The authors would like to acknowledge Caitlin Kowalewski for aiding in the editing and formatting of this publication.

1.纹影摄影<ol><li> 试验区的建立 注:测试区域存在直接在反射镜的前面。 <ol><li>夹紧在环形架夹子足够大以支撑反射镜的球面凹面镜（直径150毫米焦距1500毫米）。装上环架夹子用镜子一环站在垂直于地板。目前的研究中使用的3脚环支架，但任何高度可以，只要它是足够高，以便能够居中镜在源的观察窗中。 </li><li>放置环支架和反射镜到质谱仪的源的一侧。使平行于反射镜的面，并在同一高度，作为源。 </li><li>因此，其中心与质谱仪的中心源极区对准的镜子的位置。会发生仪器的一些重叠。 </li></ol></li><li> 截止，相机和光源 <ol><li> 隔断 <ol><li>连接金属板到三脚架的顶端。该板将作为一个平台，以保持两者的刀片和光源。剃刀刀片充当所谓的在纹影摄影的"截止"。 </li><li>附加刀片使用的磁体，使得锋利的边缘是垂直的金属板。 </li><li>放置在与镜像线的三脚架在反射镜3000毫米焦距的两倍。对齐刀片正交从反射镜反射的光的路径。 </li><li>手动调整三脚架的高度，以使刀片的锋利边缘大致与反射镜的中心对齐。 注:微调将在以后发生。 </li></ol></li><li> 相机 <ol><li>挂载数码相机300毫米长焦镜头在一个单独的三脚架。 </li><li>所以镜头（全变焦时）直接4厘米后面，并在同一平定位相机GHT作为刀片。此时不要取下镜头盖。 </li></ol></li><li> 选购的显示器 <ol><li>摄像机的视频输出连接到电脑显示器或电视机方便地查看实时的条纹现象。 注意:这是建议的过程。这个过程可以根据所用相机的类型而改变。 </li></ol></li><li> 针孔光源 <ol><li>钻一个小孔（约0.6毫米直径）为一个盖的中心（在此情况下，一个瓶盖使用了手电筒的直径相同），其可连接/贴在光源。确保该罩具有足够的直径以完全覆盖手电筒透镜。 </li><li>将盖子装在一个200流明使用铝箔胶带LED手电筒。 注:手电筒将得到温暖和建议高温胶带。 </li></ol></li><li> 光源定位 <ol><li>首先用一拉丝氨酸指针对准光源与反射镜，刀片和照相机，以确保光源的适当定位。 </li><li>放在旁边的刀片的金属板的激光指针。 </li><li>所以该光束射到反射镜的中心手动移动激光指针。根据需要进行调整，以确保使束的大约一半被阻塞反射光束垂直相交的剃刀刀片。 </li><li>手动调节后视镜的位置，激光指示器的光束直接在刀片瞄准如果光束对准不是在1.2.5.3实现。 小心！不要直视激光指针或反射光束。 </li><li>确保激光束被集中在透镜上，同时保持摄像机的镜头盖。 </li><li>而一切都对准与覆盖手电筒更换激光指针。确保该手电筒是在相同的方向的激光指针。 </li><li>打开手电筒和，用一张白纸，在截止观察的反射光。确保该光束是在截止的小的聚焦光斑。 </li><li>使任何垂直调整必要阻止大约一半的反射光束与截止的。 </li><li>在相机上取下镜头盖并专注于镜子。 注:建议相机/透镜在手动对焦模式中使用。 </li></ol></li></ol></li></ol> 2.示例测试对象:质谱电离源<ol><li>手动对齐测试区域内的质谱离子源，具有该喷嘴的端部和质谱仪的入口之间10mm的距离。 </li><li>手动打开针阀到环境源允许氮气通过源流动。 </li><li>打开用于控制质谱仪的软件。在这项研究中，所使用的软件是"SQ驱动程序"。点击网络连接然后勒-open-选择适当的调谐文件。 </li><li>一旦手动调谐打开应用所有电压和温度的环境源。每个质谱仪都将拥有自己的这一步软件。对于目前的研究，一旦手动调谐处于打开状态，点击按钮"电源电压为关"和按钮"所有的气体和加热器关闭"来执行此任务。 </li><li>观察排出用数码相机随着温度的增加的观点屏幕上的纹影装置的喷嘴的流动的外观。观察气流（见"结果"部分中描述）现身喷嘴的结束。该气体流可以在摄像机的背面看的，或者它可以直接在液晶监视器上观看。 </li><li>由任一记录来自摄像机的视频，或者服用所述气体流的图片收集图像，一旦所希望的图像被相机实时可视化。 </li><li>传送所收集的图像（多个）到计算机与CAMER存储卡或USB连接，并查看图像与您选择的软件。 </li></ol>从采集到的图像喷雾半角3.测定<ol><li>打开使用图像浏览软件采集到的图像并打印出采集到的图像（S）。 </li><li>绘制限定平行的气流的中心轴流动的用尺子的方向上的印刷图像（多个）上的线。 </li><li>绘制沿着用尺子打印的图像（多个）上的可视化气流的边缘的线。这可能由于微光存在于视频格式被可视化从记录的视频更好;用它来帮助识别打印图像的边缘。标记气流的外边缘以获得用于喷雾半角的范围内。 </li><li>测量中心轴和用量角器在3.2绘制的线之间产生的角度。 </li></ol>

