This work was performed at GeoSoilEnviroCARS (Sector 13), Partnership for Extreme Crystallography program (PX^2), Advanced Photon Source (APS), and Argonne National Laboratory. GeoSoilEnviroCARS is supported by the National Science Foundation-Earth Sciences (EAR-1128799) and Department of Energy-Geosciences (DE-FG02-94ER14466). The PX^2 program is supported by COMPRES under NSF Cooperative Agreement EAR 11-57758. Use of the Advanced Photon Source was supported by the US Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. DE-C02-6CH11357. Use of the COMPRES-GSECARS gas loading system was supported by COMPRES under NSF Cooperative Agreement EAR 11-57758 and by GSECARS through NSF grant EAR-1128799 and DOE grant DE-FG02-94ER14466. We would also like to thank Prof. R. T. Downs at the University of Arizona for kindly providing the samples from RRUFF collections.

1. Sample Preparation
NOTE: The sample preparation process includes three major steps: preparing the empty DAC, loading the sample and loading the inert gas pressure transmitting medium. DAC preparation and sample loading have been described in detail in Lavina et al. 10, and pressure transmitting medium loading has been described in Rivers et al. 14 Here we briefly describe the typical sample preparation process.
<ol>
	<li>Select a pair of conical diamonds with matching backing plates.
	<ol>
		<li>Ensure that the flat tips of the diamond anvils (culets) are identical to each other. 
		NOTE: The diameter of the culet depends on the target maximum pressure of the experiment.</li>
		<li>Ensure that the conical housing of the backing plate matches the shape of the conical diamond. Ensure that the height of the diamond, the diameter of the back side of the diamond, the opening angle of the backing plate and the opening angle of the DAC are compatible with each other, so as to maximize the angular access for diffraction 9,15.</li>
	</ol>
	</li>
	<li>Clean the diamonds and backing plates in an acetone ultrasonic bath for 3-5 min. Set the ultrasonic cleaner to &#34;Sonic&#34; mode. Examine the diamonds and backing plates under an optical microscope, and remove all visible dust or debris.</li>
	<li>Place the diamond in the conical housing of the backing plate, and the assembly in a mounting jig.</li>
	<li>Apply a few kg of load to the diamond with compressing screws, so that it is in intimate contact with the backing plate. Then, apply about 0.1 g of epoxy around the outer circumference of the diamond. Heat the assembly to 70 &#176;C for 8 h to cure the epoxy.</li>
	<li>Clean the interior of the DAC, the back side of the backing plates and the culets of the diamonds with acetone, and place the backing plates with diamonds in the DAC.</li>
	<li>Adjust the positions of the diamonds with set screws that hold the backing plate in place, so that the anvil culets are concentric when the two opposing diamonds come into contact.</li>
	<li>Check the tilt angle between the two culets looking for interference fringes under an optical microscope. Adjust the tilt angle of the diamonds by fine-tuning the load of the set screws, so that the number of fringes is minimized 10.</li>
	<li>Place a piece of 250 &#181;m thick rhenium (Re) metal foil between the two diamonds, and fix it in place with wax. 
	NOTE: This Re foil acts as the gasket of the DAC.</li>
	<li>Apply load uniformly and slowly by tightening the four compressing screws of the DAC, and monitor the thickness change of the Re gasket with a micrometer by measuring the total thickness of the DAC assembly.</li>
	<li>When the thickness of the Re gasket is ~40 &#181;m, remove the load of the compressing screws slowly, and take out the pre-indented gasket.</li>
	<li>Use a laser milling machine to drill a hole at the center of the pre-indentation, whose diameter is at least 2/3 of the culet diameter. Align the gasket with an optical microscope, and set the intended diameter of the gasket hole in the laser milling machine user interface. Press the &#34;scan&#34; button to drill the hole.</li>
	<li>Soak the drilled gasket into acetone and clean it with an ultrasonic cleaner in normal mode. Clean the culets of the diamonds with acetone. Place the drilled gasket back in the DAC. 
	NOTE: The hole in the gasket serves as the sample chamber.</li>
	<li>Place a single crystal sample at the center of the gasket hole, which should also be the center of the diamond culet. 
	NOTE: The optimal sample size is 20 &#181;m x 20 &#181;m x 5 &#181;m (length x width x thickness).</li>
	<li>Place a small ruby sphere (~10 &#181;m in diameter) close to the sample. 
	NOTE: The ruby sphere serves as the pressure marker.</li>
	<li>Place the DAC with the sample inside the COMPRES/GSECARS gas loading vessel and load compressed helium (He) or neon (Ne) gas to a maximum pressure of about 25,000 psi to fill the vessel 14.</li>
	<li>Increase the pressure inside the DAC sample chamber by tightening the compressing screws of the DAC, and monitor the pressure with ruby fluorescence 16.</li>
</ol>
2. Data Collection
<ol>
	<li>Place ~1 mg LaB6 powder at the rotation center of the diffractometer, and collect powder diffraction patterns at several MARCCD detector positions varying by the&#160;&#948; angle. Collect the powder diffraction patterns by clicking the &#34;Start&#34; button of the MARCCD EPICS interface. Use this diffraction pattern to calibrate the detector-sample distance and the tilt of the MARCCD detector 2.</li>
	<li>After completing the detector calibration, remove the LaB6 standard from the diffractometer. Place the DAC in the sample holder, and put it on the diffractometer&#39;s sample stage.
	<ol>
		<li>Use a clamp-type holder to hold ambient temperature DACs, a water-cooled clamp-type holder for high temperature DACs, and polymer micromesh mounted on an ACA/IUCr standard goniometer head to hold a sample at ambient pressure and temperature. 
		NOTE: In the following steps (2.3-2.8), all the motion controls are achieved with the EPICS user interface (EUI).</li>
	</ol>
	</li>
	<li>Rotate the &#966; axis so that the sample chamber is perpendicular to the viewing zoom camera by setting the &#966; angle to 120 in the EUI.</li>
	<li>Find the sample chamber with the viewing camera at the minimum magnification first. Center the sample&#39;s image by changing the sample X, Y and Z in the EUI. Focus the image of the sample by changing the microscope Z in the EUI, and then zoom-in to the maximum magnification.</li>
	<li>Align the sample chamber&#39;s image to the center of the viewing camera by changing the sample X, Y and Z in the EUI. Adjust the sample position along the camera&#39;s axis until it is in focus (using a pre-determined camera focus to estimate the position of the rotation center in this direction). Then rotate the &#966; angle to 90 in the EUI so that the sample chamber is perpendicular to the incident X-ray beam.</li>
	<li>To correct for sample displacement from the center of the instrument along the DAC axis, scan the DAC position in both horizontal and vertical directions perpendicular to the incident X-ray using the SCANW software, using motorized translations built into the goniometer, while collecting transmitted beam intensity data with a photodiode detector placed behind the sample. 
	NOTE: The photodiode detector is mounted on a pneumatic actuator and can be moved in and out of the beam remotely from the control station.</li>
	<li>Find the center position in the collected intensity scan using the &#34;center&#34; function of SCANW, corresponding to maximum transmission. This is the center of the sample chamber.</li>
