Name: custom-made microdialysis probe design
Uploaded: 2015-07-21T15:00:00.000+00:00
Duration: 5 min 38 s
Description: Watch this Scientific Journal Video about Custom-made Microdialysis Probe Design at JoVE.com

The authors are grateful to G. Barka (SunChrom GmbH, Friedrichsdorf, Germany) for his support and for providing the epoxy glue. Furthermore, the authors acknowledge Fresenius Medical Care (Bad Homburg, Germany) for supplying the Capillary Haemodiafilter FXCorDiax. Funding was obtained from Goethe University of Frankfurt.

1.准备聚四氟乙烯管<ol><li>缩短聚四氟乙烯管进入2.5厘米片。使用纸的规模估算物理尺寸。 （参见图3a） </li><li>切1结束在一个角度，以方便连接出口管。 （参见图3a） </li><li>粗糙化的PTFE管材通过使用砂纸，以允许的环氧树脂胶附着。 </li></ol> 2.准备熔融石英<ol><li>缩短熔融石英到2.25厘米片。 （参见图3a） </li></ol> 3.插入熔融石英进入聚四氟乙烯管<ol><li>插入地下30插管到中空PTFE纤维在从扁平端部的距离为1厘米刺穿它。弯曲聚四氟乙烯管，以简化流程。 （参见图3b） </li><li>将熔融二氧化硅到使用套管为指导的PTFE管材。 （参见图3c） </li><li>轻轻取出套管和安全熔融石英。 （见图3d） </li><li>熔融二氧化硅应该突出为5mm的平端。 （参见图3e） </li><li>胶代替熔融二氧化硅在穿刺部位通过氰基丙烯酸酯胶，并允许它变硬过夜。 （参见图3e） </li></ol> 4.准备环氧胶（双组份胶） <ol><li>加入30份黄色固化剂，以70份黑色环氧树脂胶通过细针的帮助下，轻轻地混合。它直接就可以使用。 </li></ol> 5.运用膜（所有进一步的步骤必须执行在显微镜下和规模纸是用来估计物理尺寸） <ol><li>拉聚砜膜小心在熔融二氧化硅，然后拉膜进入聚四氟乙烯管和用锋利的手术刀调整膜至5.5毫米。 （参见图3f） </li><li>用细笔设置在膜上的标记点定义为2毫米的交换长度从膜的端部。 </li><li>适用于少量环氧树脂胶覆盖用细针的帮助下在膜的尖端。 （参见图3g） </li><li>在步骤5.3中使用的相同细针）与环氧胶的帮助下关闭掉透析膜和PTFE纤维之间的联合。 （参见图3g） </li><li>添加在膜足够环氧树脂胶到标志点，以确保2mm的交换长度。允许它变硬过夜。 （参见图3g）测定的交换长度之前，该膜为5.5毫米。施加环氧树脂胶在膜后，只有2毫米膜可用于扩散。 </li></ol> 6.应用金属套<ol><li>通过弯曲钳帮助削减将25g针头1厘米的金属套管件。 </li><li>拉1厘米金属套筒（25 G）的到熔融二氧化硅的剩余部分。金属套后使进口管道与MICRODIA连接裂解探头。 （ 见图3H） </li><li>金属密封套筒和聚四氟乙烯管，其余环氧胶之间的联合。允许其硬化至少24小时。 （ 见图3I） </li><li>施加热胶围绕探针的分叉为额外的固定和稳定。 注:该探针的入口内部体积是0.045微升和出口内部容积为2.5微升。因此，用1微升/ min流速，需要连接入口管开始的透析液的收集之后约2.5分钟。本协议，探头装配过程产生微透析探头是95％可靠。 </li></ol>的体外回收实验7.性能<ol><li>浸探针与截止的30 kDa的和2mm的交换长度分成含有1mM葡萄糖和1mM乳酸和，在一个单独的实验脑脊液组成的储备溶液，在含有100 CSF的纳米ATP和100nM的乙酰胆碱。 </li><li>灌注探头学联。设定流速为1微升/分钟，进行在室温（22℃）的实验。允许探针平衡1小时，在原料溶液，并收集样品，每30分钟之后，并储存在-80℃直至测量。 </li><li>对于体外回收率的计算，与已知分析物浓度的储备溶液的透析液比较分析物的浓度。表达结果的物质浓度的透析液与原液比例。 </li></ol>

