Name: fabrication procedures and birefringence measurements for designing magnetically responsive lanthanide ion chelating phospholipid assemblies
Uploaded: 2018-01-03T14:00:00
Description: Watch this Scientific Journal Video about Fabrication Procedures and Birefringence Measurements for Designing Magnetically Responsive Lanthanide Ion Chelating Phospholipid Assemblies at JoVE.com

作者承认瑞士国家科学基金会资助 SMhardBi (项目编号 200021_150088/1)。在瑞士 Villigen 谢瑞 Instute 的瑞士散裂中子源 SINQ 进行了无 san 实验。作者热烈感谢 Dr. Kohlbrecher 的指导和无实验。在高磁场下的双折射测量装置是从荷兰奈梅亨型磁性实验室 HFML 的现有设置中得到启发的。我们感谢布鲁诺菲斯特的帮助, 开发的电子双折射设置, 1月 Corsano 和丹尼尔 Kiechl 为构建框架允许精细和轻便的激光对准, 和 Dr. 伯纳德科勒为持续的技术支持。

1. DMPC/甲醚-DTPA/tm3 + (摩尔比 4:1: 1) 和 DMPC/-OH/甲醚-DTPA/tm3 + (摩尔比 16:4: 5:5) 多组件的制作程序
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	<li>初步准备<ol>
		<li>用乙醇稳定的氯仿 (#62; 99% 氯仿) 冲洗所有的玻璃器皿, 用压缩空气烘干。</li>
		<li>在乙醇稳定氯仿 (#62 中生产2种不同的10毫克/毫升的 DMPC 和甲醚-DTPA 的库存溶液; 99% 氯仿), 在乙醇稳定的氯仿 (和 #62; 99% 氯仿) 和10毫米的 TmCl 的库存溶液中, 有一个10毫米的电液.3...

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详细介绍了双折射测量如何与 san 结合使用, 以评估产生高磁响应的方法在3 +螯合磷脂组件中的 Isabettini et al.23所建议的制造协议也适用于由较长的联邦和 DPPE-DTPA 磷脂组成的组件, 或用于含有其双层内化学工程类固醇衍生物的部件。11,12,17,18,19...