	<li>Rotate the sample using the goniometer &#966; axis by a few degrees using the EUI, and repeat the vertical transmission scan. Repeat the scan twice, at both positive and negative &#966; offsets.</li>
	<li>Calculate the sample position along the incident X-ray&#39;s direction with the following equation 17 
	<img alt="Equation 1" src="/files/ftp_upload/54660/54660eq1.jpg" /> 
	Where &#916;Y is the distance along the incident X-ray&#39;s direction that the sample is displaced from the center of the instrument, &#916;&#966; is the change in the &#966; angle between scans, &#916;Sx+ and &#916;Sx- are the positional offsets of X-ray&#39;s transmission profile when the &#966; angle is tilted by +&#916;&#966; and -&#916;&#966;. 
	NOTE: Several iterations of scans can be made to improve the accuracy of sample positioning.</li>
	<li>After aligning the sample, collect the single crystal diffraction data with the CCD_DC software 1.
	<ol>
		<li>At first, collect a &#966;-scan with a photodiode by clicking the &#34;scan&#34; button on the SCANW software to determine the maximum opening angle and to determine the functional-shape of the absorption effect of the diamond anvils and backing plates.</li>
		<li>After the &#966;-scan, carry out a wide &#966; exposure (during which the detector is left open while &#966; is rotated) to cover the maximum opening angle that the DAC allows, followed by a series of step &#966; exposures, each covering 1&#176;. Carry out this step by setting the &#34;total range&#34; to the maximum opening angle, and setting the &#34;Number of steps&#34; to the same number in the CCD_DC software. Collect wide &#966; scans at different detector positions by specifying the detector arm position in the&#160;&#948; and&#160;&#957; directions in the CCD_DC software, so as to allow access to more diffraction peaks. 
		NOTE: For crystals with a unit cell larger than 10 &#197;, collection of 10&#176; wide step scans covering the same angular range is also recommended. Exposure times are determined by the absorption from the diamond, and intensity of diffraction features from the sample. Usually select the exposure time that maximizes the intensities of the diffraction peaks without saturation. A typical exposure time is 1-5 s/&#176;. Typical data collection from one crystal at one detector position and one pressure takes about 30 min.</li>
	</ol>
	</li>
</ol>
3. Data Analysis
NOTE: The data analysis is carried out using the ATREX/RSV software suite 2,18. For a detailed explanation of the principles utilized in the software please see the work of Dera, et al. 2
<ol>
	<li>Process the LaB6 calibration file.
	<ol>
		<li>Open the LaB6 powder diffraction image collected at each detector position in the software. Input the wavelength of the incident X-rays (0.434 &#197;), and press the &#34;refine Cal&#34; button. 
		NOTE: The software will automatically calculate the sample-detector distance and the tilt of the detector with respect to the incident X-ray beam. Detector calibration is only conducted at detector positions &#957;=0, &#948;=0. The calibration images collected at non-zero &#957; and &#948; are used for verification of calibration quality.</li>
		<li>Save the calibration files for each detector position by manually editing the &#957; and &#948; settings, as necessary. 
		NOTE: ATREX stores detector calibration .cal files associated with each image series. When opening an image, the program checks if the associated .cal file is present. If it is not, there is the option to select such a file.</li>
	</ol>
	</li>
	<li>After saving the calibrations for all detector positions, create these associations by checking the &#34;Assign cal&#34; checkbox in the ATREX software, so that the program will remember which calibrations to use.</li>
	<li>Search for the sample&#39;s diffraction peaks, and fit the peak intensities.
	<ol>
		<li>Open the wide angle &#966; exposure in the software. Go to the &#34;Search&#34; panel, and search for the diffraction peaks in the wide angle exposure. Manually delete the over-saturated diffraction peaks from the diamond and the diffraction peaks close to the Re gasket rings.</li>
		<li>Fit the diffraction peaks to get their accurate positions and intensities. Search for the sample&#39;s diffraction peaks for all the detector positions by clicking the &#34;peak search&#34; button in software, and save the corresponding peak tables by clicking the &#34;save peaks&#34; button.</li>
	</ol>
	</li>
	<li>Reconstruct the diffraction peaks&#39; distribution in reciprocal space.
	<ol>
		<li>In the program, open the peak table for one detector position, and one image in the step &#966; scan, which is associated with the same detector position. If the detector calibration file has not yet been assigned to this file series, select the appropriate .cal file. Go to the &#34;Scan&#34; panel, and press the &#34;compute Prof. from scan&#34; button. 
		NOTE: This step will find the &#966; angle for each diffraction peak at which the peak intensity is the strongest.</li>
		<li>Save the resulting peak table .pks file. Repeat this step for all the wide rotation images at different detector positions.</li>
	</ol>
	</li>
	<li>Index the diffraction peaks.
	<ol>
		<li>Using the RSV software, open the first peak table file. Use the &#34;Append&#34; function to merge all additional peak tables. Use the RSV plugin to find the preliminary UB matrix of this crystal and to index the diffraction peaks. The software will automatically search for the most probable UB matrix.</li>
		<li>Open the preliminary UB matrix in RSV by importing the .p4p file, and refine the UB matrix with the d-spacing of each diffraction peak using the &#34;Refine w/ d-spac&#34; button. If the symmetry of the crystal is known, select appropriate crystal system constraints. 
		NOTE: When the refinement converges, the optimized UB matrix and the lattice parameters of the crystal (a, b, c, &#945;, &#946; and &#947;) are determined.</li>
		<li>Save the optimized UB matrix as a .ub file. 
		NOTE: In the initial peak search process the program might have missed some low intensity peaks that will be very valuable in the structure determination.</li>
	</ol>
	</li>
	<li>To search for these missing peaks go back to the software, and open the wide angle &#966; exposure image (the associated calibration file should be loaded automatically).</li>
	<li>In the &#34;Predict&#34; panel, open the UB matrix of the crystal and simulate the diffraction pattern. In the &#34;Peaks&#34; panel, search for the observed diffraction peaks, and remove the unobserved peaks (the text box next to the &#34;Observed&#34; button allows to specify the minimum pixel intensity threshold within the peak fitting box that is required for the peak to be considered &#34;observed&#34;). Fit the positions and intensities of the peaks by clicking the &#34;peak fit&#34; button, then save the peak table.</li>
	<li>Merge all the predicted peak tables at different detector positions with the RSV software using the &#34;append&#34; function. If different exposure settings (rotation speed in &#176;/sec) were used, use appropriate scaling factors when opening new .pks files.</li>
	<li>Export the merged peak table as a .hkl text file, which can be used to refine the crystal structure with the SHELX software package. Detailed procedures for conducting structure refinement with SHELX have been well described elsewhere 19,20.</li>
</ol>