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There are no competing financial interests with this article.

有几方面的考虑，必须之前尝试该协议来解决。除了围绕质谱仪的源和反射镜的空间，足够的开放空间必须可用于容纳所述反射镜的焦点的两倍的距离。此外，反射镜的大小由正在研究源的尺寸最终决定。如果反射镜是太小，源将不被充分显示。要注意的是一些，如果不是全部，源盖必须被除去以实现纹影摄影成像技术是重要的。 实际设置中最关键的步骤是纹影装置的各部分的取向。镜子必须垂直于地面，刀片必须在镜子的焦距长度的两倍正好放置。在这个距离，反射光将被聚焦到一个小点。封锁的剃刀刀片的光量也是我mportant。如果差图像产生的，在第一方面中，调整将与刀片的位置。当刀片不阻挡足够的光到达照相机的，没有对比形成，因此该气体将不可见。如果太多的光被阻挡的图像出现黑暗，从而难以区分从物体所研究的氮气流的更细微的信息。 该技术的一个限制是，必须有在后台的折射率和研究领域的术语一个大的差异。这将取决于所讨论的实验室的温度和湿度。室温氮是通常很难看到作为背景空气是由约78％的氮。这是在所描述的设置，因为氮的温度从其导致的折射率的变化的来源而变化克服。 总体而言，T的显著贡献他的协议是了解参与到源中电离的物理过程的能力。反过来，这将允许用户更好地调整仪器代替盲目地变化的参数，以及作为用于优化的条件下提供的推理。该技术的优点是能够使用所有的信息从物理和化学过程，以获得更好的灵敏度和选择性随环境电离源6。用户可以利用纹影图像，以确定所述源的物理性质，而质谱数据可以用来理解源的化学性质。 未来的应用将是此技术适用于任一其他各种环境电离在市场上可用的源，或一个非商业装置。这也可以应用到利用气体流的任何其它乐器/机器。

