Name: fabrication procedures and birefringence measurements for designing magnetically responsive lanthanide ion chelating phospholipid assemblies
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The authors acknowledge the Swiss National Science Foundation for financing SMhardBi (project number 200021_150088/1). The SANS experiments were performed at the Swiss spallation neutron source SINQ, Paul Scherrer Instute, Villigen, Switzerland. The authors warmly thank Dr. Joachim Kohlbrecher for his guidance with the SANS experiments. The birefringence measurement setup under high magnetic fields was inspired from the existing setup at the high-field magnetic laboratory HFML, Nijmegen, The Netherlands. We thank Bruno Pfister for his help in developing the electronics of the birefringence setup, Jan Corsano and Daniel Kiechl for constructing the frameworks permitting fine and facile alignment of the laser, and Dr. Bernhard Koller for ongoing technical support.

1. Fabrication procedure for DMPC/DMPE-DTPA/Tm3+ (molar ratio 4:1:1) and DMPC/Chol-OH/DMPE-DTPA/Tm3+ (molar ratio 16:4:5:5) polymolecular assemblies
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	<li>Preliminary preparations
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		<li>Wash all the glassware by flushing once with ethanol stabilized chloroform (&#62;99% chloroform) and dry with compressed air.</li>
		<li>Produce 2 distinct 10 mg/mL stock solutions of DMPC and DMPE-DTPA in ethanol-stabilized chloroform (&#62;99% chloroform), a 10 mM stock solution of Chol-OH in ethanol-st...

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	<li>Sanders, C. R., Hare, B. J., Howard, K. P., Prestegard, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetically-oriented+phospholipid+micelles+as+a+tool+for+the+study+of+membrane-associated+molecules.">Magnetically-oriented phospholipid micelles as a tool for the study of membrane-associated molecules.</a> Prog. Nucl. Magn. Reson. Spectrosc. 26, 421-444 (1994).</li><li>Glover, K. J., 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=Structural+evaluation+of+phospholipid+bicelles+for+solution-state+studies+of+membrane-associated+biomolecules.">Structural evaluation of phospholipid bicelles for solution-state studies of membrane-associated biomolecules.</a> Biophys. J. 81 (4), 2163-2171 (2001).</li><li>Katsaras, J. H. T. A., Pencer, J., Nieh, M. -. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term="Bicellar"+lipid+mixtures+as+used+in+biochemical+and+biophysical+studies.">"Bicellar" lipid mixtures as used in biochemical and biophysical studies.</a> Naturwissenschaften. 92 (8), 355-366 (2005).</li><li>Sanders, C. R., Prosser, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bicelles:+a+model+membrane+system+for+all+seasons?.">Bicelles: a model membrane system for all seasons?.</a> Structure. 6 (10), 1227-1234 (1998).</li><li>D&#252;rr, U. H. N., Soong, R., Ramamoorthy, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=When+detergent+meets+bilayer:+birth+and+coming+of+age+of+lipid+bicelles.">When detergent meets bilayer: birth and coming of age of lipid bicelles.</a> Prog. Nucl. Magn. Reson. Spectrosc. 69, 1-22 (2013).</li><li>D&#252;rr, U. H. N., Gildenberg, M., Ramamoorthy, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+magic+of+bicelles+lights+up+membrane+protein+structure.">The magic of bicelles lights up membrane protein structure.</a> Chem. 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Bicelle Preparation Available from: <a target="_blank" href="https://avantilipids.com/tech-support/liposome-preparation/bicelle-preparation">https://avantilipids.com/tech-support/liposome-preparation/bicelle-preparation</a> (2017)</li><li>Isabettini, S., 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=Methods+for+Generating+Highly+Magnetically+Responsive+Lanthanide-Chelating+Phospholipid+Polymolecular+Assemblies.">Methods for Generating Highly Magnetically Responsive Lanthanide-Chelating Phospholipid Polymolecular Assemblies.</a> Langmuir. 33, 6363-6371 (2017).</li><li>Nieh, M. -. P., Glinka, C. J., Krueger, S., Prosser, R. 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V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+chelate-induced+magnetic+alignment+of+biological+membranes.">Novel chelate-induced magnetic alignment of biological membranes.</a> Biophys. J. 75, 2163-2169 (1998).</li><li>Shklyarevskiy, I. 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=Magnetic+alignment+of+self-assembled+anthracene+organogel+fibers.">Magnetic alignment of self-assembled anthracene organogel fibers.</a> Langmuir. 21, 2108-2112 (2005).</li><li>Christianen, P. C. M., Shklyarevskiy, I. O., Boamfa, M. I., Maan, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alignment+of+molecular+materials+in+high+magnetic+fields.">Alignment of molecular materials in high magnetic fields.</a> Physica B: Condens. Matter. 346, 255-261 (2004).</li><li>Maret, G., Dransfeld, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biomolecules+and+polymers+in+high+steady+magnetic+fields.">Biomolecules and polymers in high steady magnetic fields.</a> Top. App. Phys. 57, 143-204 (1985).</li><li>Gielen, J. C., Shklyarevskiy, I. O., Schenning, A. P. H. J., Christianen, P. C. M., Maan, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Using+magnetic+birefringence+to+determine+the+molecular+arrangement+of+supramolecular+nanostructures.">Using magnetic birefringence to determine the molecular arrangement of supramolecular nanostructures.</a> Sci. Tech. Adv. Mater. 10 (1), 014601 (2009).</li><li>Shklyarevskiy, I. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Deformation and ordering of molecular assemblies in high magnetic fields. , (2005).</li><li>Fuller, G. 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A detailed account of how birefringence measurements were used in combination with SANS experiments to evaluate methods for generating highly magnetically responsive Ln3+ chelating phospholipids assemblies is in Isabettini et al.23 The proposed fabrication protocols are also applicable for assemblies composed of the longer DPPC and DPPE-DTPA phospholipids or for those containing chemically engineered steroid derivatives in their bilayer.11,<su...

