate the effects of single ion strikes. Au thin films have been used in studies on the effects of multibeam irradiation on microstructure evolution. Zr films have been exposed to irradiation and mechanical tension to examine creep. Ag nanopillars were subjected to simultaneous high temperature, mechanical compression, and ion irradiation to study irradiation induced creep as well. These results impact fields including: structural materials, nuclear energy, energy storage, catalysis, and microelectronics in space environments.

47 OTHER INSTRUMENTATION↗
DOE OSTI2022

Using Xe Plasma FIB for High-Quality TEM Sample Preparation

Here, a direct comparison between electron transparent transmission electron microscope (TEM) samples prepared with gallium (Ga) and xenon (Xe) focused ion beams (FIBs) is performed to determine if equivalent quality samples can be prepared with both ion species. We prepared samples using Ga FIB and Xe plasma focused ion beam (PFIB) while altering a variety of different deposition and milling parameters. The samples’ final thicknesses were evaluated using STEM-EELS $\textit{t/λ}$ data. Using the Ga FIB sample as a standard, we compared the Xe PFIB samples to the standard and to each other. We show that although the Xe PFIB sample preparation technique is quite different from the Ga FIB technique, it is possible to produce high-quality, large area TEM samples with Xe PFIB. We also describe best practices for a Xe PFIB TEM sample preparation workflow to enable consistent success for any thoughtful FIB operator. For Xe PFIB, we show that a decision must be made between the ultimate sample thickness and the size of the electron transparent region.

47 OTHER INSTRUMENTATION↗
DOE OSTI2024

Humidified sample preparation station for serial crystallography

Humidified sample preparation station for serial crystallography according to one embodiment is a humidified enclosure that delivers relative humidities above 95% and preferably above 97% in standard operation, and that can allow microscope observation of samples within. Humidified sample preparation station for serial crystallography can be used for preparation of protein crystal samples for examination using X-rays and for protein structure determination by X-ray crystallography, involving addition of liquid to the sample and removal of liquid from the sample using vacuum or suction.

Jayne, Richard↗
NASA NTRS2011

Development of a Novel Self-Enclosed Sample Preparation Device for DNA/RNA Isolation in Space

Modern biology techniques present potentials for a wide range of molecular, cellular, and biochemistry applications in space, including detection of infectious pathogens and environmental contaminations, monitoring of drug-resistant microbial and dangerous mutations, identification of new phenotypes of microbial and new life species. However, one of the major technological blockades in enabling these technologies in space is a lack of devices for sample preparation in the space environment. To overcome such an obstacle, we constructed a prototype of a DNA/RNA isolation device based on our novel designs documented in the NASA New Technology Reporting System (MSC-24811-1/3-1). This device is self-enclosed and pipette free, purposely designed for use in the absence of gravity. Our design can also be modified easily for preparing samples in space for other applications, such as flowcytometry, immunostaining, cell separation, sample purification and separation according to its size and charges, sample chemical labeling, and sample purification. The prototype of our DNA/RNA isolation device was tested for efficiencies of DNA and RNA isolation from various cell types for PCR analysis. The purity and integrity of purified DNA and RNA were determined as well. Results showed that our developed DNA/RNA isolation device offers similar efficiency and quality in comparison to the samples prepared using the standard protocol in the laboratory.

Zhang, Ye↗
DOE OSTI2019

Proteomic Sample Preparation Techniques: Towards Forensic Proteomic Applications

Proteomic sample preparation techniques rely on dynamic multi-step processes that are sample type dependent. Forensic proteomic applications comprise diverse sample types and therefore require numerous sample preparation workflows. A sample may need pre-treatment steps to solubilize proteins from a solid surface, for example - from clothing, from a swab, or off of a filter. A sample may need to be disrupted or homogenized to liberate proteins trapped in cells. Or a sample may need additional processing steps, such as organic extraction, to isolate proteins from small molecule contaminants which interfere with mass spectrometry (MS) measurements. Following these pre-treatment steps, proteins are reduced, denatured, alkylated, and enzymatically digested. The resulting peptide mixture can be desalted by solid phase extraction or fractionated prior to liquid chromatography MS-based proteomic characterization. Here we detail each step of this multi-stage sample type dependent processing.

