scholarly journals The low-cost Shifter microscope stage transforms the speed and robustness of protein crystal harvesting

2021 ◽  
Vol 77 (1) ◽  
pp. 62-74
Author(s):  
Nathan David Wright ◽  
Patrick Collins ◽  
Lizbé Koekemoer ◽  
Tobias Krojer ◽  
Romain Talon ◽  
...  
Keyword(s):  
Low Cost ◽  
Complete Removal ◽  
Protein Crystal ◽  
Fine Motor ◽  
Data Repository ◽  
X Ray ◽  
Fragment Screening ◽  
Protective Cover ◽  
Almost All ◽  
Microscope Stage

Despite the tremendous success of X-ray cryo-crystallography in recent decades, the transfer of crystals from the drops in which they are grown to diffractometer sample mounts remains a manual process in almost all laboratories. Here, the Shifter, a motorized, interactive microscope stage that transforms the entire crystal-mounting workflow from a rate-limiting manual activity to a controllable, high-throughput semi-automated process, is described. By combining the visual acuity and fine motor skills of humans with targeted hardware and software automation, it was possible to transform the speed and robustness of crystal mounting. Control software, triggered by the operator, manoeuvres crystallization plates beneath a clear protective cover, allowing the complete removal of film seals and thereby eliminating the tedium of repetitive seal cutting. The software, either upon request or working from an imported list, controls motors to position crystal drops under a hole in the cover for human mounting at a microscope. The software automatically captures experimental annotations for uploading to the user's data repository, removing the need for manual documentation. The Shifter facilitates mounting rates of 100–240 crystals per hour in a more controlled process than manual mounting, which greatly extends the lifetime of the drops and thus allows a dramatic increase in the number of crystals retrievable from any given drop without loss of X-ray diffraction quality. In 2015, the first in a series of three Shifter devices was deployed as part of the XChem fragment-screening facility at Diamond Light Source, where they have since facilitated the mounting of over 120 000 crystals. The Shifter was engineered to have a simple design, providing a device that could be readily commercialized and widely adopted owing to its low cost. The versatile hardware design allows use beyond fragment screening and protein crystallography.

scholarly journals The Low-Cost, Semi-Automated Shifter Microscope Stage Transforms Speed and Robustness of Manual Protein Crystal Harvesting

2019 ◽  
Author(s):  
Nathan David Wright ◽  
Patrick Collins ◽  
Romain Talon ◽  
Elliot Nelson ◽  
Lizbé Koekemoer ◽  
...  
Keyword(s):  
Low Cost ◽  
Complete Removal ◽  
Protein Crystal ◽  
Fine Motor ◽  
Data Repository ◽  
Control Software ◽  
X Ray ◽  
Almost All ◽  
Research Initiatives ◽  
Microscope Stage

AbstractDespite the tremendous success of x-ray cryocrystallography over recent decades, the transfer of crystals from the drops where they grow to diffractometer sample mounts, remains a manual process in almost all laboratories. Here we describe the Shifter, a semi-automated microscope stage that offers an accessible and scalable approach to crystal mounting that exploits on the strengths of both humans and machines. The Shifter control software manoeuvres sample drops beneath a hole in a clear protective cover, for human mounting under a microscope. By allowing complete removal of film seals the tedium of cutting or removing the seal is eliminated. The control software also automatically captures experimental annotations for uploading to the user’s data repository, removing the overhead of manual documentation. The Shifter facilitates mounting rates of 100-240 crystals per hour, in a more controlled process than manual mounting, which greatly extends the lifetime of drops and thus allows for a dramatic increase in the number of crystals retrievable from any given drop, without loss of X-ray diffraction quality. In 2015 the first in a series of three Shifter devices was deployed as part of the XChem fragment screening facility at Diamond Light Source (DLS), where they have since facilitated the mounting of over 100,000 crystals. The Shifter was engineered to be simple, allowing for a low-cost device to be commercialised and thus potentially transformative as many research initiatives as possible.SynopsisA motorised X/Y microscope stage is presented that combines human fine motor control with machine automation and automated experiment documentation, to transform productivity in protein crystal harvesting.

scholarly journals Surface Analysis and Roughness Parameters of Copper Metallic

2015 ◽  
Vol 57 ◽  
pp. 66-71
Author(s):  
Halo Dalshad Omar
Keyword(s):  
High Speed ◽  
Low Cost ◽  
Roughness Parameter ◽  
Fluorescence Analysis ◽  
Surface Measurement ◽  
Scanning Tunneling ◽  
Face Centered Cubic ◽  
X Rays ◽  
X Ray ◽  
Almost All

