Name: assembly and characterization of biomolecular memristors consisting of ion channel-doped lipid membranes
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国家科学基金会赠款 nsf eccs-1631472 提供了财政支助。对 g. j. t.、c. d. s.、a. b. 和 c. p. c. 的研究部分是由美国能源部的 UT-Battelle, llc 管理的橡树岭国家实验室实验室指导研究和发展方案赞助的。这项研究的一部分是在纳米酶材料科学中心进行的, 该中心是 doe 科学用户设施办公室。

1. 一般说明和注意事项
<ol><li>选择合适的、未损坏的测量器/混合玻璃器皿 (烧瓶、烧杯等) 和其他实验室器具 (飞溅、铲子等) 使用。</li>
	<li>小心处理玻璃器皿, 避免损坏, 并戴上乳胶或丁腈手套, 以避免污染玻璃制品与残留物从指尖, 并保护您的皮肤。</li>
	<li>使用洗涤剂溶液和水彻底清洁所选择的玻璃制品, 用软瓶刷擦洗, 直到清洁, 所有残留物被去除。</li>
	<li>用自来水彻底冲洗, ?...

<ol>
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本文提出了一种基于离子通道掺杂合成生物膜在油中两滴水之间形成的生物分子记忆器的组装和表征方案。设计和研究了软物质双端子器件: 1) 克服了与固态技术相关的限制, 如高噪声、高能耗、高开关电压, 2) 更密切地模仿了组合、结构, 生物突触的切换机制, 3) 探索固体装置不表现出的生物突触可塑性的机制和特征。
液滴界面双层技术21, 代表了目前技术<su...

人们普遍认识到, 生物突触是大脑高效和巨大平行的原因, 因为它们能够以高度适应性的方式学习和处理信息。这种协调的功能产生于多个高度复杂的分子机制, 这些机制驱动短期和长期突触可塑性1、2、3、4、5。神经形态计算系统的目标是在接近大脑密度、复杂性和能量效率的水平上模拟突触功能, 这是下一代类似大脑的计算机所需要的 6,7,8. 然而, 使用传统的电子电路元件复制突触特征几乎是不可能的, 而是需要设计和制造能够适应传入信号和记忆的新硬件元件信息历史9。这些类型的突触启发硬件被称为 mem-tem-mecemtem-memtemtemt"9 、10、11 (内存元素的缩写), 根据 di ventra 等人的说法, 这些硬件是被动的,双终端设备, 其电阻、电容或电感可以根据外部刺激重新配置, 并且可以记住先前的状态11。为了达到接近大脑能量消耗的水平, 这些元素应该使用类似的材料和机制来实现突触可塑性12。
迄今为止, 双端记忆电阻器13,14,15主要是使用互补金属氧化物半导体 (cmos) 技术制造的, 其特点是开关电压高、噪声高。由于高功耗和低密度, 该技术不能很好地扩展。为了解决这些限制, 最近建造了多个有机和聚合物记忆电阻器。然而, 由于通过导电聚合物基体 16,17 进行耗时的离子扩散, 这些器件的开关动力学明显变慢。因此, 基于 cmos 的记忆装置和有机记忆装置模仿突触激发的功能的机制是高度现象学的, 只包含一些突触功能, 如尖峰定时依赖可塑性 (stdp)1 8日, 同时忽视了其他关键特征, 这些特征也在使大脑成为一台强大而高效的电脑方面发挥着至关重要的作用, 比如突触前、短期可塑性19日。
最近, 我们推出了一类新型的记忆装置12 , 其特点是在仿生脂质膜中加入了电压激活肽, 模拟了生物分子组成、膜结构和离子通道触发开关生物突触的机制20。 在这里, 我们描述了如何组装和电气询问这些双终端设备, 特别侧重于如何评估短期可塑性, 以便在在线学习应用程序12中实现。器件组装是基于液滴界面双层 (dib)21方法, 近年来被广泛用于研究模型膜21和膜结合离子通道22,23的生物物理学. 24, 并作为发展刺激反应材料的基石 25,26。我们详细描述了那些对神经形态应用感兴趣, 但在生物材料或膜生物学方面经验有限的人的膜组装和审讯过程。该协议还包括对特性描述过程的完整描述, 考虑到装置27的动态和可重新配置的电气特性, 该过程与组装过程同样重要。这里描述的程序和代表性的结果是基础, 为一类新的低成本、低功耗、基于脂质界面的软记忆元件和其他生物分子, 用于神经形态计算、自主结构和系统的应用,甚至是自适应脑机接口。