Bicelles are disk-like polymolecular assemblies obtained from numerous lipid mixtures.1,2,3,4,5 They are widely used for the structural characterization of membrane biomolecules by NMR spectroscopy.6,7 However, recent efforts aim to expand the field of possible applications.5,8,9 The most studied bicelle system is composed of a mixture of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), constituting the planar part of the assembly, and 1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC) phospholipid covering the edge.1,2,3 The molecular geometry of the phospholipids composing the bilayer dictate the architecture of the self-assembled polymolecular structure.4,5 Replacing DHPC with DMPE-DTPA generates highly magnetically responsive and tunable bicelle systems.10,11 DMPC/DMPE-DTPA/Ln3+ (molar ratio 4:1:1) bicelles associate with many more paramagnetic lanthanide ions (Ln3+) on the bilayer&#39;s surface, resulting in an enhanced magnetic response.10 Moreover, replacing the water-soluble DHPC molecules with DMPE-DTPA/Ln3+ enables the formation of dilution-resistant bicelles.11
The magnetic alignability of planar polymolecular assemblies is dictated by their overall magnetic energy,
<img alt="Equation 1" src="/files/ftp_upload/56812/56812eq1v2.jpg" /> (1)
where B is the magnetic field strength, <img alt="Equation 2" src="/files/ftp_upload/56812/56812eq2.jpg" /> the magnetic constant, n the aggregation number and <img alt="Equation 3" src="/files/ftp_upload/56812/56812eq3.jpg" /> the molecular diamagnetic susceptibility anisotropy of the lipids composing the bilayer. Therefore, the response of DMPC/DMPE-DTPA/Ln3+ bicelles to magnetic fields is tailored by their size (aggregate number n) and the molecular diamagnetic susceptibility anisotropy &#916;&#967;. The latter is readily achieved by changing the nature of the chelated Ln3+.12,13,14,15 Introducing cholesterol (Chol-OH) or other steroid derivatives in the bilayer offers the possibility of tuning both the aggregate number n and the magnetic susceptibility &#916;&#967; of the assemblies.11,16,17,18,19 For a given lipid composition, larger assemblies contain more lipids capable of contributing to the Emag (larger aggregate number n), resulting in more alignable species. The size of DMPC/DHPC bicelles, for example, is conventionally controlled through optimization of the composing lipid ratio or total concentration.20,21,22 Although this is possible in DMPC/DMPE-DTPA/Ln3+ bicelles, their thermo-reversible transformation from bicelle to vesicles upon heating offers added tailoring options. Mechanical means such as extrusion through membrane filters allows shaping of the vesicles. The magnetically alignable bicelles are regenerated upon cooling to 5 &#176;C and their dimensions are dictated from the vesicle precursors.11 Herein, we focus on the potential of mechanical fabrication procedures with DMPC/DMPE-DTPA/Tm3+ (molar ratio 4:1:1) or DMPC/Chol-OH/DMPE-DTPA/Tm3+ (molar ratio 16:4:5:5) as reference systems. The process works analogously when working with other Ln3+ than Tm3+. The wide range of possibilities offered by these techniques are highlighted in Figure 1 and extensively discussed elsewhere.23
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/56812/56812fig1.jpg" /> 
Figure 1: Schematic overview of the possible fabrication procedures. The studied magnetically alignable Ln3+ chelating polymolecular assemblies are composed of either DMPC/DMPE-DTPA/Tm3+ (molar ratio 4:1:1) or DMPC/Chol-OH/DMPE-DTPA/Tm3+ (molar ratio 16:4:5:5). The dry lipid film is hydrated with a 50 mM phosphate buffer at a pH value of 7.4 and the total lipid concentration is 15 mM. An effective hydration of the lipid film requires either freeze thawing cycles (FT) or heating and cooling cycles (H&#38;C). H&#38;C cycles are necessary to regenerate samples after the last freeze thawing step, or to regenerate samples kept frozen over a prolonged period of time if they are to be used without further extrusion. These steps are extensively discussed by Isabettini et al.23 Maximally alignable polymolecular assemblies are achieved, delivering different assembly architectures based on the lipid composition. The bicelle size and magnetic alignability is tunable by extrusion (Ext) through nanopore membrane filters. The presented alignment factors Af were computed from 2D small angle neutron scattering (SANS) patterns of a DMPC/Chol-OH/DMPE-DTPA/Tm3+ (molar ratio 16:4:5:5) sample extruded through either 800, 400, 200, or 100 nm pores. SANS measurements are a complementary means of quantifying bicelle alignment that will not be covered in more detail herein.11,16 The Af ranges from -1 (parallel neutron scattering or perpendicular alignment of the bicelles with respect to the magnetic field direction) to 0 for isotropic scattering. <a href="//cloudflare.jove.com/files/ftp_upload/56812/56812fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
The structure of bicelles has been extensively studied by a wide range of characterization techniques.13 The alignment of bicelles exposed to a magnetic field has been quantified by using either NMR spectroscopy or small angle neutron scattering (SANS) experiments.5,10,11,12,13,16,17,18,19,24,25 However, the shift and broadening of the NMR peaks occurring in the presence of Ln3+ are serious limitations to the method.15,26,27,28 Although SANS experiments do not suffer from this limitation, alternative and more accessible techniques are desirable for routine quantification of magnetically induced alignment of assemblies in solution. Birefringence measurements are a viable and comparatively simple alternative. Analogously to NMR experiments, birefringence measurements reveal valuable information on lipid rearrangements and lipid phases occurring in the bilayer. Moreover, geometric transformations occurring in the polymolecular assembly with changing environmental conditions such as temperature are monitored.11,12,13,16 Magnetically induced birefringence &#916;n&#8242; has been used to study various types of phospholipid systems.13,29,30 Birefringence measurements based on the phase modulation technique in a magnetic field is a viable method to detect orientation of bicelles.12,16,18,29,31,32 The possibility of investigating bicelles with birefringence in high magnetic fields up to 35 T was also demonstrated by M. Liebi et al.13
When polarized light enters an anisotropic material, it will be refracted in an ordinary and extraordinary wave.11 The two waves have different velocities and are shifted in phase by a retardation &#948;. The degree of retardation &#948; is measured and converted into a birefringence signal&#160;<img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg" /> to quantify the degree of anisotropy in the material using
<img alt="Equation 6" src="/files/ftp_upload/56812/56812eq6.jpg" /> (2)
where &#955; is the wavelength of the laser and d is the thickness of the sample. Phospholipids are optically anisotropic and their optical axis coincides with their long molecular axes, parallel to the hydrocarbon tails.11,12 No retardation is measured if the phospholipids are randomly orientated in solution. Retardation is measured when phospholipids are aligned parallel to each other. The magnetically induced birefringence&#160;<img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg" /> can have a positive or negative sign depending on the orientation of the molecules in the magnetic field; see Figure 2. Phospholipids aligned parallel to the x-axis will result in a negative <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg" />, while those aligned along the z-axis result in a positive <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg" />. No birefringence is observed when the optical axis coincides with the direction of light propagation as the phospholipid aligns parallel to the y-axis.
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/56812/56812fig2.jpg" /> 
Figure 2: Alignment of the phospholipids and corresponding sign of the magnetically induced birefringence <img alt="Equation 12" src="/files/ftp_upload/56812/56812eq12.jpg" />. The sign of the measured <img alt="Equation 12" src="/files/ftp_upload/56812/56812eq12.jpg" /> depends on the orientation of the phospholipid in the magnetic field. Dashed lines indicate the optical axis of the molecule. The light is polarized at 45&#176; and propagates in the y direction. The magnetic field B is in the z direction. This figure has been modified from M. Liebi.11 <a href="//cloudflare.jove.com/files/ftp_upload/56812/56812fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
In the case of an isotropic colloidal suspension of bicelles, the orientation induced by the arrangement of the phospholipids in the bilayer will be lost, zeroing the retardation &#948;. The bicelles must also align in order to orientate the optically active phospholipids in their bilayers, causing a retardation &#948; of the polarized light. Consequently, birefringence is a sensitive tool to quantify the magnetic alignability of polymolecular assemblies. Bicelles aligned perpendicular to the magnetic field will yield a positive <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg" />, while those aligned parallel will yield a negative <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg" />. The sign depends on the alignment of the setup and may be checked with a reference sample.