The level of copper (Cu) distribution in the hard alloy depends little on powder material hardness although the other mechanical properties of the materials and the crystal structure of copper is face centered cubic (FCC). A discussion about simple and low cost preparation of copper powder by (Mini Mill 2 Panalytical) and preparation of the sample was rotating at 5 min and in the case of grinding of samples at high speed 300 rpm. The surface morphology of copper metallic powder was determined with Scanning Tunneling Microscope (STM) (Model: NTMDT Solver Nano.). The characterization of surface topographyand surface measurement determines surface topography and surface measurement copper includes roughness parameter. X-ray fluorescence analysis is a powerful analytical tool for the determination of almost all the elements present in a sample. The spectra obtained were analyzed using a X-ray fluorescence (XRF) (Model: Rigaku-NEX CG). Each element emits a unique spectrum of X-rays characteristic of that element. From the spectra obtained, there was the element to be present in the sample was copper (Cu).

Analytical Electron Microscopy Study of Second-Phase Particles in 7075 and 7475 Aluminum Alloys

1985 ◽  
Vol 43 ◽  
pp. 348-349
Author(s):  
M. Raghavan ◽  
J. Y. Koo ◽  
J. W. Steeds ◽  
B. K. Park
Keyword(s):  
Aluminum Alloys ◽  
Silicon Oxide ◽  
Analytical Electron Microscopy ◽  
Microscopy Study ◽  
Electron Microscopy Study ◽  
Partial Substitution ◽  
Second Phase ◽  
X Ray ◽  
Second Phase Particles ◽  
Almost All

X-ray microanalysis and Convergent Beam Electron Diffraction (CBD) studies were conducted to characterize the second phase particles in two commercial aluminum alloys -- 7075 and 7475. The second phase particles studied were large (approximately 2-5μm) constituent phases and relatively fine ( ∼ 0.05-1μn) dispersoid particles, Figures 1A and B. Based on the crystal structure and chemical composition analyses, the constituent phases found in these alloys were identified to be Al7Cu2Fe, (Al,Cu)6(Fe,Cu), α-Al12Fe3Si, Mg2Si, amorphous silicon oxide and the modified 6Fe compounds, in decreasing order of abundance. The results of quantitative X-ray microanalysis of all the constituent phases are listed in Table I. The data show that, in almost all the phases, partial substitution of alloying elements occurred resulting in small deviations from the published stoichiometric compositions of the binary and ternary compounds.

Resolution of a protein sequence ambiguity by X-ray crystallographic and mass spectrometric methods

1992 ◽  
Vol 25 (2) ◽  
pp. 205-210 ◽  
Author(s):  
L. J. Keefe ◽  
E. E. Lattman ◽  
C. Wolkow ◽  
A. Woods ◽  
M. Chevrier ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Amino Acid ◽  
Electron Density ◽  
Mass Spectrometric ◽  
Amino Acid Sequences ◽  
Data Matrix ◽  
Protein Crystal ◽  
Insertion Mutant ◽  
Laser Desorption Mass Spectrometry ◽  
X Ray

Ambiguities in amino acid sequences are a potential problem in X-ray crystallographic studies of proteins. Amino acid side chains often cannot be reliably identified from the electron density. Many protein crystal structures that are now being solved are simple variants of a known wild-type structure. Thus, cloning artifacts or other untoward events can readily lead to cases in which the proposed sequence is not correct. An example is presented showing that mass spectrometry provides an excellent tool for analyzing suspected errors. The X-ray crystal structure of an insertion mutant of Staphylococcal nuclease has been solved to 1.67 Å resolution and refined to a crystallographic R value of 0.170 [Keefe & Lattman (1992). In preparation]. A single residue has been inserted in the C-terminal α helix. The inserted amino acid was believed to be an alanine residue, but the final electron density maps strongly indicated that a glycine had been inserted instead. To confirm the observations from the X-ray data, matrix-assisted laser desorption mass spectrometry was employed to verify the glycine insertion. This mass spectrometric technique has sufficient mass accuracy to detect the methyl group that distinguishes glycine from alanine and can be extended to the more common situation in which crystallographic measurements suggest a problem with the sequence, but cannot pinpoint its location or nature.

scholarly journals Use of WetSEM® capsules for convenient multimodal scanning electron microscopy, energy dispersive X-ray analysis, and micro Raman spectroscopy characterisation of technetium oxides