Bicelles 是从许多脂质混合物中获得的圆盘状多组件。1,2,3,4,5广泛应用于膜生物分子的核磁共振波谱结构表征。6,7然而, 最近的努力旨在扩大可能的应用领域。5,8,9最受研究的 bicelle 系统由 12-二-sn-glycero-3-phosphocholine (DMPC) 组成, 构成组件的平面部分, 12-己-snglycero-3-phosphocholine (DHPC)磷脂覆盖边缘。1,2,3组成双层磷脂的分子几何决定了自组装多结构的结构。4,5用甲醚-DTPA 替换 DHPC, 生成高磁响应和可调谐的 bicelle 系统。10,11 DMPC/甲醚-DTPA/Ln3 + (摩尔比 4:1: 1) bicelles 与更多的顺磁性镧系离子 (Ln3 +) 在双层表面上相关联, 从而增强了磁响应。10此外, 用甲醚-DTPA/Ln3 +取代水溶性 DHPC 分子, 可以形成耐稀释的 bicelles。11
平面多组件的磁性 alignability 是由它们的整体磁场能量决定的,
<img alt="Equation 1" src="/files/ftp_upload/56812/56812eq1v2.jpg">(1)
其中 B 是磁场强度, 磁常数, n 的聚合数和分子磁性的易感性各向异性组成双层的脂质. <img alt="Equation 2" src="/files/ftp_upload/56812/56812eq2.jpg"> <img alt="Equation 3" src="/files/ftp_upload/56812/56812eq3.jpg">因此, DMPC/甲醚-DTPA/Ln3 + bicelles 对磁场的响应是根据它们的大小 (集合数 n) 和分子磁性磁化率各向异性Δχ来调整的。后者很容易通过改变螯合物的性质在3 +中实现。12,13,14,15在双层中引入胆固醇 (铁-OH) 或其他甾体衍生物, 可以调节聚合数 n 和Δχ的磁化率。11,16,17,18,19对于给定的脂质组合, 较大的组件含有更多的脂类, 能够对 Emag (更大的聚合数 n) 做出贡献, 从而导致更多的 alignable 物种。DMPC/DHPC bicelles 的大小, 例如, 是常规控制通过优化的组成脂质比或总浓度。20,21,22虽然这在 DMPC/甲醚-DTPA/Ln3 + bicelles 中是可能的, 但在加热时, 它们从 bicelle 到泡的热可逆转换提供了更多的剪裁选项。机械手段, 如挤压通过膜过滤器, 可以形成的囊泡。磁性 alignable bicelles 在冷却到5° c 时再生, 并且他们的维度从泡前体被口授。11在这里, 我们专注于机械制造过程的潜力, DMPC/甲醚-DTPA/tm3 + (摩尔比率 4:1: 1) 或 DMPC/, OH/甲醚-DTPA/tm3 + (摩尔比 16:4: 5:5) 作为参考系统。在使用其他 Ln3 +而不是 Tm3 +时, 该进程工作类似。这些技术提供的各种可能性在图 1中得到了突出显示, 并在其他地方进行了广泛的讨论。23
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/56812/56812fig1.jpg"> 
图 1: 可能的制造过程的示意图概述.所研究的磁性 alignable 在3 +螯合多组件是由 DMPC/甲醚-DTPA/Tm3 + (摩尔比 4:1: 1) 或 DMPC/, OH/甲醚-DTPA/Tm3 + (摩尔比 16:4: 5:5) 组成。干脂膜是水合的50毫米磷酸缓冲液 pH 值为 7.4, 总脂质浓度为15毫米。有效的脂质膜水化需要的是冻融循环 (FT) 或加热和冷却循环 (H & #38; C)。H 和 #38; C 周期是必要的, 以再生样品后, 最后冻结解冻步骤, 或再生样品保持长期冻结超过一段时间, 如果他们要使用不进一步挤压。Isabettini et al广泛讨论了这些步骤。23最大的 alignable 多组件实现, 根据脂质组成提供不同的组件体系结构。通过纳米膜过滤器, 挤出 (Ext) 可调节 bicelle 尺寸和磁性 alignability。所呈现的对齐系数f是从2D 小角中子散射 (san) 模式 (DMPC/甲醚/DTPA/Tm3 + (摩尔比率 16:4: 5:5) 中计算的, 通过800、400、200或 100 nm 孔隙进行拉伸。san 度量是量化 bicelle 对齐的补充方法, 此处不会详细介绍。11,16一个f范围从-1 (平行中子散射或垂直对准的 bicelles 与磁场方向) 到0的各向同性散射。<a href="//ecsource.jove.com/files/ftp_upload/56812/56812fig1large.jpg" target="_blank">请点击这里查看更大版本的这个数字。</a>
bicelles 的结构在广泛的表征技术中得到了广泛的研究。13使用核磁共振波谱法或小角度中子散射 (san) 实验对暴露在磁场中的 bicelles 的对准进行了量化。5,10,11,12,13,16,17,18,19,24,25但是, 在 Ln3 +的情况下发生的核磁共振峰值的移位和展宽是对该方法的严重限制。15,26,27,28尽管 san 实验不受此限制的影响, 但在解决方案中对组件的磁致对准的常规定量化方面, 还是可取的替代方法。双折射测量是一个可行的和比较简单的选择。类似核磁共振实验, 双折射测量揭示了有价值的信息, 脂质重和脂质相发生在双层。此外, 多组件中发生的几何变换, 如温度等变化的环境条件进行了监测。11,12,13,16磁致双折射Δn′用于研究各种类型的磷脂系统。13,29,30基于磁场相位调制技术的双折射测量是检测 bicelles 方位的可行方法。12,16,18,29,31,32在高磁场中 bicelles 的双折射率高达 35 T 的可能性也由 m. Liebi et al.显示13
当偏振光进入一个各向异性物质时, 它会在一个普通的和特殊的波中折射。11两个波具有不同的速度, 并在相位上由一个迟滞δ转移。延迟δ的程度被测量并且转换成双折射信号来量化材料中各向异性的程度<img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg">
<img alt="Equation 6" src="/files/ftp_upload/56812/56812eq6.jpg">(2)
其中λ是激光的波长, d 是样品的厚度。磷脂是光学的各向异性, 它们的光轴与它们的长分子轴重合, 与烃尾平行。11,12如果磷脂在溶液中是随机定向的, 则不测量延迟。当磷脂相互平行时, 就会测量迟钝。磁致双折射可根据磁场中分子的取向, 产生正负符号; <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg">请参见图 2。与 x 轴平行的磷脂将导致一个负的, 而那些沿 z-axis 对齐结果为正值的. <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg"> </img> <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg"> </img>当与 y-axis 平行时, 当光学轴与光传播的方向重合时, 不观察到双折射。
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/56812/56812fig2.jpg"> 
图 2: 磷脂的对准和磁致双折射的对应符号 <img alt="Equation 12" src="/files/ftp_upload/56812/56812eq12.jpg"> </img> .测量的的符号取决于在磁场中磷脂的方向. <img alt="Equation 12" src="/files/ftp_upload/56812/56812eq12.jpg">虚线表示分子的光轴。光在45°上极化, 并在 y 方向传播。磁场 B 在 z 方向。这一数字已从 m Liebi 中修改。11 <a href="//ecsource.jove.com/files/ftp_upload/56812/56812fig2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
在 bicelles 的各向同性胶体悬浮的情况下, 磷脂在双层中的排列所诱导的取向将会丢失, 使迟滞δ零化。bicelles 还必须对准, 以便在它们的双层中定位光学活性磷脂, 导致偏振光δ。因此, 双折射是一个敏感的工具, 量化的磁性 alignability 的多组件。垂直于磁场的 Bicelles 将产生一个正的, 而那些对齐的平行将产生一个负的. <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg"> </img> <img alt="Equation 5" src="/files/ftp_upload/56812/56812eq5.jpg"> </img>该符号取决于设置的对齐方式, 并且可以使用引用示例进行检查。