<table>
	<tbody>
		<tr>
			<td>1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)</td>
			<td>Avanti Polar Lipids</td>
			<td>850345P</td>
			<td>&#62;99%</td>
		</tr>
		<tr>
			<td>1,2-dimyristoyl-sn-glycero-3-phospho-ethanolamine-diethylene triaminepentaacetate acid hexammonium salt (DMPE-DTPA)</td>
			<td>Avanti Polar Lipids</td>
			<td>790535P</td>
			<td>&#62;99%</td>
		</tr>
		<tr>
			<td>Thulium(III) chloride</td>
			<td>Sigma-Aldrich</td>
			<td>439649</td>
			<td>anhydrous, powder, 99.9% trace metals basis</td>
		</tr>
		<tr>
			<td>Dysprosium(III) chloride</td>
			<td>Sigma-Aldrich</td>
			<td>325546</td>
			<td>anhydrous, powder, 99.9% trace metals basis</td>
		</tr>
		<tr>
			<td>Ytterbium(III) chloride</td>
			<td>Sigma-Aldrich</td>
			<td>439614</td>
			<td>anhydrous, powder, 99.9% trace metals basis</td>
		</tr>
		<tr>
			<td>Chloroform</td>
			<td>Sigma-Aldrich</td>
			<td>319988</td>
			<td>contains ethanol as stabilizer, ACS reagent, &#8805;99.8%</td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>Sigma-Aldrich</td>
			<td>34860</td>
			<td>&#8805;99.9%</td>
		</tr>
		<tr>
			<td>Cholesterol</td>
			<td>Amresco</td>
			<td>433</td>
			<td>Ultra pure grade</td>
		</tr>
		<tr>
			<td>D2O</td>
			<td>ARMAR chemicals</td>
			<td>1410</td>
			<td>99.8 atom % D</td>
		</tr>
		<tr>
			<td>Ultrapure water</td>
			<td>Millipore</td>
			<td></td>
			<td>Synergy pak2 (SYPK0SIX2), Millipack GP (MPGP02001)</td>
		</tr>
		<tr>
			<td>electronic pH meter</td>
			<td>Metrohm</td>
			<td>17440010</td>
			<td></td>
		</tr>
		<tr>
			<td>Whatmann Nuclepore 25 mm 100nm membrane filter</td>
			<td>VWR</td>
			<td>515-2028</td>
			<td></td>
		</tr>
		<tr>
			<td>Whatmann Nuclepore 25 mm 200nm membrane filter</td>
			<td>VWR</td>
			<td>515-2029</td>
			<td></td>
		</tr>
		<tr>
			<td>Whatmann Nuclepore 25 mm 400nm membrane filter</td>
			<td>VWR</td>
			<td>515-2030</td>
			<td></td>
		</tr>
		<tr>
			<td>Whatmann Nuclepore 25 mm 800nm membrane filter</td>
			<td>VWR</td>
			<td>515-2032</td>
			<td></td>
		</tr>
		<tr>
			<td>Whatmann Filter paper</td>
			<td>VWR</td>
			<td>230600</td>
			<td></td>
		</tr>
		<tr>
			<td>25 ml round bottom flask</td>
			<td>VWR</td>
			<td>201-1352</td>
			<td>14/23 NS</td>
		</tr>
		<tr>
			<td>3 ml glass snap-cup</td>
			<td>VWR</td>
			<td>548-0554</td>
			<td>ND18, 18x30mm</td>
		</tr>
		<tr>
			<td>2.5 ml glass syringe</td>
			<td>Hamilton</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium dihydrogen phosphate dihydrate</td>
			<td>Merk</td>
			<td>1.06342</td>
			<td>Salt used to make phosphate buffer</td>
		</tr>
		<tr>
			<td>di-Sodium hydrogen phosphate</td>
			<td>Merk</td>
			<td>1.06586</td>
			<td>Salt used to make phosphate buffer</td>
		</tr>
		<tr>
			<td>Liquid Nitrogen</td>
			<td>Carbagas</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Pressurized Nitrogen gas</td>
			<td>Carbagas</td>
			<td>-</td>
			<td>200 bar bottle</td>
		</tr>
		<tr>
			<td>Lipid Extruder 10 ml</td>
			<td>Lipex</td>
			<td>-</td>
			<td>Fully equipped with thermobarrel</td>
		</tr>
		<tr>
			<td>High-pressure PVC tube</td>
			<td>GR NETUM</td>
			<td>-</td>
			<td>must resist more than 4 MPa</td>