2021 ◽  
Author(s):  
D. J. Bailey ◽  
M. C. Stennett ◽  
J. Heo ◽  
N. C. Hyatt
Keyword(s):  
Raman Spectroscopy ◽  
Low Cost ◽  
Powder Materials ◽  
Spectroscopy Analysis ◽  
X Ray ◽  
Micro Raman Spectroscopy ◽  
Hazard And Risk ◽  
Sem Imaging ◽  
General User ◽  
532 Nm

AbstractSEM–EDX and Raman spectroscopy analysis of radioactive compounds is often restricted to dedicated instrumentation, within radiological working areas, to manage the hazard and risk of contamination. Here, we demonstrate application of WetSEM® capsules for containment of technetium powder materials, enabling routine multimodal characterisation with general user instrumentation, outside of a controlled radiological working area. The electron transparent membrane of WetSEM® capsules enables SEM imaging of submicron non-conducting technetium powders and acquisition of Tc Lα X-ray emission, using a low cost desktop SEM–EDX system, as well as acquisition of good quality μ-Raman spectra using a 532 nm laser.

scholarly journals Multidimensional Ln-Aminophthalate Photoluminescent Coordination Polymers

Materials ◽  
2021 ◽  
Vol 14 (7) ◽  
pp. 1786
Author(s):  
Carla Queirós ◽  
Chen Sun ◽  
Ana M. G. Silva ◽  
Baltazar de Castro ◽  
Juan Cabanillas-Gonzalez ◽  
...  
Keyword(s):  
Water Content ◽  
Coordination Polymers ◽  
Metal Ion ◽  
Low Cost ◽  
Microwave Assisted Synthesis ◽  
X Ray Diffraction ◽  
X Ray ◽  
Wide Range ◽  
Ft Ir ◽  
Ft Ir Spectroscopy

The development of straightforward reproducible methods for the preparation of new photoluminescent coordination polymers (CPs) is an important goal in luminescence and chemical sensing fields. Isophthalic acid derivatives have been reported for a wide range of applications, and in addition to their relatively low cost, have encouraged its use in the preparation of novel lanthanide-based coordination polymers (LnCPs). Considering that the photoluminescent properties of these CPs are highly dependent on the existence of water molecules in the crystal structure, our research efforts are now focused on the preparation of CP with the lowest water content possible, while considering a green chemistry approach. One- and two-dimensional (1D and 2D) LnCPs were prepared from 5-aminoisophthalic acid and Sm3+/Tb3+ using hydrothermal and/or microwave-assisted synthesis. The unprecedented LnCPs were characterized by single-crystal X-ray diffraction (SCRXD), powder X-ray diffraction (PXRD), Fourier transform infrared (FT-IR) spectroscopy and scanning electron microscopy (SEM), and their photoluminescence (PL) properties were studied in the solid state, at room temperature, using the CPs as powders and encapsulated in poly(methyl methacrylate (PMMA) films, envisaging the potential preparation of devices for sensing. The materials revealed interesting PL properties that depend on the dimensionality, metal ion, co-ligand used and water content.

scholarly journals Monitoring protein precipitates by in-house X-ray powder diffraction

2013 ◽  
Vol 28 (S2) ◽  
pp. S458-S469 ◽  
Author(s):  
Kenny Ståhl ◽  
Christian G. Frankær ◽  
Jakob Petersen ◽  
Pernille Harris
Keyword(s):  
Powder Diffraction ◽  
Protein Crystal ◽  
Data Sets ◽  
Crystal Forms ◽  
X Ray ◽  
Cell Parameters ◽  
X Ray Scattering ◽  
Source Organism ◽  
Peak Asymmetry ◽  
Overlapping Peaks

Powder diffraction from protein powders using in-house diffractometers is an effective tool for identification and monitoring of protein crystal forms and artifacts. As an alternative to conventional powder diffractometers a single crystal diffractometer equipped with an X-ray micro-source can be used to collect powder patterns from 1 µl samples. Using a small-angle X-ray scattering (SAXS) camera it is possible to collect data within minutes. A streamlined program has been developed for the calculation of powder patterns from pdb-coordinates, and includes correction for bulk-solvent. A number of such calculated powder patterns from insulin and lysozyme have been included in the powder diffraction database and successfully used for search-match identification. However, the fit could be much improved if peak asymmetry and multiple bulk-solvent corrections were included. When including a large number of protein data sets in the database some problems can be foreseen due to the large number of overlapping peaks in the low-angle region, and small differences in unit cell parameters between pdb-data and powder data. It is suggested that protein entries are supplied with more searchable keywords as protein name, protein type, molecular weight, source organism etc. in order to limit possible hits.