质谱法，可用于分子质量鉴定的分析工具，已经成为最强大的分析技术的最新之一。在过去的十年新的环境电离源的整个主机已经成为可用的质谱检测。用于本手稿收集的数据，直接样品分析（DSA）源被利用。虽然这些来源是非常灵活的，需要用于其优化和目的的延长的物理电离过程的更详细的了解。该实验的目的是通过使用一种称为纹影摄影技术的设备上的氮气流的可视化的环境源中，以便更好地理解电离过程。 科学研究往往通过观察，这是困难的，如果研究的对象是透明的，肉眼启动。条纹照相是一种技术，可以让无形通过依靠透明介质1内的折射率变化变得可见。的折射率的不均匀性引起的光允许可视化的歪曲。纹影技术已经在各种专业领域，包括弹道学建模，航天工程，一般气体检测和流量监测，并且有时被常规地用于显现蛋白质条带在凝胶电泳2-5。 大多数环境电离源，以促进电离使用气体流。广泛范围的条件下可以存在为源的选择，然而该实验的参数必须涉及的气体与从周围实验室空气不同的折射率的利用率。这种特殊的研究利用热氮气。应该指出，只有在折射率的差小，从气流和空气纯氮之间观察到在RT 6，主要是因为IR主要是氮组成。此问题是在这种情况下，由于其产生的折射率显著足以改变为要观察的气体的气流中的纯氮气的高温克服。 其他质谱来源，如大气解吸化学电离（DAPCI）7，实时（DART）流动气压余辉（FAPA）8-10，并直接分析11电离源使用了条纹摄影。此协议的目的是讨论如何使用基本条纹照相配置学习环境电离。这种技术，然而，也适用于任何数量的涉及气流不同的分析技术。

<table border="0" cellpadding="0" cellspacing="0" width="606">
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:303px;">Flashlight</td>
			<td style="width:109px;">EAGTAC</td>
			<td style="width:108px;">D25A Ti</td>
			<td style="width:87px;">or equvilent&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Spherical Concave Mirror</td>
			<td>Anchor Optics</td>
			<td>27633</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Rebel EOS T2i</td>
			<td>Canon</td>
			<td>4462B001</td>
			<td>or equvilent&#160;</td>
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		<tr height="20">
			<td height="20" style="height:20px;">300 mm telephoto lens</td>
			<td>Canon</td>
			<td>6473A003</td>
			<td>or equvilent&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Direct Sample Analysis (DSA) Ionization Source</td>
			<td>PerkinElmer</td>
			<td>MZ300560</td>
			<td>or equvilent&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Sq 300 MS with SQ Driver Software</td>
			<td>PerkinElmer</td>
			<td>N2910801</td>
			<td>or equvilent&#160;</td>
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		<tr height="20">
			<td height="20" style="height:20px;">Ring Stand</td>
			<td>Fisher Scientific</td>
			<td>11-474-207</td>
			<td>or equvilent&#160;</td>
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			<td height="20" style="height:20px;">Laser Pointer</td>
			<td>Apollo</td>
			<td>MP1200</td>
			<td>or equvilent&#160;</td>
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		<tr height="20">
			<td height="20" style="height:20px;">razor blade</td>
			<td>Blue Hawk</td>
			<td>34112</td>
			<td>or equvilent&#160;</td>
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		<tr height="20">
			<td height="20" style="height:20px;">small drill bit #73</td>
			<td>CML Supply</td>
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			<td height="20" style="height:20px;">Protractor</td>
			<td>Sterling&#160;</td>
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			<td height="20" style="height:20px;">Hose Clamp</td>
			<td>Trident</td>
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	</tbody>
</table>

visualization of ambient mass spectrometry with the use of schlieren photography

这份手稿概述了如何使用条纹照相可视化质谱环境电离源。为了正确地优化质谱仪，有必要表征和理解源的物理原理。大多数商业环境电离源利用氮气，氦气，或大气空气的射流，以促进分析物的电离。因此，纹影摄影可用于通过利用​​在所述流和环境空气用于实时可视化之间的折射率的差异来可视化气流。基本设置需要一个摄像头，镜子，手电筒，和刀片。如果配置正确，源的实时图像通过观察它的反射观察。这允许洞察在源的作用机制，以及通路到它的优化可以得到阐明。光棚上的不可见的情况。