		</tr>
		<tr>
			<td>Serto adaptors</td>
			<td>Sertot</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Nitrile gloves</td>
			<td>VWR</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>2 ml glass pipettes</td>
			<td>VWR</td>
			<td>612-1702</td>
			<td>230 mm long</td>
		</tr>
		<tr>
			<td>Diode Laser</td>
			<td>Newport</td>
			<td>LPM635-25C</td>
			<td></td>
		</tr>
		<tr>
			<td>DSP Dual Phase Lock-in Amplifier</td>
			<td>SRS</td>
			<td>SR830</td>
			<td></td>
		</tr>
		<tr>
			<td>Photodiode Detector</td>
			<td>Silonex Inc.</td>
			<td>SLSD-71N5</td>
			<td>5mm2, Silicon, photo-conductive</td>
		</tr>
		<tr>
			<td>5.5 T Cryogenic Magnetic</td>
			<td>Cryogenic/Oerlikon AG</td>
			<td>-</td>
			<td>12 bar He-cooled. RW4000/6000 compressor, RGD 5/100 TA cryo-head</td>
		</tr>
		<tr>
			<td>Second order low pass filter</td>
			<td>home-built</td>
			<td>-</td>
			<td>Linear power supply 24V DC, second order, Sallen Key, cut-off frequency 360 Hz, +/- 12V, max 10 mA</td>
		</tr>
		<tr>
			<td>Photoelastic modulator</td>
			<td>Hinds instruments</td>
			<td>PEM-90</td>
			<td></td>
		</tr>
		<tr>
			<td>Glan-Thompson Calcite Polarizer</td>
			<td>Newport</td>
			<td>10GT04</td>
			<td>25.4mm diameter</td>
		</tr>
		<tr>
			<td>Quartz sample cuvette</td>
			<td>Hellma</td>
			<td>165-10-40</td>
			<td>temperature controlled cell, 0.8 ml, 10mm path length</td>
		</tr>
		<tr>
			<td>Temperature probe</td>
			<td>Thermocontrol</td>
			<td>-</td>
			<td>Type K, 0.5mm diameter, Thermocoax</td>
		</tr>
		<tr>
			<td>Non-polarizing mirrors</td>
			<td>Newport</td>
			<td>50326-1002</td>
			<td>25.4mm</td>
		</tr>
		<tr>
			<td>RS 232 cables</td>
			<td>National Instruments</td>
			<td>189284-02</td>
			<td>For Connecting to the RS-232 Port on the front of Compact FieldPoint Controllers</td>
		</tr>
		<tr>
			<td>BNC 50 &#937; cable and connectors</td>
			<td>National Instruments</td>
			<td>763389-01</td>
			<td></td>
		</tr>
		<tr>
			<td>cFP-AI-110</td>
			<td>National Instruments</td>
			<td>777318-110</td>
			<td>8-Channel Analog Voltage and Current Input Module for Compact FieldPoint</td>
		</tr>
		<tr>
			<td>cFP-CB-1</td>
			<td>National Instruments</td>
			<td>778618-01</td>
			<td>Integrated Connector Block for Wiring to Compact FieldPoint I/O</td>
		</tr>
		<tr>
			<td>cFP-CB-3</td>
			<td>National Instruments</td>
			<td>778618-03</td>
			<td>Integrated Isothermal Connector Block for Wiring Thermocouples to the cFP-TC-120 Module</td>
		</tr>
		<tr>
			<td>cFP-TC-120</td>
			<td>National Instruments</td>
			<td>777318-120</td>
			<td>8-Channel Thermocouple Input Module for Compact FieldPoint</td>
		</tr>
		<tr>
			<td>cFP-1804</td>
			<td>National Instruments</td>
			<td>779490-01</td>
			<td>Ethernet/Serial Interface for NI Compact FieldPoint</td>
		</tr>
		<tr>
			<td>LabView 2010</td>
			<td>National Instruments</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Industrial power supply</td>
			<td>Traco Power</td>
			<td>TCL 060-124</td>
			<td>100-240V AC</td>
		</tr>
		<tr>
			<td>Waterbath</td>
			<td>Julabo</td>
			<td>FP40-HE</td>
			<td>refrigerated/Heating Circulator</td>
		</tr>
	</tbody>
</table>