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	<li>Feldberg, W., Malcom, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experiments+on+the+site+of+action+of+tubocurarine+when+applied+via+the+cerebral+ventricles.">Experiments on the site of action of tubocurarine when applied via the cerebral ventricles.</a> J Physiol. 149, 58-77 (1959).</li><li>Gaddum, 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=Push-pull+cannulae.">Push-pull cannulae.</a> J Physiol. 155 (1 P), (1961).</li><li>Bito, L., Davson, H., Levin, E., Murray, M., Snider, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+concentrations+of+free+amino+acids+and+other+electrolytes+in+cerebrospinal+fluid,+in+vivo+dialysate+of+brain,+and+blood+plasma+of+the+dog.">The concentrations of free amino acids and other electrolytes in cerebrospinal fluid, in vivo dialysate of brain, and blood plasma of the dog.</a> J Neurochem. 13 (11), 1057-1067 (1966).</li><li>Delgado, J. M., Lerma, J., Mart&#237;n del R&#237;o, R., Sol&#237;s, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dialytrode+technology+and+local+profiles+of+amino+acids+in+the+awake+cat+brain.">Dialytrode technology and local profiles of amino acids in the awake cat brain.</a> J Neurochem. 42 (5), 1218-1228 (1984).</li><li>Ungerstedt, U., Pycock, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+correlates+of+dopamine+neurotransmission.">Functional correlates of dopamine neurotransmission.</a> Bull Schweiz Akad Med Wiss. 30 (1-3), 44-55 (1974).</li><li>Ungerstedt, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microdialysis--principles+and+applications+for+studies+in+animals+and+man.">Microdialysis--principles and applications for studies in animals and man.</a> J Intern Med. 230 (4), 365-373 (1991).</li><li>Bourne, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracerebral+microdialys:+30+years+as+a+tool+for+the+neuroscientist.">Intracerebral microdialys: 30 years as a tool for the neuroscientist.</a> Clin Exp Pharmacol Physiol. 30 (1-2), 16-24 (2003).</li><li>Lietsche, J., Gorka, J., Hardt, S., Karas, M., Klein, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Self-built+microdialysis+probes+with+improved+recoveries+of+ATP+and+neuropeptides.">Self-built microdialysis probes with improved recoveries of ATP and neuropeptides.</a> J Neurosci Methods. 237, 1-8 (2014).</li><li>Chefer, V. I., Thompson, A. C., Zapata, A., Shippenberg, 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=Overview+of+Brain+Microdialysis.">Overview of Brain Microdialysis.</a> Curr Protoc Neurosci. 7, Unit 7.1 (2009).</li><li>Neurochem, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Brain+Microdialysis.">Brain Microdialysis.</a> J. Neurochem. 52 (6), 1667-1679 (1989).</li><li>Jolly, D., Vezina, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+microdialysis+in+the+rat:+low+cost+and+low+labor+construction+of+a+small+diameter,+removable,+concentric-style+microdialysis+probe+system.">In vivo microdialysis in the rat: low cost and low labor construction of a small diameter, removable, concentric-style microdialysis probe system.</a> J Neurosci Methods. 68 (2), 259-267 (1996).</li><li>Sumbria, R., Klein, J., Bickel, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+depression+of+energy+metabolism+after+microdialysis+probe+implantation+is+distinct+from+ischemia-induced+changes+in+mouse+brain.">Acute depression of energy metabolism after microdialysis probe implantation is distinct from ischemia-induced changes in mouse brain.</a> Neurochem Res. 36, 109-116 (2010).</li></ol>

The authors have nothing to disclose.

图3示出的示例性探针组件。正确内外径尺寸必须被密切油管观察（OD1.6毫米;编号350微米），熔融二氧化硅（外径105微米;编号40微米）和渗析膜（外径245微米;编号210微米）。同样重要的是，保持隔膜和（105微米）和膜和管（105微米）以及之间熔融二氧化硅之间的空间。如果值不同，在探头压力可以上升，并导致该膜的泄漏。 这里我们介绍一种微透析探针以2mm的交...