<ol>
	<li>Boffa Ballaran, T., Kurnosov, A., Trots, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single-crystal+X-ray+diffraction+at+extreme+conditions:+a+review.">Single-crystal X-ray diffraction at extreme conditions: a review.</a> High Pressure Res. 33 (3), 453-465 (2013).</li><li>Dera, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+pressure+single-crystal+micro+X-ray+diffraction+analysis+with+GSE_ADA/RSV+software.">High pressure single-crystal micro X-ray diffraction analysis with GSE_ADA/RSV software.</a> High Pressure Res. 33 (3), 466-484 (2013).</li><li>Hejny, C., Minkov, V. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-pressure+crystallography+of+periodic+and+aperiodic+crystals.">High-pressure crystallography of periodic and aperiodic crystals.</a> IUCrJ. 2 (2), 218-229 (2015).</li><li>Lavina, B., Dera, P., Downs, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modern+X-ray+Diffraction+Methods+in+Mineralogy+and+Geosciences.">Modern X-ray Diffraction Methods in Mineralogy and Geosciences.</a> Spectroscopic Methods in Mineralology and Materials Sciences. 78, 1-31 (2014).</li><li>McMahon, M. I., Loa, I., Stinton, G. W., Lundegaard, L. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determining+complex+crystal+structures+from+high+pressure+single-crystal+diffraction+data+collected+on+synchrotron+sources.">Determining complex crystal structures from high pressure single-crystal diffraction data collected on synchrotron sources.</a> High Pressure Res. 33 (3), 485-500 (2013).</li><li>Dera, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Metastable+high-pressure+transformations+of+orthoferrosilite+Fs(82).">Metastable high-pressure transformations of orthoferrosilite Fs(82).</a> Phys Earth Planet Inter. 221, 15-21 (2013).</li><li>Finkelstein, G. J., Dera, P. K., Duffy, T. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phase+transitions+in+orthopyroxene+(En(90))+to+49+GPa+from+single-crystal+X-ray+diffraction.">Phase transitions in orthopyroxene (En(90)) to 49 GPa from single-crystal X-ray diffraction.</a> Phys Earth Planet Inter. 244, 78-86 (2015).</li><li>Zhang, J. S., Dera, P., Bass, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+high-pressure+phase+transition+in+natural+Fe-bearing+orthoenstatite.">A new high-pressure phase transition in natural Fe-bearing orthoenstatite.</a> Am Mineral. 97 (7), 1070-1074 (2012).</li><li>Kantor, I., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=BX90:+A+new+diamond+anvil+cell+design+for+X-ray+diffraction+and+optical+measurements.">BX90: A new diamond anvil cell design for X-ray diffraction and optical measurements.</a> Rev Sci Instrum. 83 (12), (2012).</li><li>Lavina, B., Dera, P., Meng, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthesis+and+Microdiffraction+at+Extreme+Pressures+and+Temperatures.">Synthesis and Microdiffraction at Extreme Pressures and Temperatures.</a> J Vis Exp. (80), (2013).</li><li>Miletich, R., Allan, D. R., Kuhs, W. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-Pressure+Single-Crystal+Techniques.">High-Pressure Single-Crystal Techniques.</a> Rev Mineral Geochem. 41 (1), 445-519 (2000).</li><li>Thorkildsen, G., Mathiesen, R. H., Larsen, H. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Angle+calculations+for+a+six-circle+kappa+diffractometer.">Angle calculations for a six-circle kappa diffractometer.</a> J Appl Crystallogr. 32, 943-950 (1999).</li><li>You, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Angle+calculations+for+a+'4S+2D'+six-circle+diffractometer.">Angle calculations for a '4S+2D' six-circle diffractometer.</a> J Appl Crystallogr. 32, 614-623 (1999).</li><li>Rivers, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+COMPRES/GSECARS+gas-loading+system+for+diamond+anvil+cells+at+the+Advanced+Photon+Source.">The COMPRES/GSECARS gas-loading system for diamond anvil cells at the Advanced Photon Source.</a> High Pressure Res. 28 (3), 273-292 (2008).</li><li>Boehler, R., De Hantsetters, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+anvil+designs+in+diamond-cells.">New anvil designs in diamond-cells.</a> High Pressure Res. 24 (3), 391-396 (2004).</li><li>Mao, H. K., Xu, J., Bell, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calibration+of+the+Ruby+Pressure+Gauge+to+800-Kbar+under+Quasi-Hydrostatic+Conditions.">Calibration of the Ruby Pressure Gauge to 800-Kbar under Quasi-Hydrostatic Conditions.</a> J Geophys Solid Earth Planets. 91, 4673-4676 (1986).</li><li>Smith, J. S., Desgreniers, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selected+techniques+in+diamond+anvil+cell+crystallography:+centring+samples+using+X-ray+transmission+and+rocking+powder+samples+to+improve+X-ray+diffraction+image+quality.">Selected techniques in diamond anvil cell crystallography: centring samples using X-ray transmission and rocking powder samples to improve X-ray diffraction image quality.</a> J Synchrotron Res. 16, 83-96 (2009).</li><li>. ATREX: IDL code for single crystal XRD data processing Available from: <a target="_blank" href="https://github.com/pdera/GSE_ADA">https://github.com/pdera/GSE_ADA</a> (2016)</li><li>. Refinement tutorial Available from: <a target="_blank" href="https://github.com/pdera/GSE_ADA/blob/master/Documentation/Refinement%20tutorial.pdf">https://github.com/pdera/GSE_ADA/blob/master/Documentation/Refinement%20tutorial.pdf</a> (2016)</li><li>Sheldrick, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+short+history+of+SHELX.">A short history of SHELX.</a> Acta Crystallographica Section A. 64, 112-122 (2008).</li><li>Pandolfo, F., Nestola, F., Camara, F., Domeneghetti, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-pressure+behavior+of+space+group+P2/n+omphacite.">High-pressure behavior of space group P2/n omphacite.</a> Am Mineral. 97 (2-3), 407-414 (2012).</li><li>Angel, R. J., Bujak, M., Zhao, J., Gatta, G. D., Jacobsen, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effective+hydrostatic+limits+of+pressure+media+for+high-pressure+crystallographic+studies.">Effective hydrostatic limits of pressure media for high-pressure crystallographic studies.</a> J Appl Crystallogr. 40, 26-32 (2007).</li><li>Mccormick, T. C., Hazen, R. M., Angel, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Compressibility+of+Omphacite+to+60+Kbar+-+Role+of+Vacancies.">Compressibility of Omphacite to 60 Kbar - Role of Vacancies.</a> Am Mineral. 74 (11-12), 1287-1292 (1989).</li><li>Srajer, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extraction+of+accurate+structure-factor+amplitudes+from+Laue+data:+wavelength+normalization+with+wiggler+and+undulator+X-ray+sources.">Extraction of accurate structure-factor amplitudes from Laue data: wavelength normalization with wiggler and undulator X-ray sources.</a> J Synchrotron Radiat. 7, 236-244 (2000).</li><li>Tschauner, O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Discovery+of+bridgmanite,+the+most+abundant+mineral+in+Earth,+in+a+shocked+meteorite.">Discovery of bridgmanite, the most abundant mineral in Earth, in a shocked meteorite.</a> Science. 346 (6213), 1100-1102 (2014).</li><li>Dera, P., Weidner, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Mineral Physics 2016 Long-range Planning Report: Harnessing the Extremes: From Atoms and Bonds to Earthquakes and Plate Tectonics. , (2016).</li><li>Rothkirch, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single-crystal+diffraction+at+the+extreme+conditions+beamline+P02.2:+procedure+for+collecting+and+analyzing+high-pressure+single-crystal+data.">Single-crystal diffraction at the extreme conditions beamline P02.2: procedure for collecting and analyzing high-pressure single-crystal data.</a> J synchrotron rad. 20 (5), 711-720 (2013).</li><li>Merlini, M., Hanfland, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single-crystal+diffraction+at+megabar+conditions+by+synchrotron+radiation.">Single-crystal diffraction at megabar conditions by synchrotron radiation.</a> High Pressure Res. 33 (3), 511-522 (2013).</li></ol>

The authors declare no conflict of interest.