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			<td height="21" style="height:22px;width:377px;">1,2-diphytanoy-sn-glycero-3-phosphocholine (DPhPC)</td>
			<td style="width:206px;">Avanti Polar Lipids</td>
			<td style="width:183px;">850356P/850356C</td>
			<td style="width:663px;">Purchased as lyophilized powder (P) or in chloroform (C)&#160;</td>
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		<tr height="21">
			<td height="21" style="height:22px;width:377px;">Agarose&#160;</td>
			<td style="width:206px;">Sigma-Aldrich</td>
			<td style="width:183px;">A9539</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Agarose (0.5g Agarose Tablets)</td>
			<td style="width:206px;">Benchmark</td>
			<td style="width:183px;">A2501</td>
			<td>You can either use the powder form or the tablets&#160;</td>
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			<td height="21" style="height:21px;width:377px;">Alamethicin&#160;</td>
			<td style="width:206px;">AG Scientific</td>
			<td style="width:183px;">A-1286</td>
			<td></td>
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			<td height="21" style="height:21px;width:377px;">Analytical balance&#160;</td>
			<td style="width:206px;">Mettler Toledo</td>
			<td>ME204TE/00</td>
			<td></td>
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			<td height="21" style="height:21px;width:377px;">Axopatch 200B Amplifier&#160;</td>
			<td style="width:206px;">Molecular Devices</td>
			<td style="width:183px;">-</td>
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			<td height="21" style="height:22px;width:377px;">BK Precision 4017B 10 MHz DDs Sweep/Function Generator</td>
			<td style="width:206px;">Digi-Key</td>
			<td style="width:183px;">BK4017B-ND</td>
			<td></td>
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			<td height="21" style="height:21px;width:377px;">Borosilicate Glass Capillaries</td>
			<td style="width:206px;">World Precision Instruments</td>
			<td style="width:183px;">1B100F-4</td>
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			<td height="21" style="height:21px;width:377px;">Brain Total Lipid Extracts (Porcine)</td>
			<td style="width:206px;">Avanti Polar Lipids</td>
			<td style="width:183px;">131101</td>
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			<td height="21" style="height:21px;width:377px;">DigiData 1440A system</td>
			<td style="width:206px;">Molecular Devices</td>
			<td style="width:183px;">-</td>
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			<td height="21" style="height:21px;width:377px;">Extruder Set With Holder/Heating Block&#160;</td>
			<td style="width:206px;">Avanti Polar Lipids</td>
			<td style="width:183px;">610000</td>
			<td>This includes a mini-extruder, 2 syringes, 100 PC membranes, 100 filter supports, and 1 holder/heating block</td>
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			<td height="21" style="height:21px;width:377px;">Freezer (-20 &#176;C)</td>
			<td style="width:206px;">VWR International</td>
			<td style="width:183px;">SCUCBI0420AD</td>
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			<td height="21" style="height:21px;width:377px;">Glassware</td>
			<td style="width:206px;">VWR International</td>
			<td style="width:183px;">-</td>
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			<td height="21" style="height:21px;width:377px;">Hexadecane, 99%</td>
			<td style="width:206px;">Sigma-Aldrich</td>
			<td style="width:183px;">544-76-3</td>
			<td></td>
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			<td height="21" style="height:21px;width:377px;">Isopropyl Alcohol</td>
			<td style="width:206px;">VWR International</td>
			<td style="width:183px;">BDH1133-4LP</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Microelectrode Holder&#160;</td>
			<td style="width:206px;">World Precision Instruments</td>
			<td style="width:183px;">MEH1S</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">MOPS</td>
			<td style="width:206px;">Sigma-Aldrich</td>
			<td style="width:183px;">M1254</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:25px;width:377px;">Nitrogen (N2) Gas</td>
			<td style="width:206px;">Airgas</td>
			<td style="width:183px;">UN1066</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Parafilm M All-Purpose Laboratory Film</td>
			<td style="width:206px;">Parafilm</td>
			<td style="width:183px;">PM999</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Powder Free Soft Nitrile Examination Gloves&#160;</td>
			<td style="width:206px;">VWR International</td>
			<td style="width:183px;">CA89-38-272</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Precleaned Microscope Sildes&#160;</td>
			<td style="width:206px;">Fisher Scientific&#160;</td>
			<td style="width:183px;">22-267-013</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Refrigirator (4 &#176;C)</td>
			<td style="width:206px;">VWR International</td>
			<td style="width:183px;">SCUCFS-0504G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Silver wire</td>
			<td style="width:206px;">GoodFellow</td>
			<td>147-346-94</td>
			<td>Different diameters could be used depending on the application&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Sodium Chloride (KCl)</td>
			<td style="width:206px;">Sigma-Aldrich</td>
			<td style="width:183px;">P3911</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">Stirring Hot Plate</td>
			<td style="width:206px;">Thermo Scientific&#160;</td>
			<td style="width:183px;">SP131325</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">VWR Light-Duty Tissue Wipers</td>
			<td style="width:206px;">VWR International</td>
			<td style="width:183px;">82003-820</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:377px;">VWR Scientific 50D Ultrasonic Cleaner</td>
			<td style="width:206px;">VWR International</td>
			<td style="width:183px;">13089</td>
			<td></td>
		</tr>
	</tbody>
</table>