微透析是在神经科学研究中一个常用的技术。在过去的50年中，最小侵入性的微透析技术不断完善，成为一个行之有效的方法来监测外空间的局部浓度的小分子量化合物。几乎每间质液可以自由移动的动物进行调查。 Gaddum介绍，在20世纪60年代推拉技术。他修饰的方法从费尔德等人在其筒箭毒物通过套管终止于侧脑室和通过套管1收集的流出物也灌注。 Gaddum开发，其中套管由两个同心钢针植入在不同脑区和灌注通过溶液而同时除去来自周围的刀片2的神经元释放的神经递质的推挽技术。不幸的是，组织DAMA引起的尖端周围套管戈限于该方法的应用。作为这种方法的一个进一步推进，尾藤和同事提出了透析袋的方法，其中所收集的溶液，从周围组织中通过透析膜隔开。它们注入透析囊到狗脖子的皮下组织。透析袋的含量无蛋白质的，并且可以为离子和氨基酸3许多周后进行分析。接下来的发展是dialytrode，一个原始微透析探针，其最初由Delgado的于1972年4所述。最后，Ungerstedt和同事改善微透析探针的设计，以便它是较小的和位移较少的组织5。 同心微透析探针的行为类似于一个毛细血管。该系统由一个解决方案，包括对周围组织的流体的离子组成，同时缺乏所关注分析物不断灌注。钍Ë透析膜暴露于外部溶液或组织是半渗透性的。它允许的物质的被动扩散进入沿其浓度梯度6探头。渗透率依赖于许多变量，如分子量，形状，电荷和pH的化合物。它也被限制，由于该膜材料的性能，孔膜的尺寸和流速7。 图1和2示出了同心微透析探针。经由入口管导入金属套筒，其围绕熔融二氧化硅的灌注流体进入。内的金属套筒，它流顺着熔融二氧化硅和叶子它在它的前端。在透析膜和聚四氟乙烯（PTFE）-tubing（如特氟隆）之间的空间内的灌注液，然后向上流动。在这里，从组织的物质发生扩散环绕该膜。透析液通过PTF离开探头电子管道，其连接到出口管并可以收集。 微透析技术具有相对于其它体内技术的几个优点。探针构成与透析液不含有酶或细胞的结果的物理屏障。因此，没有必要进行分析前的洗脱物的纯化，以及分析物的无酶促降解发生。氧化降解可发生在管道的分析物的通过期间，但这通常可以通过加入抗氧化剂（ 例如抗坏血酸）的灌注液被阻止。可替换地，氧化性损伤的神经肽，例如，被有效地通过用尖更换出口管路，收集透析液8抑制。透析液可以直接与几乎任何类型的分析方法进行研究和多种分析物可以同时被收集。该系统可在清醒的动物中使用，几乎所有的大脑区域可以是检查。此外，通过探针灌注药物是可能的（retrodialysis）。然而，也有在微透析技术的局限性。偏低的时间分辨率不提供关于神经递质变化的实时信息。因为它是一种侵入性的技术，探针植入引起的手术创伤和麻醉，这会影响神经递质的浓度，在该步骤期间7,9,10是必需的。 在透析液的化合物浓度仅包括少量在细胞外流体中的实际化合物浓度。对于在细胞外流体中的未知化合物浓度的计算中，相对于在体外恢复已被计算。相对于个人在体外回收率为每个探头和每个化合物的测定是在开始体内实验前有必要的。为了这个目的，将探针浸入含的溶液的利益而灌注流体分析物是缺乏的感兴趣的分析物相同的溶液。在透析液测定化合物的浓度后，此数据必须被提到其在周围流体的浓度。几种物质在体外恢复判定可以同时9来实现。 许多神经科学的研究小组利用微透析技术来研究神经递质和代谢物在不同脑区的细胞外空间。因此，商业探针在神经科学研究环境有很大的需求。市售探头一大优点是高功能可靠性。该实验装置用于商业探头是公认和验证众多知名神经递质及代谢产物。然而，尽管可商购的微透析设备价格昂贵，并且在应用程序11的弹性较小，在第是工作一个微透析探针组件中详细介绍，它可以适用于任何应用，并且可以为不到10美元来制造。这种定制的探头是测试在各个脑区8调查同心圆微透析探针。 体外探针回收率的不同分子量（的100-1,600大范围）中，用物质不同的物理化学性质进行了比较。 在体外回收的葡萄糖，乳酸和乙酰胆碱测定，其分子量小于200道尔顿，ATP的具有分子量约500 Da和神经肽血管紧张素II，P物质和生长抑素具有分子量高于1,000达被执行。