In this report we show the detailed procedure for carrying out single crystal diffraction experiments with DACs at the GSECARS 13-BM-C beamline. BX-90 type DACs with BA-type diamond anvils and backing plates are recommended for single crystal diffraction experiments 2,9,15. The advantage of the BX-90 type DAC is its wider angular access compared to the traditional symmetric DACs, which provides for effective sampling of many diffraction peaks 9,15. The wide angular access becomes critical for samples with lower symmetry and with smaller unit cells: the former require more diffraction peaks to constrain the lattice parameters accurately, and the latter give fewer diffraction peaks within the given angular access 2. The more angular access one reaches in the experiment, the more accurate atomic positional parameters one measures 2,4. Restricted angular access may result in a two dimensional reciprocal vector dataset, making reliable data interpretation mathematically impossible 2.
One important, yet often overlooked step is to select suitable pressure transmission medium. Though pressure media such as argon, silicone oil or methanol-ethanol-water solution were used in previous single crystal diffraction experiments that did not exceed 10 GPa 21-23, these pressure media become significantly nonhydrostatic between 5-10 GPa 22, and greatly reduce the quality of the crystal during compression 2,22. Our general experience has been that only He and Ne result in high quality experiments up to 50 GPa (e.g., references6,7). At the APS, these gases can be conveniently loaded into DACs with the use of GSECARS/COMPRES gas-loading apparatus 14. When He or Ne is chosen as the pressure medium, the sample chamber shrinks during the gas loading (Figure 2). Once the sample directly touches the gasket, it breaks easily during the compression. So it is important to drill a big enough sample chamber, whose diameter is at least 2/3 of the culet diameter, to avoid the contact between the sample and the gasket after gas loading.
The synchrotron-based monochromatic single crystal diffraction setup at PX^2 is unique. Compared to the laboratory diffractometers, the synchrotron X-ray source provides a much higher flux (&#62;104) 4,27,28, which significantly improves the signal-to-noise ratio and reduces the data collection time 4,27,28. Synchrotron based powder diffraction is also commonly used to determine the structure of materials at high pressures through the Rietveld approach 4. Single crystal diffraction has advantages over the Rietveld approach, because it decouples the fitting of lattice parameters and structural parameters 2,4. Powder diffraction with Rietveld fitting usually requires fitting both lattice parameters and structural parameters at the same time, while the number of independent observations is typically much lower than in single crystal diffraction 4. Another common structure determination method is Laue diffraction, which uses polychromatic radiation with an area detector 4. Compared to monochromatic data collection at PX^2, the reduction of Laue method data requires additional terms including harmonic deconvolution and intensity normalization, which adds additional difficulties in the data analysis 4,24. Monochromatic single crystal diffraction is a straightforward way of solving structures, yet it has its own limitations. An ideal dataset of monochromatic single crystal diffraction requires a defect-less crystal with a size of tens of &#181;m, and the crystal quality needs to preserve at high pressures. These requirements can be difficult to meet for some non-quenchable minerals, such as bridgmanite 25.
Time resolved single crystal diffraction is capable of capturing the transient metastable states and transformation kinetics during pressure induced structural transitions, and is one of the future research directions for PX^2 26. Quantitative characterization of defects and lattice dynamics, based on analysis of X-ray diffuse scattering at high pressures is also under development at the PX^2 26. A compact optical platform for laser-heated high-pressure single crystal diffraction is being built, and will enable the earth-science community to study the behavior of materials under deep-earth conditions 26.

Single crystal X-ray diffraction is one of the most efficient and well-established ways to determine the chemical composition and structure of a crystalline material at different experimental conditions. Recently there have been a number 1-5 of developments in high-pressure single crystal diffraction. Pressure is one of the major factors that influence the behavior and properties of Earth and planetary materials. High-pressure experiments routinely reveal new polymorphs of common materials and can uncover ways to synthesize chemicals which are impossible to make at ambient conditions. Recently, several new silicate polymorphs have been identified with high-pressure single crystal diffraction, which provide new insight into the properties of Earth&#39;s mantle 6-8.
Different from single crystal diffraction at atmospheric pressure, high-pressure single crystal diffraction requires a pressure vessel to generate and maintain pressure during data collection. The most common pressure vessel used in high-pressure single crystal diffraction is the diamond anvil cell (DAC), which is composed of a pair of diamond anvils held together by a metal frame/metal gasket, and a pressure transmitting medium to provide a hydrostatic environment in the sample chamber 4,9-11. Single crystal diffraction using a diamond anvil cell differs from diffraction at ambient conditions in several important ways. First, the coverage of reciprocal space is significantly reduced due to limited X-ray angular access through the body of the DAC and the backing plates. Second, the angle-dependent absorption of X-rays by the diamonds and backing plates must be determined and used to correct the diffraction signal so that accurate structure factors can computed. Third, any overlap of the sample&#39;s diffraction signal with scatter or diffraction from the DAC components, such as the diamonds, gasket and pressure transmitting medium, must be eliminated. Fourth, aligning the sample in the DAC to the center of the goniometer is difficult. The direction perpendicular to the load axis of the DAC is always blocked by the gasket, and is not accessible to either the optical microscope or the X-ray beam. In the axial direction, the optical microscope can only visualize a displaced image of the sample because of the high refractive index of the diamond. These differences require the invention of new high-pressure single crystal diffraction measurement methods.
The Partnership for Extreme Xtallography (PX^2) project is a new research initiative dedicated to high-pressure single crystal diffraction with DACs. The project is hosted at the GeoSoilEnviroCARS experimental station 13-BM-C at the APS, which provides most of the infrastructure including detectors, focused X-rays and a 6-circle heavy duty diffractometer 12,13 optimized for a variety of advanced crystallography experiments. The diffractometer has six angular degrees of freedom, four sample-orienting (&#181;, &#951;, &#967; and &#966;) and two detector-orienting (&#948; and &#965;). The angular conventions from You 13 are used to describe the motion of the sample and the detector, although the &#951;, &#967; and &#966; motions are pseudo-angles derived from the instrument&#39;s kappa geometry real motors. The experimental procedures have been optimized for high-pressure single crystal diffraction with DACs, and a suite of data processing and analysis software packages has been developed. In this manuscript, we present a detailed protocol for a typical high-pressure single crystal diffraction experiment using the BX-90 type DAC 9, as a guide to collect and analyze data at PX^2.