<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 是可调谐的圆盘状多组件, 由大量的脂质混合物组成。应用范围从膜蛋白结构研究的核磁共振 (NMR), 以纳米的发展, 包括形成光学活性和磁性可转换凝胶。这些技术需要对装配尺寸、磁响应和热阻进行高度控制。混合物 12-二-sn-glycero-3-phosphocholine (DMPC) 及其镧系离子 (Ln3 +) 螯合磷脂共轭, 12-二-snglycero-3-phospho-ethanolamine-diethylene triaminepentaacetate (甲醚-DTPA), 组装成高磁响应的组件, 如 DMPC/甲醚-DTPA/Ln3 + (摩尔比率 4:1: 1) bicelles。在双层中引入胆固醇 (-OH) 和类固醇衍生物会导致另一组提供独特物理化学性质的组件。对于某一特定的脂质成分, 磁性 alignability 与 bicelle 大小成正比。在3 +中的络合结果在大小和对准方向方面都产生了前所未有的磁响应。热可逆崩溃的圆盘状结构成泡后, 加热允许裁剪装配的尺寸, 通过挤出膜过滤器的定义孔径大小。磁性 alignable bicelles 通过冷却到5° c 再生, 导致装配尺寸由囊泡前体定义。在此, 解释了这个制造过程, 并通过在 5.5 T 磁场下的双折射测量来量化组件的磁性 alignability。双折射信号, 源自磷脂双层, 进一步使监测的多变化发生在双层。这种简单的技术是互补的核磁共振实验, 通常用于表征 bicelles。

non-extruded DMPC/甲醚-DTPA/Tm3 +的双折射信号 (摩尔比 4:1: 1) 在一个 5.5 T 磁场的加热和冷却循环期间, 从5到40° c 和背部以1° c/分钟 (图 6) 的速度监测样品。双折射结果在5° c 时确认了高磁对齐, 其值为 1.5 x 10-5, 为所报告的挤出系统的两倍强。6,7,23 DMPC 在24° c 以上的 T<s...

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

介绍了高磁响应稀土离子螯合多组件的制备方法。磁响应由装配尺寸决定, 它是通过纳米膜的挤压而定制的。采用双折射测量、核磁共振和小角度中子散射技术对组件的磁 alignability 和温度诱发的结构变化进行监测。

设计磁响应稀土离子螯合磷脂组件的制备方法及双折射测量

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Layer semiconductors with easily processed two-dimensional (2D) structures exhibit indirect-to-direct bandgap transitions and superior transistor ...

Synthesis and Functionalization of 3D Nano-graphene Materials: Graphene Aerogels and Graphene Macro Assemblies

合成和3D纳米石墨烯材料的功能化：石墨烯气凝胶和石墨烯宏组件

Efforts to assemble graphene into three-dimensional monolithic structures have been hampered by the high cost and poor processability of graphene. ...

Synthesis of Ionic Liquid Based Electrolytes, Assembly of Li-ion Batteries, and Measurements of Performance at High Temperature

离子液体基电解质的合成，装配锂离子电池，并在高温性能的测量

The chemical instability of the traditional electrolyte remains a safety issue in widely used energy storage devices such as Li-ion batteries. Li-ion ...

Magnetically Induced Rotating Rayleigh-Taylor Instability

磁感应旋转瑞利 - 泰勒不稳定性

Classical techniques for investigating the Rayleigh-Taylor instability include using compressed gasses1, rocketry2 or linear electric motors3 to reverse ...

Focused Ion Beam Fabrication of LiPON-based Solid-state Lithium-ion Nanobatteries for In Situ Testing

Focused Ion Beam Fabrication of LiPON-based Solid-state Lithium-ion Nanobatteries for In Situ Testing

基于 LiPON 的固态锂离子 Nanobatteries 的聚焦离子束制备方法在原位测试中的研究

Solid-state electrolytes are a promising replacement for current organic liquid electrolytes, enabling higher energy densities and improved safety of ...

Three-electrode Coin Cell Preparation and Electrodeposition Analytics for Lithium-ion Batteries

锂离子电池三电极硬币电池制备及电沉积分析

As lithium-ion batteries find use in high energy and power applications, such as in electric and hybrid-electric vehicles, monitoring the degradation and ...

Measuring Magnetically-Tuned Ferroelectric Polarization in Liquid Crystals

液晶中磁调谐铁电极化的测量

Materials showing coupling phenomena between magnetism and (ferro)electricity, i.e., magnetoelectric effects, have attracted a great deal of attention due ...