<table><tbody><tr><td>Epoxy glue</td><td>SunChrom GmbH, Fiedrichsdorf, Germany</td><td></td><td>&amp;nbsp;&amp;nbsp;&amp;nbsp;</td></tr><tr><td>Fused silica ID 40 &amp;micro;m OD 105 &amp;micro;m</td><td>Ziemer-Chromatographie, Mannheim, Germany</td><td>Art. No: 6.040105</td><td></td></tr><tr><td>Polysulfone membrane (haemodialysis filter FX Cor Diax 600)</td><td>Fresenius Medical Care AG &amp;amp; Co. KGaA, Bad Homburg, Germany</td><td>REF: F00001593</td><td></td></tr><tr><td>Cyanacrylate glue (Pattex&amp;reg; superglue plastic)</td><td>Henkel AG&amp;amp;Co. KGaA, D&amp;uuml;sseldorf, Germany</td><td></td><td></td></tr><tr><td>TEFLON-tubing 1.6 x 0.35 mm</td><td>SunChrom&amp;reg; GmbH, Friedberg, Germany</td><td>Art. No: 969-195.219</td><td></td></tr><tr><td>Scalpel (Feather&amp;reg; Surgical Blade No. 10)</td><td>pfm medical ag, cologne, Germany</td><td>Art. No: 07310</td><td></td></tr><tr><td>Microscope</td><td>MEIJI Techno&amp;nbsp;</td><td>EMZ-8TR</td><td></td></tr><tr><td>30 G x 11&quot; (0.3 x 25 mm) cannula&amp;nbsp; Sterican&amp;reg; Z</td><td>B.Braun Melsungen AG, Melsungen, Germany</td><td>REF: 9324500</td><td></td></tr><tr><td>25 G x 11/2&quot; (0.5 x 40 mm) cannula 100 Sterican&amp;reg;</td><td>B.Braun Melsungen AG, Melsungen, Germany</td><td>REF: 9186166</td><td></td></tr><tr><td>Fine pen (Stabilo point 88 fine 0.4)</td><td>Schwan-STABILO Schwanh&amp;auml;u&amp;szlig;er GmbH &amp;amp; Co. KG</td><td>Art. No: 88/36</td><td></td></tr><tr><td>Hot glue&amp;nbsp; (Glue sticks ULTRA Power x 11 mm)</td><td>Steinel&amp;reg; GmbH, Herzebrock-Clarholz, Germany</td><td>Art. No: 4007841046910</td><td></td></tr><tr><td>Sandpaper P60 230 x 280</td><td>Robert Bosch GmbH, Gerlingen-Schillerh&amp;ouml;he, Germany</td><td>Catalog Number: 2608605397</td><td></td></tr></tbody></table>

custom-made microdialysis probe design

微透析是在神经科学研究中一个常用的技术。因此，商业探头的需求量很大，监测生理，药理和病理变化的脑脊液。不幸的是，商业探头昂贵的研究小组在公共机构。在这项工作中，一个探针组件进行详细说明，以建立一个可靠的，同心的，定制的微透析探针为低于10美元。的微透析探针由聚砜膜具有分子截止为30 kDa的的。探针的体外回收率与不同分子量（在100-1,600大的范围内）和不同的物理化学性质的物质进行比较。探头产生一个体外回收大约20％的小分子化合物的葡萄糖，乳酸盐，乙酰胆碱和ATP。 体外回收率为神经肽与1,000-1,600达量之间的分子量为2-6％。因此，尽管在较高分子量瓦特神经肽八个降低体外恢复值，化合物的透析中的分子量的较低范围（高达500道尔顿）对体外回收率没有很大的影响。本方法允许利用透析膜与其他截止值和膜材料。因此，这种定制的探针组件具有足够的灵活性的优点透析物质在宽的分子量范围。在这里，我们引入一个微透析探针以2mm的交换长度，这是适用于微透析在小鼠和大鼠的大脑区域。然而，探针的尺寸可以很容易地适应较大交换长度在较大的动物中使用。

同心定制微透析探针是由聚砜膜具有分子截止为30 kDa的的。探针组件示于图3。 它显示出在体外恢复为19.10±1.2％和21.2±1.6％的小能量代谢葡萄糖（180.16道尔顿）和乳酸（112.06道尔顿），分别。用于带正电的乙酰胆碱与181.66大的分子量，它表现出22.6±1.4％ 的体外回收价值。 ATP与551.62达和三个负电荷的磷酸残基的分子量，得到的体外恢复到一个?...