<table border="0" cellpadding="0" cellspacing="0" width="1063">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Diamond</td>
			<td style="width:160px;">Almax</td>
			<td style="width:136px;">P01037</td>
			<td style="width:520px;">Boehler-Almax type diamond</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Backing plate</td>
			<td style="width:160px;">Almax</td>
			<td style="width:136px;">P01289</td>
			<td>Backing plate&#39;s design should match the diamond&#39;s design</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">Re gasket</td>
			<td style="width:160px;">Alfa Aesar</td>
			<td style="width:136px;">10309</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Epoxy</td>
			<td style="width:160px;">Henkel Loctite</td>
			<td style="width:136px;">Stycast 2651</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Polymer micromesh</td>
			<td style="width:160px;">MiTeGen</td>
			<td style="width:136px;">M3-L18SP-25</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Goniometer head</td>
			<td style="width:160px;">Hampton Research</td>
			<td style="width:136px;">HR4-647</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Software: ATREX</td>
			<td style="width:160px;">Open source software</td>
			<td style="width:136px;"></td>
			<td>Website: https://github.com/pdera/GSE_ADA</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Software: RSV</td>
			<td style="width:160px;">Open source software</td>
			<td style="width:136px;"></td>
			<td>Website: https://github.com/pdera/RSV</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Software: cell_now</td>
			<td style="width:160px;">Bruker Corporation</td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Software: CCD_DC</td>
			<td style="width:160px;">Free software</td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

high pressure single crystal diffraction at px^2

In this report we describe detailed procedures for carrying out single crystal X-ray diffraction experiments with a diamond anvil cell (DAC) at the GSECARS 13-BM-C beamline at the Advanced Photon Source. The DAC program at 13-BM-C is part of the Partnership for Extreme Xtallography (PX^2) project. BX-90 type DACs with conical-type diamond anvils and backing plates are recommended for these experiments. The sample chamber should be loaded with noble gas to maintain a hydrostatic pressure environment. The sample is aligned to the rotation center of the diffraction goniometer. The MARCCD area detector is calibrated with a powder diffraction pattern from LaB6. The sample diffraction peaks are analyzed with the ATREX software program, and are then indexed with the RSV software program. RSV is used to refine the UB matrix of the single crystal, and with this information and the peak prediction function, more diffraction peaks can be located. Representative single crystal diffraction data from an omphacite (Ca0.51Na0.48)(Mg0.44Al0.44Fe2+0.14Fe3+0.02)Si2O6 sample were collected. Analysis of the data gave a monoclinic lattice with P2/n space group at 0.35 GPa, and the lattice parameters were found to be: a = 9.496 &#177;0.006 &#197;, b = 8.761 &#177;0.004 &#197;, c = 5.248 &#177;0.001 &#197;, &#946; = 105.06 &#177;0.03&#186;, &#945; = &#947; = 90&#186;.

We show one representative example of high-pressure single crystal diffraction on the silicate mineral omphacite (Ca0.51Na0.48)(Mg0.44Al0.44Fe2+0.14Fe3+0.02)Si2O6. The omphacite sample was loaded in a BX-90 type DAC with Boehler-Almax (BA) type diamond anvils and backing plates (Figure 1). The sample chamber was filled with a noble-gas pressure transmitting medium (helium in this case) to ensure a hydrostatic pressure environment. The pressure of the sample chamber was 0.35 GPa, determined by ruby fluorescence. The sample was aligned with the rotation center of the diffraction goniometer (Figures 3, 4). We calibrated the position and tilt of the MARCCD detector at &#957; = 0, &#948; = 0 with a LaB6 powder standard (Figure 5). During the experiment, &#951;, &#967; and &#181; angles were fixed at 0. The diffraction peaks of the sample were first analyzed using the &#34;Search&#34; function of the ATREX software (Figure 6). Then, the lattice parameters and the UB matrix of the omphacite single crystal were refined using the RSV software (Figure 7). With the refined UB matrix of the crystal, more diffraction peaks were found using the &#34;Predict&#34; function of the software (Figure 8). The refined lattice parameters of this omphacite single crystal at this pressure are: a = 9.496 &#177;0.006 &#197;, b = 8.761&#177;0.004 &#197;, c = 5.248 &#177;0.001 &#197;, &#946; = 105.06 &#177;0.03&#186;, &#945; = &#947; = 90&#186; (Tab. 1). The omphacite crystal was found to have a monoclinic lattice in the P2/n space group. Our refined lattice parameters are consistent with the published lattice parameters of omphacite with a similar chemical composition and at a similar pressure: P = 0.449 GPa, a = 9.5541 &#177;0.0005 &#197;, b = 8.7481 &#177;0.0007 &#197;, c = 5.2482 &#177;0.0003 &#197;, &#946; = 106.895 &#177;0.004&#186; 21.
<img alt="Figure 1" src="/files/ftp_upload/54660/54660fig1.jpg" /> 
Figure 1: Components of BX-90 DAC which is used for high pressure single crystal diffraction. (a) Boehler-Almax (BA) type diamond; (b) Re gasket; (c) BA type backing plate; (d) BA type diamond glued on BA type backing plate; (e) cylinder part of the BX-90 DAC; (f) piston part of the BX-90 DAC; (g) left-handed (black oxide finish) and right-handed (stainless-steel finish) compressing screws: (h) right-handed compressing screw with disk spring washers; (i) BX-90 DAC assembly ready for high pressure single crystal diffraction experiment. <a href="http://cloudflare.jove.com/files/ftp_upload/54660/54660fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" src="/files/ftp_upload/54660/54660fig2.jpg" /> 
Figure 2: Microscope image of the DAC sample chamber before and after noble gas pressure transmitting medium loading. After the gas pressure transmitting medium loading, the sample chamber hole shrank by ~30% in diameter. <a href="http://cloudflare.jove.com/files/ftp_upload/54660/54660fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" src="/files/ftp_upload/54660/54660fig3.jpg" /> 
Figure 3: Experimental setup for high-pressure single crystal diffraction at PX^2. The six angular degrees of freedom (&#181;, &#951;, &#967;, &#966;, &#948; and &#965;) and the three translational directions (x, y and z) are labeled. The notation for angles follows the angular convention of You 13. <a href="http://cloudflare.jove.com/files/ftp_upload/54660/54660fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" src="/files/ftp_upload/54660/54660fig4.jpg" /> 
Figure 4: Align the sample chamber to the rotation center. Left: sample chamber scans at the X-ray normal direction (blue) and &#966;-rotation by +&#916;&#966; (green) and -&#916;&#966; (red). Right: X-ray transmission profiles of the sample chamber scans at different &#966; angles. The offsets of the X-ray transmission profiles are used to calculate the positional correction along the incident X-ray direction. <a href="http://cloudflare.jove.com/files/ftp_upload/54660/54660fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" src="/files/ftp_upload/54660/54660fig5.jpg" /> 
Figure 5: Calibrating the MARCCD detector using the data analysis software. LaB6 powder diffraction pattern is used to carry out the calibration. <a href="http://cloudflare.jove.com/files/ftp_upload/54660/54660fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" src="/files/ftp_upload/54660/54660fig6.jpg" /> 
Figure 6: Diffraction peak search using the data analysis software. In total 63 diffraction peaks were found in this wide exposure image. <a href="http://cloudflare.jove.com/files/ftp_upload/54660/54660fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" src="/files/ftp_upload/54660/54660fig7.jpg" /> 
Figure 7: Indexing the diffraction peaks and calculating the UB matrix of the sample using the RSV software. The indexing is carried out automatically by the software. <a href="http://cloudflare.jove.com/files/ftp_upload/54660/54660fig7large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 8" src="/files/ftp_upload/54660/54660fig8.jpg" /> 
Figure 8: Predicting the diffraction peaks with the data analysis software. 112 diffraction peaks were found with the same diffraction image as in Figure 6 using the peak-prediction function. <a href="http://cloudflare.jove.com/files/ftp_upload/54660/54660fig8large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Lattice parameter</td>
			<td>Value</td>
		</tr>
		<tr>
			<td>a</td>
			<td>9.496 &#177;0.006 &#197;</td>
		</tr>
		<tr>
			<td>b</td>
			<td>8.761 &#177;0.004 &#197;</td>
		</tr>
		<tr>
			<td>c</td>
			<td>5.248 &#177;0.001 &#197;</td>
		</tr>
		<tr>
			<td>a</td>
			<td>90&#186;</td>
		</tr>
		<tr>
			<td>b</td>
			<td>105.06 &#177;0.03&#186;</td>
		</tr>
		<tr>
			<td>g</td>
			<td>90&#186;</td>
		</tr>
	</tbody>
</table>
Table 1: Lattice parameters of omphacite (Ca0.51Na0.48)(Mg0.44Al0.44Fe2+0.14Fe3+0.02)Si2O6 at 0.35 GPa. The omphacite crystal was found to have a monoclinic lattice in the P2/n space group.