assembly and characterization of biomolecular memristors consisting of ion channel-doped lipid membranes

能够在合成电路元素中重现突触功能对于寻求模仿具有同等效率和密度的大脑认知能力的神经形态计算系统至关重要。迄今为止, 硅基三端晶体管和双端记忆电阻器在神经形态电路中得到了广泛的应用, 这在很大程度上是由于它们能够进行同地定位的信息处理和存储器。然而, 这些设备无法实现大脑的互连性和复杂性, 因为它们耗电, 无法模拟关键的突触功能, 并受到高噪声和高开关电压的影响。为了克服这些限制, 我们开发并表征了一种生物分子记忆电阻器, 该记忆器模拟了生物突触的组成、结构和开关特性。在这里, 我们描述了生物分子记忆器的组装和表征过程, 其中包括 5 nw 厚的脂质双层在油中的脂质功能化水滴之间形成的, 并掺杂了电压活化的醛氧西林肽。虽然类似的组装协议已被用来研究生物物理特性的液支持脂膜和膜结合离子通道, 本文的重点是关键的修改液滴接口双层法必不可少的实现稳定的电阻器性能。具体而言, 我们描述了脂质体的制备过程和在脂质双层膜中加入类似霉素肽的过程, 以及每个成分的适当浓度及其对记忆电阻器整体响应的影响。我们还详细介绍了生物分子记忆器的表征过程, 包括测量和分析通过循环伏安法获得的记忆电流-电压关系, 以及短期可塑性和学习响应阶跃化电压脉冲列车。

图 1显示了用于组装和表征生物分子记忆器的实验装置。图1b所示, 将电极的自由端降低到油层底部, 有助于最大限度地减少电极和液滴的振动, 从而导致测量电流和双层面积的变化, 特别是在情况下在那里加热油可以产生油的对流流动。图 2显示了组装 ag/agcl 电线、类毛细血管、电极和微型移液器支架的过程和结果。该装置位于正?...

Watch this Scientific Journal Video about Assembly and Characterization of Biomolecular Memristors Consisting of Ion Channel-doped Lipid Membranes at JoVE.com

Assembly and Characterization of Biomolecular Memristors Consisting of Ion Channel-doped Lipid Membranes

软、低功耗、生物分子记忆电阻利用生物突触的相似成分、结构和切换机制。本文提出了一种从油中水滴之间形成的绝缘脂质双层层中获得的生物分子记忆器的组装和表征方案。电压活化的阿米西林肽的加入导致膜上的记忆离子电导。

离子通道掺杂脂质膜组成的生物分子记忆器的组装与表征

The ability to recreate synaptic functionalities in synthetic circuit elements is essential for neuromorphic computing systems that seek to emulate the ...