Watch this Scientific Journal Video about Custom-made Microdialysis Probe Design at JoVE.com

Custom-made Microdialysis Probe Design

微透析是在神经科学研究中一个常用的技术。因此商用探头有很大demand.In这项工作探针组件进行了详细建立一个可靠的，同心，定做微透析探针为不到10美元解释。

定制的微透析探针设计

Microdialysis is a commonly used technique in neuroscience research. Therefore commercial probes are in great demand to monitor physiological, ...

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Neuroscience

12.9K 次观看.  Goethe University of Frankfurt. 微透析是在神经科学研究中一个常用的技术。因此商用探头有很大demand.In这项工作探针组件进行了详细建立一个可靠的，同心，定做微透析探针为不到10美元解释。 

视频: Custom-made Microdialysis Probe Design

Hippocampal Insulin Microinjection and In vivo Microdialysis During Spatial Memory Testing

Glucose metabolism is a useful marker for local neural activity, forming the basis of methods such as 2-deoxyglucose and functional magnetic resonance imaging. However, use of such methods in animal models requires anesthesia and hence both alters the brain state and prevents behavioral measures. An alternative method is the use of in vivo microdialysis to take continuous measurement of brain extracellular fluid concentrations of glucose, lactate, and related metabolites in awake, unrestrained animals. This technique is especially useful when combined with tasks designed to rely on specific brain regions and/or acute pharmacological manipulation; for example, hippocampal measurements during a spatial working memory task (spontaneous alternation) show a dip in extracellular glucose and rise in lactate that are suggestive of enhanced glycolysis1-3,4-5, and intrahippocampal insulin administration both improves memory and increases hippocampal glycolysis6. Substances such as insulin can be delivered to the hippocampus via the same microdialysis probe used to measure metabolites. The use of spontaneous alternation as a measure of hippocampal function is designed to avoid any confound from stressful motivators (e.g. footshock), restraint, or rewards (e.g. food), all of which can alter both task performance and metabolism; this task also provides a measure of motor activity that permits control for nonspecific effects of treatment. Combined, these methods permit direct measurement of the neurochemical and metabolic variables regulating behavior.

Hippocampal Insulin Microinjection and In vivo Microdialysis During Spatial Memory Testing

Modulation of hippocampally-dependent spatial working memory by direct intrahippocampal microinjection, accompanied and followed by in vivo microdialysis for metabolites in conscious, behaving animals.

海马胰岛素显微注射和<em在体内</em&gt;微透析过程中空间记忆测试

Glucose metabolism is a useful marker for local neural activity, forming the basis of methods such as 2-deoxyglucose and functional magnetic resonance ...

科研

神经科学

Dissolved Solute Sampling Across an Oxic-Anoxic Soil-Water Interface Using Microdialysis Profilers

Biogeochemical processes shift rapidly in both spatial (millimeter scale) and temporal (hour scale to day scale) dimensions at the oxic-anoxic interface in response to disturbances. Deciphering the rapid biogeochemical changes requires in situ, minimally invasive tools with high spatial and temporal sampling resolution. However, the available passive sampling devices are not very useful in many cases either due to their disposable nature or the complexity and extensive workload for sample preparation.
To address this problem, a microdialysis profiler with 33 individual polyethersulfone nanomembrane tubes (semipermeable, &#60;20 nm pore size) integrated into the one-dimensional skeleton (60 mm) was established to iteratively sample the dissolved compounds in porewater across the soil-water interface at a high resolution of 1.8 mm (outer diameter plus one spacing, i.e., 0.1 mm between probes). The sampling mechanism is based on the principle of concentration gradient diffusion. The automatic loading of degassed water allows minimal disturbance to the chemical species across the oxic-anoxic interface.
This paper describes the procedures of device setup and continuous porewater sampling across the soil-water interface on a daily basis. Concentration-depth profiles were selectively measured before (on Day 6) and after (on Day 7) disturbances induced by irrigation. The results showed that concentration-depth profiles were undergoing rapid changes, especially for redox-sensitive elements (i.e., iron and arsenic). These protocols can help investigate the biogeochemical responses across the soil-water interface under various disturbances caused by physical, chemical, and biological factors. The paper thoroughly discusses the advantages and disadvantages of this method for potential use in the environmental sciences.

A microdialysis profiler is described to sample dissolved porewater solutes across an oxic-anoxic soil-water interface in situ with minimal disturbance. This device is designed to capture rapid changes in concentration-depth profiles induced by disturbances at the soil-water interface and beyond.

