Name: pressure-polishing pipettes for improved patch-clamp recording
Uploaded: 2008-10-22T00:00:00
Description: Watch this Scientific Journal Video about Pressure-polishing Pipettes for Improved Patch-clamp Recording at JoVE.com

<p>压力抛光技术的开发与SR Lockery合作。 </p>

<p class="jove_step">拉移液器</p><p>使用灯丝一个车夫，如萨特的P - 97火焰山/布朗微电极拖轮，拉约10-20移液器。 </p><ol><li>选择你的玻璃。我们使用硼硅（BF150 - 86 - 10，OD = 1.5毫米，编号= 0.86毫米）。商店的玻璃，仔细，所以保持清洁和无尘。 </li><li>设计一个拉动的程序。其主要目的是尖端张开约所需的大小，这将取决于应用程序。萨特<em>的</em> P <em>- 97移液器食谱</em> <sup>2</sup>是一个很好的参考。 </li><li>在显微镜下?...

<ol>
	<li>Goodman, M. B., Lockery, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pressure+polishing:+a+method+for+re-shaping+patch+pipettes+during+fire+polishing. .">Pressure polishing: a method for re-shaping patch pipettes during fire polishing. .</a> <em style='font-style: italic'>J Neurosci Methods</em>. <b>100</b>, 13-15 (2000).</li></ol>

<p>我们最初开发这种方法使移液器有用2-3μm的神经元<em>在</em> C片钳记录<em>线虫</em> <sup>1。</sup>它是在古德曼实验室常规使用。我们的压力不仅波兰移液器录制从微小的<em>C。线虫</em>的神经元，但也获得macropatch表示在爪蟾卵母细胞的离子通道和单通道录音。对于一个给定的尖端开口直径，生硬的造型产生一个低电阻的吸管。下吸液管电阻会导致较低的串联电阻和低噪声录音。 </p>

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		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Glass Capillaries</td>
<td>Consumable</td>
<td>Sutter Instruments</td>
<td>BF150-85-10 (or similar)</td>
<td>The method works with any type of capillary except those made from quartz.  However, we recommend using capillaries with a filament.</td>
</tr>
<tr>
<td>Inverted Microscope</td>
<td>Microscope</td>
<td>Leica</td>
<td>DM IL (or similar)</td>
<td>Any simple inverted microscope will do.  However, it is essential to have sufficient magnification to see pipettes during polishing.  We use a 100x metallurgical lens with a long working distance for this purpose.</td>
</tr>
<tr>
<td>Microforge Kit</td>
<td>Tool</td>
<td>ALA Science</td>
<td>CPM-2</td>
<td>Supplies polishing coil, controller and foot pedal.</td>
</tr>
<tr>
<td>Air pressure source</td>
<td>Tool</td>
<td>ALA Science</td>
<td>PR-60</td>
<td>Accessory for providing high pressure, includes a suitable valve.  Alternatively, compressed air from the building can be used together with a filter/regulator (set to 40 PSI) and a 4-way valve (see below)</td>
</tr>
<tr>
<td>4-way valve</td>
<td>Tool</td>
<td>Cole-Parmer</td>
<td>A-98150-01</td>
<td>Fittings will be needed to couple valve to tubing</td>
</tr>
<tr>
<td>Pipette Holder with Luer Connector</td>
<td>Tool</td>
<td>World Precision Instruments</td>
<td>MPH-6S20</td>
<td>Use a male luer-to-barb fitting to connect pipette holder to high-pressure tubing</td>
</tr>
<tr>
<td>Tygon&#174; tubing</td>
<td>Tool</td>
<td>Cole-Parmer</td>
<td>06408-62</td>
<td>Use this tubing to connect pipette holder to 4-way valve
</td>
</tr>		</tbody>
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pressure-polishing pipettes for improved patch-clamp recording

<p class="jove_content">压力抛光塑造膜片钳记录玻璃移液器的方法。首先，我们开发了适合小（&lt;3米）的神经元细胞体的录音编造移液器此方法。基本原理是类似玻璃吹制，并结合空气的压力和热量，修改形状由传统的微量车夫准备的补丁移液器。它可以应用到所谓的软（碱石灰）和硬（硼硅）眼镜。一般来说，抛光压力，可以减少25％，吸液管电阻不减少的尖端张开“（Goodman和Lockery，2000）直径。它可以适用于几乎任何类型的玻璃，只需要增加一个高压阀门及配件一个microforge。这种技术是必要的记录，从超小细胞（&lt;5米）和，还可以通过单声道录音，最大限度地减少吸液管电阻。钝的形状也很有用穿孔膜片钳记录，因为这在一个较大的膜穿孔的气泡的笔尖形状结果。 </p>