fabrication procedures and birefringence measurements for designing magnetically responsive lanthanide ion chelating phospholipid assemblies

Bicelles are tunable disk-like polymolecular assemblies formed from a large variety of lipid mixtures. Applications range from membrane protein structural studies by nuclear magnetic resonance (NMR) to nanotechnological developments including the formation of optically active and magnetically switchable gels. Such technologies require high control of the assembly size, magnetic response and thermal resistance. Mixtures of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and its lanthanide ion (Ln3+) chelating phospholipid conjugate, 1,2-dimyristoyl-sn-glycero-3-phospho-ethanolamine-diethylene triaminepentaacetate (DMPE-DTPA), assemble into highly magnetically responsive assemblies such as DMPC/DMPE-DTPA/Ln3+ (molar ratio 4:1:1) bicelles. Introduction of cholesterol (Chol-OH) and steroid derivatives in the bilayer results in another set of assemblies offering unique physico-chemical properties. For a given lipid composition, the magnetic alignability is proportional to the bicelle size. The complexation of Ln3+ results in unprecedented magnetic responses in terms of both magnitude and alignment direction. The thermo-reversible collapse of the disk-like structures into vesicles upon heating allows tailoring of the assemblies&#39; dimensions by extrusion through membrane filters with defined pore sizes. The magnetically alignable bicelles are regenerated by cooling to 5 &#176;C, resulting in assembly dimensions defined by the vesicle precursors. Herein, this fabrication procedure is explained and the magnetic alignability of the assemblies is quantified by birefringence measurements under a 5.5 T magnetic field. The birefringence signal, originating from the phospholipid bilayer, further enables monitoring of polymolecular changes occurring in the bilayer. This simple technique is complementary to NMR experiments that are commonly employed to characterize bicelles.