Nicora, Carrie D.↗
DOE OSTI2023

Alkaline-SDS cell lysis of microbes with acetone protein precipitation for proteomic sample preparation in 96-well plate format

Plate-based proteomic sample preparation offers a solution to the large sample throughput demands in the biotechnology field where hundreds or thousands of engineered microbes are constructed for testing is routine. Meanwhile, sample preparation methods that work efficiently on broader microbial groups are desirable for new applications of proteomics in other fields, such as microbial communities. Here, we detail a step-by-step protocol that consists of cell lysis in an alkaline chemical buffer (NaOH/SDS) followed by protein precipitation with high-ionic strength acetone in 96-well format. The protocol works for a broad range of microbes ( e . g ., Gram-negative bacteria, Gram-positive bacteria, non-filamentous fungi) and the resulting proteins are ready for tryptic digestion for bottom-up quantitative proteomic analysis without the need for desalting column cleanup. The yield of protein using this protocol increases linearly with respect to the amount of starting biomass from 0.5–2.0 OD*mL of cells. By using a bench-top automated liquid dispenser, a cost-effective and environmentally-friendly option to eliminating pipette tips and reducing reagent waste, the protocol takes approximately 30 minutes to extract protein from 96 samples. Tests on mock mixtures showed expected results that the biomass composition structure is in close agreement with the experimental design. Lastly, we applied the protocol for the composition analysis of a synthetic community of environmental isolates grown on two different media. This protocol has been developed to facilitate rapid, low-variance sample preparation of hundreds of samples and allow flexibility for future protocol development.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Sample Preparation Methods for Targeted Single-Cell Proteomics

We compared three cell isolation and two proteomic sample preparation methods for single-cell and near-single-cell analysis. Whole blood was used to quantify hemoglobin (Hb) and glycated-Hb (gly-Hb) in erythrocytes using targeted mass spectrometry and stable isotope-labeled standard peptides. Each method differed in cell isolation and sample preparation as follows: 1) FACS and automated preparation in one-pot for trace samples (autoPOTS); 2) limited dilution via microscopy and a novel rapid one-pot sample preparation method that circumvented the need for the solid-phase extraction, low-volume liquid handling instrumentation and humidified incubation chamber; and 3) CellenONE-based cell isolation and the same one-pot sample preparation method used for limited dilution. Only the CellenONE device routinely isolated single-cells from which Hb was measured to be 540–660 amol per red blood cell (RBC), which was comparable to the calculated SI reference range for mean corpuscular hemoglobin (390–540 amol/RBC). FACSAria sorter and limited dilution could routinely isolate single-digit cell numbers, to reliably quantify CMV-Hb heterogeneity. Finally, we observed that repeated measures, using 5–25 RBCs obtained from N = 10 blood donors, could be used as an alternative and more efficient strategy than single RBC analysis to measure protein heterogeneity, which revealed multimodal distribution, unique for each individual.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

A lathe system for micrometre-sized cylindrical sample preparation at room and cryogenic temperatures

A simple two-spindle based lathe system for the preparation of cylindrical samples intended for X-ray tomography is presented. The setup can operate at room temperature as well as under cryogenic conditions, allowing the preparation of samples down to 20 and 50 µm in diameter, respectively, within minutes. Case studies are presented involving the preparation of a brittle biomineral brachiopod shell and cryogenically fixed soft brain tissue, and their examination by means of ptychographic X-ray computed tomography reveals the preparation method to be mainly free from causing artefacts. Since this lathe system easily yields near-cylindrical samples ideal for tomography, a usage for a wide variety of otherwise challenging specimens is anticipated, in addition to potential use as a time- and cost-saving tool prior to focused ion-beam milling. Fast sample preparation becomes especially important in relation to shorter measurement times expected in next-generation synchrotron sources.