Watch this Scientific Journal Video about Pressure-polishing Pipettes for Improved Patch-clamp Recording at JoVE.com

Pressure-polishing Pipettes for Improved Patch-clamp Recording

<p class="jove_content">这是一个指导修改形状的玻璃微量。具体来说，通过热和空气压力的锥度不增加尖端张开扩大，导致较低的吸液管电阻。这是至关重要的，以获得小细胞低噪音录音，但在许多应用中非常有用。</p>

改进的膜片钳记录的压力抛光移液器

Pressure-polishing is a method for shaping glass pipettes for patch-clamp recording.  We first developed this method for fabricating pipettes suitable for ...

Research

JoVE Journal

Biology

Methods for Patch Clamp Capacitance Recordings from the Calyx

膜片钳从花萼电容录音方法

We demonstrate the basic techniques for presynaptic patch clamp recording at the calyx of Held, a mammalian central nervous system nerve terminal. ...

科研

生物学

Patch Clamp Recording of Ion Channels Expressed in  Xenopus Oocytes

补丁钳爪蟾卵母细胞中表达的离子通道

Since its development by Sakmann and Neher 1, 2, the patch clamp has become established as an extremely useful technique for electrophysiological ...

Making Patch-pipettes and Sharp Electrodes with a Programmable Puller

膜片吸液管和夏普电极与一个可编程的拖轮

Glass microelectrodes (also called pipettes) have been a workhorse of electrophysiology for decades.  Today, such pipettes are made from glass capillaries ...

Patch Clamp and Perfusion Techniques for Studying Ion Channels Expressed in <em>Xenopus</em> oocytes

Patch Clamp and Perfusion Techniques for Studying Ion Channels Expressed in Xenopus oocytes

膜片钳和灌注技术研究离子通道的表达<em>爪蟾</em>卵母细胞

The protocol presented here is designed to study the activation of the large conductance, voltage- and Ca2+-activated K+ (BK) channels. The protocol may ...

Aseptic Laboratory Techniques: Volume Transfers with Serological Pipettes and Micropipettors

无菌实验室技术：量与血清学移液器和Micropipettors的转移

Microorganisms are everywhere - in the air, soil, and human body as well as on inanimate surfaces like laboratory benches and computer keyboards. The ...

Exploring Arterial Smooth Muscle Kv7 Potassium Channel Function using Patch Clamp Electrophysiology and Pressure Myography

探讨动脉平滑肌的KV7钾离子通道功能使用膜片钳电生理和压力Myography

Contraction  or relaxation of smooth muscle cells within the walls of resistance arteries  determines the artery diameter and thereby controls flow of ...

Giant Liposome Preparation for Imaging and Patch-Clamp Electrophysiology

巨脂质体制剂成像和膜片钳电

The reconstitution of ion channels into chemically defined lipid membranes for electrophysiological recording has been a powerful technique to identify ...

One-channel Cell-attached Patch-clamp Recording

单通道细胞贴附膜片钳记录

Ion channel proteins are universal devices for fast communication across biological membranes. The temporal signature of the ionic flux they generate ...

Reconstitution of a Transmembrane Protein, the Voltage-gated Ion Channel, KvAP, into Giant Unilamellar Vesicles for Microscopy and Patch Clamp Studies

跨膜蛋白的重组，电压门控离子通道，KvAP，进入巨人单层囊的显微镜和膜片钳研究

Giant Unilamellar Vesicles (GUVs) are a popular biomimetic system for studying membrane associated phenomena. However, commonly used protocols to grow ...

Implementing Patch Clamp and Live Fluorescence Microscopy to Monitor Functional Properties of Freshly Isolated PKD Epithelium

实施膜片钳和活荧光显微镜监视新鲜分离的PKD上皮细胞的功能特性

Cyst initiation and expansion during polycystic kidney disease is a complex process characterized by abnormalities in tubular cell proliferation, luminal ...