使用微透析剖面仪在氧化-缺氧土壤-水界面上进行溶解溶质采样

Biogeochemical processes shift rapidly in both spatial (millimeter scale) and temporal (hour scale to day scale) dimensions at the oxic-anoxic interface ...

环境科学

The Microfluidic Probe: Operation and Use for Localized Surface Processing

Microfluidic devices allow assays to be performed using minute amounts of sample and have recently been used to control the microenvironment of cells. Microfluidics is commonly associated with closed microchannels which limit their use to samples that can be introduced, and cultured in the case of cells, within a confined volume. On the other hand, micropipetting system have been used to locally perfuse cells and surfaces, notably using push-pull setups where one pipette acts as source and the other one as sink, but the confinement of the flow is difficult in three dimensions. Furthermore, pipettes are fragile and difficult to position and hence are used in static configuration only. 
The microfluidic probe (MFP) circumvents the constraints imposed by the construction of closed microfluidic channels and instead of enclosing the sample into the microfluidic system, the microfluidic flow can be directly delivered onto the sample, and scanned across the sample, using the MFP. . The injection and aspiration openings are located within a few tens of micrometers of one another so that a microjet injected into the gap is confined by the hydrodynamic forces of the surrounding liquid and entirely aspirated back into the other opening. The microjet can be flushed across the substrate surface and provides a precise tool for localized deposition/delivery of reagents which can be used over large areas by scanning the probe across the surface. 
In this video we present the microfluidic probe1 (MFP). We explain in detail how to assemble the MFP, mount it atop an inverted microscope, and align it relative to the substrate surface, and finally show how to use it to process a substrate surface immersed in a buffer.

In this video we present the microfluidic probe1 (MFP). We explain in detail how to assemble the MFP, mount it atop an inverted microscope, and align it relative to the substrate surface, and finally show how to use it to process a substrate surface immersed in a buffer solution.

微探针：操作和使用的局部表面处理

Microfluidic devices allow assays to be performed using minute amounts of sample and have recently been used to control the microenvironment of cells. ...

生物学

Integrated Cell Manipulation Platform Coupled with the Single-probe for Mass Spectrometry Analysis of Drugs and Metabolites in Single Suspension Cells

Single cell mass spectrometry (SCMS) enables sensitive detection and accurate analysis of broad ranges of cellular species on the individual-cell level. The single-probe, a microscale sampling and ionization device, can be coupled with a mass spectrometer for on-line, rapid SCMS analysis of cellular constituents under ambient conditions. Previously, the single-probe SCMS technique was primarily used to measure cells immobilized onto a substrate, limiting the types of cells for studies. In the current study, the single-probe SCMS technology has been integrated with a cell manipulation system, typically used for in vitro fertilization. This integrated cell manipulation and analysis platform uses a cell-selection probe to capture identified individual floating cells and transfer the cells to the single-probe tip for microscale lysis, followed by immediate mass spectrometry analysis. This capture and transfer process removes the cells from the surrounding solution prior to analysis, minimizing the introduction of matrix molecules in the mass spectrometry analysis. This integrated setup is capable of SCMS analysis of targeted patient-isolated cells present in body fluids samples (e.g., urine, blood, saliva, etc.), allowing for potential applications of SCMS analysis to human medicine and disease biology.

An integrated cell manipulation platform is developed for use in conjunction with a single-probe mass spectrometry setup for the on-line analysis of individual suspension cells under ambient conditions.

集成细胞操作平台与单悬架细胞中药物和代谢物质谱分析的单探针相结合

Single cell mass spectrometry (SCMS) enables sensitive detection and accurate analysis of broad ranges of cellular species on the individual-cell level. ...

化学

In Vivo Microdialysis Method to Collect Large Extracellular Proteins from Brain Interstitial Fluid with High-molecular Weight Cut-off Probes

In vivo microdialysis is a powerful technique to collect ISF from awake, freely-behaving animals based on a dialysis principle. While microdialysis is an established method that measures relatively small molecules including amino acids or neurotransmitters, it has been recently used to also assess dynamics of larger molecules in ISF using probes with high molecular weight cut off membranes. Upon using such probes, microdialysis has to be run in a push-pull mode to avoid pressure accumulated inside of the probes. This article provides step-by-step protocols including stereotaxic surgery and how to set up microdialysis lines to collect proteins from ISF. During microdialysis, drugs can be administered either systemically or by direct infusion into ISF. Reverse microdialysis is a technique to directly infuse compounds into ISF. Inclusion of drugs in the microdialysis perfusion buffer allows them to diffuse into ISF through the probes while simultaneously collecting ISF. By measuring tau protein as an example, the author shows how its levels are altered upon stimulating neuronal activity by reverse microdialysis of picrotoxin. Advantages and limitations of microdialysis are described along with the extended application by combining other in vivo methods.