我们的协议描述了如何组装和电气特征肽掺杂生物膜，密切模仿生物突触的组成，结构和传输特性，并表现出可调调的记忆电阻。该技术使用户能够根据与生物突触和离子通道相关的时间尺度和激发水平评估工程系统中的活动依赖性、记忆电阻和短期可塑性。该技术为描述含有电压激活离子通道的仿生膜提供了一个框架，使其适用于各种细胞传输过程的表征，包括神经元中的细胞传输过程。我们向新研究人员的建议是，首先要熟练地制备脂质溶液，并在线型电极上组装液滴界面双层。亲眼看到液滴点胶和电极上的定位过程简化了这种双层形成技术，使所有人都能立即获得这种技术。演示这个程序将是约瑟夫·纳杰姆博士，一个博士后从我的实验室。首先，在微离心管中准备阿拉米西辛库存溶液，将乙醇中的阿拉米辛肽粉溶解到每毫升2.5毫克的最终浓度。涡流管短暂混合，并储存在零下20摄氏度的冰柜中的库存溶液。在1.5毫升安全锁管中，在溶液 A 的 99 微升中加入一微升阿拉米辛库存溶液，以实现脂体悬浮液中最终的阿拉米辛浓度为 13 微摩尔。漩涡混合良好。由此产生的肽脂体溶液是溶液B.Mix117微升溶液B与10微升溶液B，实现最终阿拉米西明素浓度的一微摩尔，然后涡流混合良好。将生成的解决方案称为 C.将解决方案 B 和 C 存储在 4 摄氏度。将一毫米厚的 25 倍 75 毫米玻璃侧放在倒置显微镜的舞台上。将几滴六烷油分配到玻璃滑梯的中心，然后将储油罐直接放在玻璃滑梯上的油上。完全用六烷油向储油罐加注。确保储液罐位于客观透镜上方。然后将电极支架插入安装在微操纵器上的电流放大器的头部阶段。微操纵器可最大限度地降低电极长度和电噪声。然后，将用第二根银氯化锡线的玻璃微管支架安装到另一个微操纵器上。使用微操纵器，定位电极，使银-银-氯化线涂覆的尖端完全浸入油藏中，形成相似的垂直平面。对齐两个电极，将其分开几毫米。要形成脂质双层，请将电极垂直移动到油相中。使用微管在每根导线上沉积 200 纳米的脂质溶液 A。等待三到五分钟，以便水油界面上自发进行脂质单层组装。如果周围的油密度足够低，液滴可能会下垂。之后，降低电极以重新下潜，直到两个电极的末端几乎接触储油罐的底部。然后形成双层，水平移动电极，使液滴接触。为了获得捏合、歇斯底里、电流-电压的关系，请使用函数发生器将三角形或正弦电压波形应用到与溶液 A. 组装的无阿拉米辛脂质膜上。要记录面间脂双层层的大小，请测量计算机上的脂质膜的直径，或记录由 10 赫兹、10 毫伏三角波产生的峰峰值电流振幅来计算膜面积。将导线从油相中取出，以去除不含阿拉米西辛的液滴。使用溶液C添加新的水滴，形成脂质双层液。根据方波电流振幅，使用微操纵器调整液滴之间的接触，使双层器具有与早期形成的相似区域。然后应用 10 赫兹和 10 毫伏电压波形式，并记录诱导电流响应。要使用定制编程软件和模拟电压源进行脉冲实验，请及时和关闭时间生成具有特定高振幅和低振幅的电压脉冲。记录电流以响应应用的脉冲。电流与电压的图显示电压偏置对无阿拉米辛脂质双层体应用时的非零电流响应。添加 0.017 赫兹，该频率的阻抗由膜电阻控制。对于高绝缘膜，显示低欧姆电流响应。在含有阿拉米辛肽的两个液滴之间形成的脂质双层体图显示，在高于100毫伏的插入阈值的电压下，电流呈指数级增长。在高电压下，位于脂质双层体表面的阿拉米西辛肽插入膜并聚集形成导电孔隙。两极的对称电流响应是由于从膜的对面插入和聚合了单独的肽群。电容电流必须从总电流中减去，才能获得压流、挤压的滞后电流电压响应。生物分子记忆器对后续电压脉冲的响应，尽管在每个关闭时间间歇性地恢复绝缘状态，但同时提高导电性。目前的刺激措施和先前的刺激措施都促成了目前的增长。两个液滴上的粘合单层必须形成，然后将它们聚集在一起形成双层体。如果水滴太早地聚集在一起，它们就会聚集在一起，并且没有形成双层液。我们现在在设计和制造由固态神经元组成的微流体基础神经网络，这些神经元由基于膜的突触连接，支持 ORNL 高性能超级计算机的网络同化优化。这些分子是第一个具有生物突触的组成、结构、开关机制和离子传输的。因此，它们提供了一个生物分子基础，为大脑一样的计算和记忆提供了补充。