Watch this Scientific Journal Video about High Pressure Single Crystal Diffraction at PX^2 at JoVE.com

High Pressure Single Crystal Diffraction at PX^2

In this report, we describe detailed procedures for carrying out single crystal X-ray diffraction experiments with a diamond anvil cell at the GSECARS 13-BM-C beamline at the Advanced Photon Source. ATREX and RSV programs are used to analyze the data.

In this report we describe detailed procedures for carrying out single crystal X-ray diffraction experiments with a diamond anvil cell (DAC) at the ...

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21.3K Views.  University of Hawai’i at Manoa. In this report, we describe detailed procedures for carrying out single crystal X-ray diffraction experiments with a diamond anvil cell at the GSECARS 13-BM-C beamline at the Advanced Photon Source. ATREX and RSV programs are used to analyze the data.

Video: High Pressure Single Crystal Diffraction at PX^2

High-pressure Sapphire Cell for Phase Equilibria Measurements of CO2/Organic/Water Systems

The high pressure sapphire cell apparatus was constructed to visually determine the composition of multiphase systems without physical sampling. Specifically, the sapphire cell enables visual data collection from multiple loadings to solve a set of material balances to precisely determine phase composition. Ternary phase diagrams can then be established to determine the proportion of each component in each phase at a given condition. In principle, any ternary system can be studied although ternary systems (gas-liquid-liquid) are the specific examples discussed herein. For instance, the ternary THF-Water-CO2 system was studied at 25 and 40&#160;&#176;C and is described herein. Of key importance, this technique does not require sampling. Circumventing the possible disturbance of the system equilibrium upon sampling, inherent measurement errors, and technical difficulties of physically sampling under pressure is a significant benefit of this technique. Perhaps as important, the sapphire cell also enables the direct visual observation of the phase behavior. In fact, as the CO2 pressure is increased, the homogeneous THF-Water solution phase splits at about 2 MPa. With this technique, it was possible to easily and clearly observe the cloud point and determine the composition of the newly formed phases as a function of pressure.
The data acquired with the sapphire cell technique can be used for many applications. In our case, we measured swelling and composition for tunable solvents, like gas-expanded liquids, gas-expanded ionic liquids and Organic Aqueous Tunable Systems (OATS)1-4. For the latest system, OATS, the high-pressure sapphire cell enabled the study of (1) phase behavior as a function of pressure and temperature, (2) composition of each phase (gas-liquid-liquid) as a function of pressure and temperature and (3) catalyst partitioning in the two liquid phases as a function of pressure and composition. Finally, the sapphire cell is an especially effective tool to gather accurate and reproducible measurements in a timely fashion.

High-pressure Sapphire Cell for Phase Equilibria Measurements of CO2/Organic/Water Systems

The high pressure sapphire cell apparatus is a unique tool to study, without sampling, phase behavior under a wide range of pressures. Using a cathetometer, very precise volume measurements can be recorded to measure liquid expansion and phase composition. Thus, this synthetic method enables the study of (1) phase equilibria of multi-component mixtures and (2) the partition behavior of catalyst or model compounds as a function of pressure.

The high pressure sapphire cell apparatus was constructed to visually determine the composition of multiphase systems without physical sampling. ...

Metal-silicate Partitioning at High Pressure and Temperature: Experimental Methods and a Protocol to Suppress Highly Siderophile Element Inclusions

Estimates of the primitive upper mantle (PUM) composition reveal a depletion in many of the siderophile (iron-loving) elements, thought to result from their extraction to the core during terrestrial accretion. Experiments to investigate the partitioning of these elements between metal and silicate melts suggest that the PUM composition is best matched if metal-silicate equilibrium occurred at high pressures and temperatures, in a deep magma ocean environment. The behavior of the most highly siderophile elements (HSEs) during this process however, has remained enigmatic. Silicate run-products from HSE solubility experiments are commonly contaminated by dispersed metal inclusions that hinder the measurement of element concentrations in the melt. The resulting uncertainty over the true solubility and metal-silicate partitioning of these elements has made it difficult to predict their expected depletion in PUM. Recently, several studies have employed changes to the experimental design used for high pressure and temperature solubility experiments in order to suppress the formation of metal inclusions. The addition of Au (Re, Os, Ir, Ru experiments) or elemental Si (Pt experiments) to the sample acts to alter either the geometry or rate of sample reduction respectively, in order to avoid transient metal oversaturation of the silicate melt. This contribution outlines procedures for using the piston-cylinder and multi-anvil apparatus to conduct solubility and metal-silicate partitioning experiments respectively. A protocol is also described for the synthesis of uncontaminated run-products from HSE solubility experiments in which the oxygen fugacity is similar to that during terrestrial core-formation. Time-resolved LA-ICP-MS spectra are presented as evidence for the absence of metal-inclusions in run-products from earlier studies, and also confirm that the technique may be extended to investigate Ru. Examples are also given of how these data may be applied.