In Vivo Microdialysis Method to Collect Large Extracellular Proteins from Brain Interstitial Fluid with High-molecular Weight Cut-off Probes

In vivo microdialysis has enabled collection of molecules present in brain interstitial fluid (ISF) from awake, freely-behaving animals. In order to analyze relatively large molecules in ISF, the current article specifically focuses on the microdialysis protocol using probes with high molecular weight cut off membranes.

体内高分子量截离法从脑间质液中提取大细胞外蛋白的微透析方法

In vivo microdialysis is a powerful technique to collect ISF from awake, freely-behaving animals based on a dialysis principle. While microdialysis is an ...

Microdialysis of Excitatory Amino Acids During EEG Recordings in Freely Moving Rats

Microdialysis is a well-established neuroscience technique that correlates the changes of neurologically active substances diffusing into the brain interstitial space with the behavior and/or with the specific outcome of a pathology (e.g., seizures for epilepsy). When studying epilepsy, the microdialysis technique is often combined with short-term or even long-term video-electroencephalography (EEG) monitoring to assess spontaneous seizure frequency, severity, progression and clustering. The combined microdialysis-EEG is based on the use of several methods and instruments. Here, we performed in vivo microdialysis and continuous video-EEG recording to monitor glutamate and aspartate outflow over time, in different phases of the natural history of epilepsy in a rat model. This combined approach allows the pairing of changes in the neurotransmitter release with specific stages of the disease development and progression. The amino acid concentration in the dialysate was determined by liquid chromatography. Here, we describe the methods and outline the principal precautionary measures one should take during in vivo microdialysis-EEG, with particular attention to the stereotaxic surgery, basal and high potassium stimulation during microdialysis, depth electrode EEG recording and high-performance liquid chromatography analysis of aspartate and glutamate in the dialysate. This approach may be adapted to test a variety of drug or disease induced changes of the physiological concentrations of aspartate and glutamate in the brain. Depending on the availability of an appropriate analytical assay, it may be further used to test different soluble molecules when employing EEG recording at the same time.

Here, we describe a method for in vivo microdialysis to analyze aspartate and glutamate release in the ventral hippocampus of epileptic and non-epileptic rats, in combination with EEG recordings. Extracellular concentrations of aspartate and glutamate may be correlated with the different phases of the disease.

自由运动大鼠脑电图记录过程中兴奋氨基酸的微透析

Microdialysis is a well-established neuroscience technique that correlates the changes of neurologically active substances diffusing into the brain ...

Microinjectrode System for Combined Drug Infusion and Electrophysiology

This microinjectrode system is designed for drug infusion, electrophysiology, and delivery and retrieval of experimental probes, such as microelectrodes and nanosensors, optimized for repeated use in awake, behaving animals. The microinjectrode system can be configured for multiple purposes: (1) simple arrangement of the cannula for placement of an experimental probe that would otherwise be too fragile to penetrate the dura mater, (2) microfluidic infusion of a drug, either independently or coupled to a cannula containing an experimental probe (i.e., microelectrode, nanosensor). In this protocol we explain the step by step construction of the microinjectrode, its coupling to microfluidic components, and the protocol for use of the system in vivo. The microfluidic components of this system allow for delivery of volumes on the nanoliter scale, with minimal penetration damage. Drug infusion can be performed independently or simultaneously with experimental probes such as microelectrodes or nanosensors in an awake, behaving animal. Applications of this system range from measuring the effects of a drug on cortical electrical activity and behavior, to understanding the function of a specific region of cortex in the context of behavioral performance based on probe or nanosensor measurements. To demonstrate some of the capabilities of this system, we present an example of muscimol infusion for reversible inactivation of the frontal eye field (FEF) in rhesus macaque during a working memory task.

We present a microinjectrode system designed for electrophysiology and assisted delivery of experimental probes (i.e., nanosensors, microelectrodes), with optional drug infusion. Widely available microfluidic components are coupled to a cannula containing the probe. A step-by-step protocol for microinjectrode construction is included, with results during muscimol infusion in macaque cortex.