纹影设置包括质谱电离源的示意图在图1中。当所有的纹影部件正确对齐可以发现，在测试区域内的气体可以被看作是对比明暗区域。 图2示出了这种对比如何能用于观察如何从质谱源变化的氮气喷流的形状喷嘴尺寸减小。 源和气流的完整，未裁剪纹影图像可在图3中找到。该图像示出了测试的取向对象相对于反射镜。 图3中的图像还示出了当适量，约50％，光被所述刀片切断什么应该预计。如果截止要么是过高（ 图4），或过低（ 图5 </STRONG&gt;），图像不佳将导致。 一旦设置完成，可以一边看自己的摄像机的视频屏幕上的效果调整各种质谱仪参数。这个图像，沿着质谱仪的实际信号，允许将迅速达到由于气体流的新的理解最佳条件。 然后这些图像可以被用来计算氮气流的喷射一半的角度。喷雾半角告诉用户的氮气流的整体尺寸。这个角度是由喷嘴，以及该气体的压力和温度的直径进行。 图6是半角测量以恒定的喷嘴的尺寸和在气体压力变化的表示。正如所料，半角度的增加的压力相应地增加，表示该气体的整体尺寸增加流。 图7是与恒压半角的表示，同时改变喷嘴直径。正如所料，半角度与增加喷嘴直径增大。这意味着在氮气喷射出来的源作为喷嘴直径的尺寸的总体比例增加增加。 <img alt="图1" src="/files/ftp_upload/54195/54195fig1.jpg" /> 图1.条纹示意图（重新打印与参考7权限）。与质谱离子源的条纹照相设备的示意图。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54195/54195fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图2" src="/files/ftp_upload/54195/54195fig2.jpg" /> 图2.尼特可视化<html>罗根流（再打印与来自参考7许可）从与（A）4.8毫米，（B）。3.2毫米，（C）的 1.5毫米，（D）的 0.5的不同喷嘴的内径电离源气体流量。纹影照片毫米。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54195/54195fig2large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图3" src="/files/ftp_upload/54195/54195fig3.jpg" /> 图3.环境资源的可视化，与截止正确定位电离源的广角纹影照片。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54195/54195fig3large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <html> g4.jpg"/&gt; 图4.可视化与低截止差。纹影照片与定位过低截止。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54195/54195fig4large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图5" src="/files/ftp_upload/54195/54195fig5.jpg" /> 图5.可怜的可视化与高截止。纹影照片与定位过高截止。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54195/54195fig5large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图6" src="/files/ftp_upload/54195/54195fig6.jpg" /> 图6.半角与瓦斯压力，一个图表，描述在与不同的气体压力恒定的喷嘴尺寸喷雾半角的变化。="htt​​ps://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54195/54195fig6large.jpg"目标="_空白"&gt;点击此处查看该图的放大版本。 <img alt="图7" src="/files/ftp_upload/54195/54195fig7.jpg" /> 图7.半角VS喷嘴尺寸，一个图表，描述在与不同的喷嘴尺寸定压喷射半角的变化。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54195/54195fig7large.jpg" target="_blank">请点击此处查看该图的放大版本。</a>

Watch this Scientific Journal Video about Visualization of Ambient Mass Spectrometry with the Use of Schlieren Photography at JoVE.com

Visualization of Ambient Mass Spectrometry with the Use of Schlieren Photography

This paper presents a protocol for the visualization of gaseous streams of an ambient ionization source using schlieren photography and mass spectrometry.

环境质谱可视化纹影摄影的使用

This manuscript outlines how to visualize mass spectrometry ambient ionization sources using schlieren photography. In order to properly optimize the mass ...

visualization-ambient-mass-spectrometry-with-use-schlieren

Nanyang Technological University

Research

JoVE Journal

Chemistry

8.2K 次观看.  University of Maryland, Baltimore County. This paper presents a protocol for the visualization of gaseous streams of an ambient ionization source using schlieren photography and mass spectrometry.