We present a procedure to determine the metal-silicate partitioning of siderophile elements, emphasizing techniques that suppress the formation of metal inclusions in experiments for the noble metals. The results of these experiments are used to demonstrate the effect of core-formation on the highly siderophile element composition of the mantle.

Estimates of the primitive upper mantle (PUM) composition reveal a depletion in many of the siderophile (iron-loving) elements, thought to result from ...

Flow-pattern Guided Fabrication of High-density Barcode Antibody Microarray

Antibody microarray as a well-developed technology is currently challenged by a few other established or emerging high-throughput technologies. In this report, we renovate the antibody microarray technology by using a novel approach for manufacturing and by introducing new features. The fabrication of our high-density antibody microarray is accomplished through perpendicularly oriented flow-patterning of single stranded DNAs and subsequent conversion mediated by DNA-antibody conjugates. This protocol outlines the critical steps in flow-patterning DNA, producing and purifying DNA-antibody conjugates, and assessing the quality of the fabricated microarray. The uniformity and sensitivity are comparable with conventional microarrays, while our microarray fabrication does not require the assistance of an array printer and can be performed in most research laboratories. The other major advantage is that the size of our microarray units is 10 times smaller than that of printed arrays, offering the unique capability of analyzing functional proteins from single cells when interfacing with generic microchip designs. This barcode technology can be widely employed in biomarker detection, cell signaling studies, tissue engineering, and a variety of clinical applications.

This protocol outlines the fabrication of a large-scale, multiplexed two-dimensional DNA or antibody array, with potential applications in cell signaling studies and biomarker detection.

Antibody microarray as a well-developed technology is currently challenged by a few other established or emerging high-throughput technologies. In this ...

X-ray Powder Diffraction in Conservation Science: Towards Routine Crystal Structure Determination of Corrosion Products on Heritage Art Objects

The crystal structure determination and refinement process of corrosion products on historic art objects using laboratory high-resolution X-ray powder diffraction (XRPD) is presented in detail via two case studies.
The first material under investigation was sodium copper formate hydroxide oxide hydrate, Cu4Na4O(HCOO)8(OH)2&#8729;4H2O (sample 1) which forms on soda glass/copper alloy composite historic objects (e.g., enamels) in museum collections, exposed to formaldehyde and formic acid emitted from wooden storage cabinets, adhesives, etc. This degradation phenomenon has recently been characterized as &#34;glass induced metal corrosion&#34;.
For the second case study, thecotrichite, Ca3(CH3COO)3Cl(NO3)2&#8729;6H2O (sample 2), was chosen, which is an efflorescent salt forming needlelike crystallites on tiles and limestone objects which are stored in wooden cabinets and display cases. In this case, the wood acts as source for acetic acid which reacts with soluble chloride and nitrate salts from the artifact or its environment.
The knowledge of the geometrical structure helps conservation science to better understand production and decay reactions and to allow for full quantitative analysis in the frequent case of mixtures.

Modern high resolution X-ray powder diffraction (XRPD) in the laboratory is used as an efficient tool to determine crystal structures of long-known corrosion products on historic objects.

The crystal structure determination and refinement process of corrosion products on historic art objects using laboratory high-resolution X-ray powder ...

Rapid High-throughput Species Identification of Botanical Material Using Direct Analysis in Real Time High Resolution Mass Spectrometry

We demonstrate that direct analysis in real time-high resolution mass spectrometry can be used to produce mass spectral profiles of botanical material, and that these chemical fingerprints can be used for plant species identification. The mass spectral data can be acquired rapidly and in a high throughput manner without the need for sample extraction, derivatization or pH adjustment steps. The use of this technique bypasses challenges presented by more conventional techniques including lengthy chromatography analysis times and resource intensive methods. The high throughput capabilities of the direct analysis in real time-high resolution mass spectrometry protocol, coupled with multivariate statistical analysis processing of the data, provide not only class characterization of plants, but also yield species and varietal information. Here, the technique is demonstrated with two psychoactive plant products, Mitragyna speciosa (Kratom) and Datura (Jimsonweed), which were subjected to direct analysis in real time-high resolution mass spectrometry followed by statistical analysis processing of the mass spectral data. The application of these tools in tandem enabled the plant materials to be rapidly identified at the level of variety and species.

A method for species identification of botanical material by direct analysis in real time-high resolution mass spectrometry and multivariate statistical analysis is presented.

We demonstrate that direct analysis in real time-high resolution mass spectrometry can be used to produce mass spectral profiles of botanical material, ...

A Straightforward Method for Glucosinolate Extraction and Analysis with High-pressure Liquid Chromatography (HPLC)

Glucosinolates are a well-studied and highly diverse class of natural plant compounds. They play important roles in plant resistance, rapeseed oil quality, food flavoring, and human health. The biological activity of glucosinolates is released upon tissue damage, when they are mixed with the enzyme myrosinase. This results in the formation of pungent and toxic breakdown products, such as isothiocyanates and nitriles. Currently, more than 130 structurally different glucosinolates have been identified. The chemical structure of the glucosinolate is an important determinant of the product that is formed, which in turn determines its biological activity. The latter may range from detrimental (e.g., progoitrin) to beneficial (e.g., glucoraphanin). Each glucosinolate-containing plant species has its own specific glucosinolate profile. For this reason, it is important to correctly identify and reliably quantify the different glucosinolates present in brassicaceous leaf, seed, and root crops or, for ecological studies, in their wild relatives. Here, we present a well-validated, targeted, and robust method to analyze glucosinolate profiles in a wide range of plant species and plant organs. Intact glucosinolates are extracted from ground plant materials with a methanol-water mixture at high temperatures to disable myrosinase activity. Thereafter, the resulting extract is brought onto an ion-exchange column for purification. After sulfatase treatment, the desulfoglucosinolates are eluted with water and the eluate is freeze-dried. The residue is taken up in an exact volume of water, which is analyzed by high-pressure liquid chromatography (HPLC) with a photodiode array (PDA) or ultraviolet (UV) detector. Detection and quantification are achieved by conducting comparisons of the retention times and UV spectra of commercial reference standards. The concentrations are calculated based on a sinigrin reference curve and well-established response factors. The advantages and disadvantages of this straightforward method, when compared to faster and more technologically advanced methods, are discussed here.

A Straightforward Method for Glucosinolate Extraction and Analysis with High-pressure Liquid Chromatography HPLC

Here, we describe in great detail an established and robust protocol for the extraction of glucosinolates from ground plant materials. After an on-column sulfatase treatment of the extracts, the desulfoglucosinolates are eluted and analyzed by high-pressure liquid chromatography.

Glucosinolates are a well-studied and highly diverse class of natural plant compounds. They play important roles in plant resistance, rapeseed oil ...