联合药物输液和电生理学的显微注射系统

This microinjectrode system is designed for drug infusion, electrophysiology, and delivery and retrieval of experimental probes, such as microelectrodes ...

Applications of the Single-probe: Mass Spectrometry Imaging and Single Cell Analysis under Ambient Conditions

Mass spectrometry imaging (MSI) and in-situ single cell mass spectrometry (SCMS) analysis under ambient conditions are two emerging fields with great potential for the detailed mass spectrometry (MS) analysis of biomolecules from biological samples. The single-probe, a miniaturized device with integrated sampling and ionization capabilities, is capable of performing both ambient MSI and in-situ SCMS analysis. For ambient MSI, the single-probe uses surface micro-extraction to continually conduct MS analysis of the sample, and this technique allows the creation of MS images with high spatial resolution (8.5 &#181;m) from biological samples such as mouse brain and kidney sections. Ambient MSI has the advantage that little to no sample preparation is needed before the analysis, which reduces the amount of potential artifacts present in data acquisition and allows a more representative analysis of the sample to be acquired. For in-situ SCMS, the single-probe tip can be directly inserted into live eukaryotic cells such as HeLa cells, due to the small sampling tip size (&lt; 10 &#181;m), and this technique is capable of detecting a wide range of metabolites inside individual cells at near real-time. SCMS enables a greater sensitivity and accuracy of chemical information to be acquired at the single cell level, which could improve our understanding of biological processes at a more fundamental level than previously possible. The single-probe device can be potentially coupled with a variety of mass spectrometers for broad ranges of MSI and SCMS studies.

Here, we present protocols to perform both ambient mass spectrometry imaging (MSI) of tissues and in-situ live single cell MS (SCMS) analysis using the single-probe, which is a miniaturized multifunctional device for MS analysis.

单探头的应用:根据环境条件质谱成像和单细胞分析

Mass spectrometry imaging (MSI) and in-situ single cell mass spectrometry (SCMS) analysis under ambient conditions are two emerging fields with great ...

研究人类微血管功能障碍的新机制

Intradermal Microdialysis: An Approach to Investigating Novel Mechanisms of Microvascular Dysfunction in Humans

The cutaneous vasculature is an accessible tissue that can be used to assess microvascular function in humans. Intradermal microdialysis is a minimally invasive technique used to investigate mechanisms of vascular smooth muscle and endothelial function in the cutaneous circulation. This technique allows for the pharmacological dissection of the pathophysiology of microvascular endothelial dysfunction as indexed by decreased nitric oxide-mediated vasodilation, an indicator of cardiovascular disease development risk. In this technique, a microdialysis probe is placed in the dermal layer of the skin, and a local heating unit with a laser Doppler flowmetry probe is placed over the probe to measure the red blood cell flux. The local skin temperature is clamped or stimulated with direct heat application, and pharmacological agents are perfused through the probe to stimulate or inhibit intracellular signaling pathways in order to induce vasodilation or vasoconstriction or to interrogate mechanisms of interest (co-factors, antioxidants, etc.). The cutaneous vascular conductance is quantified, and mechanisms of endothelial dysfunction in disease states can be delineated.

作者聚焦:一种揭示心血管疾病风险人群机制的药物解剖方法 

Intradermal microdialysis is a minimally invasive technique used to investigate microvascular function in health and disease. Both dose-response and local heating protocols can be utilized for this technique to explore mechanisms of vasodilation and vasoconstriction in the cutaneous circulation.

皮内微透析:研究人类微血管功能障碍新机制的方法

The cutaneous vasculature is an accessible tissue that can be used to assess microvascular function in humans. Intradermal microdialysis is a minimally ...

定制的微透析探针设计

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海马胰岛素显微注射和

使用微透析剖面仪在氧化-缺氧土壤-水界面上进行溶解溶质采样

微探针：操作和使用的局部表面处理

集成细胞操作平台与单悬架细胞中药物和代谢物质谱分析的单探针相结合

体内高分子量截离法从脑间质液中提取大细胞外蛋白的微透析方法

自由运动大鼠脑电图记录过程中兴奋氨基酸的微透析

联合药物输液和电生理学的显微注射系统

单探头的应用:根据环境条件质谱成像和单细胞分析

皮内微透析:研究人类微血管功能障碍新机制的方法

皮内微透析:研究人类微血管功能障碍新机制的方法