scholarly journals Synthesis of Exosome-Based Fluorescent Gold Nanoclusters for Cellular Imaging Applications

2021 ◽  
Vol 22 (9) ◽  
pp. 4433
Author(s):  
Eun Sung Lee ◽  
Byung Seok Cha ◽  
Seokjoon Kim ◽  
Ki Soo Park
Keyword(s):  
Photoelectron Spectroscopy ◽  
Low Cost ◽  
Stokes Shift ◽  
Gold Nanoclusters ◽  
Human Colon ◽  
Cellular Imaging ◽  
Metal Nanoclusters ◽  
Human Colon Cancer ◽  
X Ray ◽  
High Concentrations

In recent years, fluorescent metal nanoclusters have been used to develop bioimaging and sensing technology. Notably, protein-templated fluorescent gold nanoclusters (AuNCs) are attracting interest due to their excellent fluorescence properties and biocompatibility. Herein, we used an exosome template to synthesize AuNCs in an eco-friendly manner that required neither harsh conditions nor toxic chemicals. Specifically, we used a neutral (pH 7) and alkaline (pH 11.5) pH to synthesize two different exosome-based AuNCs (exo-AuNCs) with independent blue and red emission. Using field-emission scanning electron microscopy, energy dispersive X-ray microanalysis, nanoparticle tracking analysis, and X-ray photoelectron spectroscopy, we demonstrated that AuNCs were successfully formed in the exosomes. Red-emitting exo-AuNCs were found to have a larger Stokes shift and a stronger fluorescence intensity than the blue-emitting exo-AuNCs. Both exo-AuNCs were compatible with MCF-7 (human breast cancer), HeLa (human cervical cancer), and HT29 (human colon cancer) cells, although blue-emitting exo-AuNCs were cytotoxic at high concentrations (≥5 mg/mL). Red-emitting exo-AuNCs successfully stained the nucleus and were compatible with membrane-staining dyes. This is the first study to use exosomes to synthesize fluorescent nanomaterials for cellular imaging applications. As exosomes are naturally produced via secretion from almost all types of cell, the proposed method could serve as a strategy for low-cost production of versatile nanomaterials.

DEPOSITION OF SINGLE-PHASE CuInSe2 THIN FILMS UNDER LOW VACUUM LEVEL BY A TWO-STAGE GROWTH TECHNIQUE

2009 ◽  
Vol 16 (03) ◽  
pp. 381-386 ◽  
Author(s):  
J. B. CHU ◽  
H. B. ZHU ◽  
Z. A. WANG ◽  
Z. Q. BIAN ◽  
Z. SUN ◽  
...  
Keyword(s):  
Solar Cells ◽  
Single Phase ◽  
Low Cost ◽  
Control Process ◽  
Rf Magnetron Sputtering ◽  
Chalcopyrite Structure ◽  
Sputtering Deposition ◽  
Vacuum Level ◽  
X Ray ◽  
High Efficient

Single-phase CuInSe 2 films were grown by high vapor selenization of CuIn alloy precursors within a partially closed graphite box. The CuIn precursors were prepared using Cu x In y alloy targets with different composition rates under low vacuum level by a homemade sputtering system. The Cu and In composition rates of the used targets are 11:9, 10:10, and 9:11, respectively. The metallic precursor films were selenized using a two-step temperature profile, i.e. at 250°C and 400–500°C, respectively. The influence of the temperature at the second selenization step on the quality of the CIS absorbing layers was investigated. The CIS films were characterized by X-ray diffractometry, scanning electron microscopy, energy dispersive X-ray analysis, and Raman spectroscopy. The deposited CIS absorbers selenized at a high temperature of 500°C for 30 min exhibited a single-phase chalcopyrite structure with a preferential orientation in the (112) direction. These layers display uniform, large, and densely packed crystals with a grain size of about 3–5 μm. Cadmium sulfide buffer layer was manufactured by chemical bath deposition method. Bilayers ZnO / ZnO : Al were prepared by RF magnetron sputtering deposition. CIS solar cells with an efficiency of about 6.5% were produced without antireflective films. The method to fabricate CIS solar cells by a combination of the low vacuum sputtering deposition and the graphite box selenization process has provided a simple control process and shown a promising potential for developing high efficient and low-cost CuInSe 2 solar cells.

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