Measurements of Long-range Electronic Correlations During Femtosecond Diffraction Experiments Performed on Nanocrystals of Buckminsterfullerene

The precise details of the interaction of intense X-ray pulses with matter are a topic of intense interest to researchers attempting to interpret the results of femtosecond X-ray free electron laser (XFEL) experiments. An increasing number of experimental observations have shown that although nuclear motion can be negligible, given a short enough incident pulse duration, electronic motion cannot be ignored. The current and widely accepted models assume that although electrons undergo dynamics driven by interaction with the pulse, their motion could largely be considered &#39;random&#39;. This would then allow the supposedly incoherent contribution from the electronic motion to be treated as a continuous background signal and thus ignored. The original aim of our experiment was to precisely measure the change in intensity of individual Bragg peaks, due to X-ray induced electronic damage in a model system, crystalline C60. Contrary to this expectation, we observed that at the highest X-ray intensities, the electron dynamics in C60 were in fact highly correlated, and over sufficiently long distances that the positions of the Bragg reflections are significantly altered. This paper describes in detail the methods and protocols used for these experiments, which were conducted both at the Linac Coherent Light Source (LCLS) and the Australian Synchrotron (AS) as well as the crystallographic approaches used to analyse the data.

We describe an experiment designed to probe the electronic damage induced in nanocrystals of Buckminsterfullerene (C60) by intense, femtosecond pulses of X-rays. The experiment found that, surprisingly, rather than being stochastic, the X-ray induced electron dynamics in C60 are highly correlated, extending over hundreds of unit cells within the crystals1.

The precise details of the interaction of intense X-ray pulses with matter are a topic of intense interest to researchers attempting to interpret the ...

Microfluidic Chips for In Situ Crystal X-ray Diffraction and In Situ Dynamic Light Scattering for Serial Crystallography

This protocol describes fabricating microfluidic devices with low X-ray background optimized for goniometer based fixed target serial crystallography. The devices are patterned from epoxy glue using soft lithography and are suitable for in situ X-ray diffraction experiments at room temperature. The sample wells are lidded on both sides with polymeric polyimide foil windows that allow diffraction data collection with low X-ray background. This fabrication method is undemanding and inexpensive. After the sourcing of a SU-8 master wafer, all fabrication can be completed outside of a cleanroom in a typical research lab environment. The chip design and fabrication protocol utilize capillary valving to microfluidically split an aqueous reaction into defined nanoliter sized droplets. This loading mechanism avoids the sample loss from channel dead-volume and can easily be performed manually without using pumps or other equipment for fluid actuation. We describe how isolated nanoliter sized drops of protein solution can be monitored in situ by dynamic light scattering to control protein crystal nucleation and growth. After suitable crystals are grown, complete X-ray diffraction datasets can be collected using goniometer based in situ fixed target serial X-ray crystallography at room temperature. The protocol provides custom scripts to process diffraction datasets using a suite of software tools to solve and refine the protein crystal structure. This approach avoids the artefacts possibly induced during cryo-preservation or manual crystal handling in conventional crystallography experiments. We present and compare three protein structures that were solved using small crystals with dimensions of approximately 10-20 &#181;m grown in chip. By crystallizing and diffracting in situ, handling and hence mechanical disturbances of fragile crystals is minimized. The protocol details how to fabricate a custom X-ray transparent microfluidic chip suitable for in situ serial crystallography. As almost every crystal can be used for diffraction data collection, these microfluidic chips are a very efficient crystal delivery method.

Microfluidic Chips for In Situ Crystal X-ray Diffraction and In Situ Dynamic Light Scattering for Serial Crystallography

This protocol describes in detail how to fabricate and operate microfluidic devices for X-ray diffraction data collection at room temperature. Additionally, it describes how to monitor protein crystallization by dynamic light scattering and how to process and analyze obtained diffraction data.

This protocol describes fabricating microfluidic devices with low X-ray background optimized for goniometer based fixed target serial crystallography. The ...

High Resolution Physical Characterization of Single Metallic Nanoparticles

Individual molecules can be detected and characterized by measuring the degree by which they reduce the ionic current flowing through a single nanometer-scale pore. The signal is characteristic of the molecule&#39;s physicochemical properties and its interactions with the pore. We demonstrate that the nanopore formed by the bacterial protein exotoxin Staphylococcus aureus alpha hemolysin (&#945;HL) can detect polyoxometalates (POMs, anionic metal oxygen clusters), at the single molecule limit. Moreover, multiple degradation products of 12-phosphotungstic acid POM (PTA, H3PW12O40) in solution are simultaneously measured. The single molecule sensitivity of the nanopore method allows for POMs to be characterized at significantly lower concentrations than required for nuclear magnetic resonance (NMR) spectroscopy. This technique could serve as a new tool for chemists to study the molecular properties of polyoxometalates or other metallic clusters, to better understand POM synthetic processes, and possibly improve their yield. Hypothetically, the location of a given atom, or the rotation of a fragment in the molecule, and the metal oxidation state could be investigated with this method. In addition, this new technique has the advantage of allowing the real-time monitoring of molecules in solution.&#160;

Here, we present a protocol to detect discrete metal oxygen clusters, polyoxometalates (POMs), at the single molecule limit using a biological nanopore-based electronic platform. The method provides a complementary approach to traditional analytical chemistry tools used in the study of these molecules.

Individual molecules can be detected and characterized by measuring the degree by which they reduce the ionic current flowing through a single ...

Improving High Viscosity Extrusion of Microcrystals for Time-resolved Serial Femtosecond Crystallography at X-ray Lasers

High-viscosity micro-extrusion injectors have dramatically reduced sample consumption in serial femtosecond crystallographic experiments (SFX) at X-ray free electron lasers (XFELs). A series of experiments using the light-driven proton pump bacteriorhodopsin have further established these injectors as a preferred option to deliver crystals for time-resolved serial femtosecond crystallography (TR-SFX) to resolve structural changes of proteins after photoactivation. To obtain multiple structural snapshots of high quality, it is essential to collect large amounts of data and ensure clearance of crystals between every pump laser pulse. Here, we describe in detail how we optimized the extrusion of bacteriorhodopsin microcrystals for our recent TR-SFX experiments at the Linac Coherent Light Source (LCLS). The goal of the method is to optimize extrusion for a stable and continuous flow while maintaining a high density of crystals to increase the rate at which data can be collected in a TR-SFX experiment. We achieve this goal by preparing lipidic cubic phase with a homogenous distribution of crystals using a novel three-way syringe coupling device followed by adjusting the sample composition based on measurements of the extrusion stability taken with a high-speed camera setup. The methodology can be adapted to optimize the flow of other microcrystals. The setup will be available for users of the new Swiss Free Electron Laser facility.

The success of a time-resolved serial femtosecond crystallography experiment is dependent on efficient sample delivery. Here, we describe protocols to optimize the extrusion of bacteriorhodopsin microcrystals from a high viscosity micro-extrusion injector. The methodology relies on sample homogenization with a novel three-way coupler and visualization with a high-speed camera.

High-viscosity micro-extrusion injectors have dramatically reduced sample consumption in serial femtosecond crystallographic experiments (SFX) at X-ray ...