36 MATERIALS SCIENCE↗
DOE OSTI2018

Estimation of Dose from Potential Radionuclide Emissions from the Sample Preparation Laboratory at the Materials and Fuels Complex

The Sample Preparation Laboratory (SPL) at the Materials and Fuels Complex will perform sample preparation, examination, and mechanical-property testing of highly irradiated materials. The purpose of this report is to document calculation of the estimated dose at public receptor locations from potential radionuclide releases to the atmosphere from SPL. Atmospheric dispersion and dose calculations were performed with the computer model CAP88-PC Version 4 in accordance with the requirements of Code of Federal Regulations (CFR), Title 40, “Protection of the Environment,” Part 61, “National Emission Standards for Hazardous Air Pollutants (NESHAPs),” Subpart H, “National Emission Standards for Emissions of Radionuclides Other than Radon from Department of Energy Facilities” (40 CFR 61, Subpart H 2010). Emission potentials were calculated based on the methodology in 40 CFR Part 61, Appendix D and additional guidance/approval by EPA Region 10 [see letter from Donald Dossett (EPA Region 10) to Tim Safford (DOE-ID), Oct 19, 2017 (CCN 241475)] for solid materials that undergo heating. Estimates of total effective dose are based on low-level chronic exposure. The potential maximum dose to a member of the public was calculated to be 0.02 mrem/year based on an estimated 16 g of the 30 kg source term (0.053%) being exposed to very high temperatures associated with electrical-discharge machining (EDM) and the remaining material heated to 100°C. Calculations show the maximum dose could be as high as 0.05 mrem/year if the remaining material were heated to 1400°C (i.e., the melting point of stainless steel). The potential maximum dose to a public receptor from emissions at SPL is not expected to exceed 0.1 mrem/year (1% of the standard) so long as the material impacted by EDM does not exceed 56 g (3.5 times the expected mass of 16 g and less than 0.2% of the total source term), and the remaining material is not heated to temperatures greater than 1400°C. It should be noted that credit for HEPA filtration does not impact the dose results because the dose is dominated by radionuclides that have exceeded their boiling point; thus, no credit for HEPA filtration may be applied to the emission factor.

99 GENERAL AND MISCELLANEOUS↗
DOE OSTI2020

Sample preparation with sucrose cryoprotection dramatically alters Zn distribution in the rodent hippocampus, as revealed by elemental mapping

Transition metal ions (Fe, Mn, Cu, Zn) are essential for healthy brain function, but altered concentration, distribution, or chemical form of the metal ions has been implicated in numerous brain pathologies. Currently, it is not possible to image the cellular or sub-cellular distribution of metal ions in vivo and therefore, studying brain-metal homeostasis largely relies on ex vivo in situ elemental mapping. Sample preparation methods that accurately preserve the in vivo elemental distribution are essential if one wishes to translate the knowledge of elemental distributions measured ex vivo toward increased understanding of chemical and physiological pathways of brain disease. Furthermore, the choice of sample preparation is particularly important for metal ions that exist in a labile or mobile form, for which the in vivo distribution could be easily distorted by inappropriate sample preparation. One of the most widely studied brain structures, the hippocampus, contains a large pool of labile and mobile Zn. Herein, we describe how sucrose cryoprotection, the gold standard method of preparing tissues for immuno-histochemistry or immuno-fluorescence, which is also often used as a sample preparation method for elemental mapping studies, drastically alters hippocampal Zn distribution. Based on the results of this study, in combination with a comparison against the strong body of published literature that has used either rapid plunge freezing of brain tissue, or sucrose cryo-protection, we strongly urge investigators in the future to cease using sucrose cryoprotection as a method of sample preparation for elemental mapping, especially if Zn is an analyte of interest.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS1996