The birefringence signal of a non-extruded DMPC/DMPE-DTPA/Tm3+ (molar ratio 4:1:1) sample was monitored under a 5.5 T magnetic field during a heating and cooling cycle from 5 to 40 &#176;C and back at a rate of 1 &#176;C/min (Figure 6). The birefringence results confirmed high magnetic alignments at 5 &#176;C with a value of 1.5 x 10-5, twice as strong as for the reported extruded systems.6,<sup class="...

Watch this Scientific Journal Video about Fabrication Procedures and Birefringence Measurements for Designing Magnetically Responsive Lanthanide Ion Chelating Phospholipid Assemblies at JoVE.com

Fabrication Procedures and Birefringence Measurements for Designing Magnetically Responsive Lanthanide Ion Chelating Phospholipid Assemblies

Fabrication procedures for highly magnetically responsive lanthanide ion chelating polymolecular assemblies are presented. The magnetic response is dictated by the assembly size, which is tailored by extrusion through nanopore membranes. The assemblies&#39; magnetic alignability and temperature-induced structural changes are monitored by birefringence measurements, a complimentary technique to nuclear magnetic resonance and small angle neutron scattering.

Bicelles are tunable disk-like polymolecular assemblies formed from a large variety of lipid mixtures. Applications range from membrane protein structural ...

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