Research

Education

JoVE Core

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Bioengineering

Unit 1

Alternating-Current Circuits

SCIENTISTS IN ACTION

Biomolecular Detection employing the Interferometric Reflectance Imaging Sensor (IRIS)

Biomolecular Detection employing the Interferometric Reflectance Imaging Sensor IRIS

生物分子检测的干涉反射成像传感器（IRIS）

The sensitive measurement of biomolecular interactions has use in many fields and industries such as basic biology and microbiology, ...

科研

生物工程

Synthesis, Assembly, and Characterization of Monolayer Protected Gold Nanoparticle Films for Protein Monolayer Electrochemistry

单层的合成，装配和表征蛋白质单分子膜的电化学保护的金纳米粒子薄膜

Colloidal gold nanoparticles protected with alkanethiolate ligands called monolayer protected gold clusters (MPCs) are synthesized and subsequently ...

Isolation of Cellular Lipid Droplets: Two Purification Techniques Starting from Yeast Cells and Human Placentas

隔离细胞脂滴：两个纯化技术从酵母细胞与人类的胎盘开始

Lipid droplets are dynamic organelles that can be found in most eukaryotic and certain prokaryotic cells. Structurally, the droplets consist of a core of ...

Isolation and Characterization Of Chimeric Human Fc-expressing Proteins Using Protein A Membrane Adsorbers And A Streamlined Workflow

使用蛋白质膜吸收器和流线型工作流的奇美人 Fc 表达蛋白质的隔离和特征

Laboratory scale to industrial scale purification of biomolecules from cell culture supernatants and lysed cell solutions can be accomplished using ...

Micropatterning and Assembly of 3D Microvessels

缩微和3D微血管大会

In vitro platforms to study endothelial cells and vascular biology are largely limited to 2D endothelial cell culture, flow chambers with polymer or glass ...

Merging Ion Concentration Polarization between Juxtaposed Ion Exchange Membranes to Block the Propagation of the Polarization Zone

并列的合并离子交换膜之间的离子浓差极化，以块极化区的传播

The ion concentration polarization (ICP) phenomenon is one of the most prevailing methods to preconcentrate low-abundance biological samples. The ICP ...

A Method for Growing Bio-memristors from Slime Mold

一种从煤泥霉菌中生长生物器的方法

Our research is aimed at gaining a better understanding of the electronic properties of organisms in order to engineer novel bioelectronic systems and ...

Detection of Endotoxin in Nano-formulations Using Limulus Amoebocyte Lysate (LAL) Assays

Detection of Endotoxin in Nano-formulations Using Limulus Amoebocyte Lysate LAL Assays

用利穆鲁布莱巴细胞裂解液 (lal) 法检测纳米配方中的内毒素

When present in pharmaceutical products, a Gram-negative bacterial cell wall component endotoxin (often also called lipopolysaccharide) can cause ...

Growth and Characterization of Irradiated Organoids from Mammary Glands

乳腺辐照有机体的生长与表征

Organoids derived from the digested tissue are multicellular three-dimensional (3D) constructs that better recapitulate in vivo conditions than cell ...

Focused Ion Beam Lithography to Etch Nano-architectures into Microelectrodes

聚焦 Ion 光束平版印刷到蚀刻纳米架构到微电极

With advances in electronics and fabrication technology, intracortical microelectrodes have undergone substantial improvements enabling the production of ...