Advancement of Solidification Processing Technology Through Real Time X-Ray Transmission Microscopy: Sample Preparation

Two types of samples were prepared for the real time X-ray transmission microscopy (XTM) characterization. In the first series directional solidification experiments were carried out to evaluate the critical velocity of engulfment of zirconia particles in the Al and Al-Ni eutectic matrix under ground (l-g) conditions. The particle distribution in the samples was recorded on video before and after the samples were directionally solidified. In the second series samples of the above two type of composites were prepared for directional solidification runs to be carried out on the Advanced Gradient Heating Facility (AGHF) aboard the space shuttle during the LMS mission in June 1996. X-ray microscopy proved to be an invaluable tool for characterizing the particle distribution in the metal matrix samples. This kind of analysis helped in determining accurately the critical velocity of engulfment of ceramic particles by the melt interface in the opaque metal matrix composites. The quality of the cast samples with respect to porosity and instrumented thermocouple sheath breakage or shift could be easily viewed and thus helped in selecting samples for the space shuttle experiments. Summarizing the merits of this technique it can be stated that this technique enabled the use of cast metal matrix composite samples since the particle location was known prior to the experiment.

Stefanescu, D. M.↗
DOE OSTI2024

Robust surfactant-assisted one-pot sample preparation for label-free single-cell and nanoscale proteomics

With advanced mass spectrometry (MS)-based proteomics, genome-scale proteome coverage can be achieved from bulk cells. However, such bulk measurement obscures cell to cell heterogeneity, precluding proteome profiling of single cells and small numbers of cells of interest. To address this issue, in recent 5 years there are a surge of small sample preparation methods developed for robust effective collection and processing of single cells and small numbers of cells for in-depth MS-based proteome profiling. Based on their broad accessibility, they can be categorized into two types: specific device- and standard PCR tube- or multi-well plate-based methods. Herein we describe the detailed protocol of our recently developed, easily adoptable, Surfactant-assisted One-Pot (SOP) sample preparation coupled with MS method termed SOP-MS for label-free single-cell and nanoscale proteomics. SOP-MS capitalizes on the combination of a MS-compatible surfactant, DDM (n-Dodecyl-ß-D-maltoside), and standard low-bind PCR tube or multi-well plate for ‘all-in-one’ one-pot sample preparation without sample transfer. With its robust and convenient features, SOP-MS can be readily implemented in any MS laboratory for single-cell and nanoscale proteomics. With further improvements in MS detection sensitivity and sample throughput, we believe that SOP-MS could open an avenue for single-cell proteomics with broad applicability in the biological and biomedical research.

Single-cell proteomics, nanoscale proteomics, SOP-↗
NASA NTRS1993

Synchrotron/crystal sample preparation

The Center for Applied Optics (CAO) of the University of Alabama in Huntsville (UAH) prepared this final report entitled 'Synchrotron/Crystal Sample Preparation' in completion of contract NAS8-38609, Delivery Order No. 53. Hughes Danbury Optical Systems (HDOS) is manufacturing the Advanced X-ray Astrophysics Facility (AXAF) mirrors. These thin-walled, grazing incidence, Wolter Type-1 mirrors, varying in diameter from 1.2 to 0.68 meters, must be ground and polished using state-of-the-art techniques in order to prevent undue stress due to damage or the presence of crystals and inclusions. The effect of crystals on the polishing and grinding process must also be understood. This involves coating special samples of Zerodur and measuring the reflectivity of the coatings in a synchrotron system. In order to gain the understanding needed on the effect of the Zerodur crystals by the grinding and polishing process, UAH prepared glass samples by cutting, grinding, etching, and polishing as required to meet specifications for witness bars for synchrotron measurements and for investigations of crystals embedded in Zerodur. UAH then characterized these samples for subsurface damage and surface roughness and figure.

Johnson, R. Barry↗