视频: Visualization of Ambient Mass Spectrometry with the Use of Schlieren Photography

Analyzing Protein Dynamics Using Hydrogen Exchange Mass Spectrometry

All cellular processes depend on the functionality of proteins. Although the functionality of a given protein is the direct consequence of its unique amino acid sequence, it is only realized by the folding of the polypeptide chain into a single defined three-dimensional arrangement or more commonly into an ensemble of interconverting conformations. Investigating the connection between protein conformation and its function is therefore essential for a complete understanding of how proteins are able to fulfill their great variety of tasks. One possibility to study conformational changes a protein undergoes while progressing through its functional cycle is hydrogen-1H/2H-exchange in combination with high-resolution mass spectrometry (HX-MS). HX-MS is a versatile and robust method that adds a new dimension to structural information obtained by e.g. crystallography. It is used to study protein folding and unfolding, binding of small molecule ligands, protein-protein interactions, conformational changes linked to enzyme catalysis, and allostery. In addition, HX-MS is often used when the amount of protein is very limited or crystallization of the protein is not feasible. Here we provide a general protocol for studying protein dynamics with HX-MS and describe as an example how to reveal the interaction interface of two proteins in a complex. &#160;&#160;

Protein conformation and dynamics are key to understanding the relationship between protein structure and function. Hydrogen exchange coupled with high-resolution mass spectrometry is a versatile method to study the conformational dynamics of proteins as well as characterizing protein-ligand and protein-protein interactions, including contact interfaces and allosteric effects.

使用氢交换质谱分析蛋白质动力学

All cellular processes depend on the functionality of proteins. Although the functionality of a given protein is the direct consequence of its unique ...

科研

化学

A Study of the Complexation of Mercury(II) with Dicysteinyl Tetrapeptides by Electrospray Ionization Mass Spectrometry

In this study we evaluated a method for the characterization of complexes, formed in different relative ratios of mercury(II) to dicysteinyl tetrapeptide, by electrospray ionization orbitrap mass spectrometry. This strategy is based on previous successful characterization of mercury-dicysteinyl complexes involving tripeptides by utilizing mass spectrometry among other techniques. Mercury(II) chloride and a dicysteinyl tetrapeptide were incubated in a degassed buffered medium at varying stoichiometric ratios. The complexes formed were subsequently analyzed on an electrospray mass spectrometer consisting of a hybrid linear ion- and orbi- trap mass analyzer. The electrospray ionization mass spectrometry (ESI-MS) spectra were acquired in the positive mode and the observed peaks were then analyzed for distinct mercury isotopic distribution patterns and associated monoisotopic peak. This work demonstrates that an accurate stoichiometry of mercury and peptide in the complexes formed under specified electrospray ionization conditions can be determined by using high resolution ESI MS based on distinct mercury isotopic distribution patterns.

A Study of the Complexation of MercuryII with Dicysteinyl Tetrapeptides by Electrospray Ionization Mass Spectrometry

The characterization of complexes formed in different relative ratios of mercury(II) to dicysteinyl tetrapeptides by electrospray ionization orbitrap mass spectrometry is presented.

汞（II）与Dicysteinyl四肽的络合的电喷雾质​​谱研究

In this study we evaluated a method for the characterization of complexes, formed in different relative ratios of mercury(II) to dicysteinyl tetrapeptide, ...

Fizzy Extraction of Volatile Organic Compounds Combined with Atmospheric Pressure Chemical Ionization Quadrupole Mass Spectrometry

Chemical analysis of volatile and semivolatile compounds dissolved in liquid samples can be challenging. The dissolved components need to be brought to the gas phase, and efficiently transferred to a detection system. Fizzy extraction takes advantage of the effervescence phenomenon. First, a carrier gas (here, carbon dioxide) is dissolved in the sample by applying overpressure and stirring the sample. Second, the sample chamber is decompressed abruptly. Decompression leads to the formation of numerous carrier gas bubbles in the sample liquid. These bubbles assist the release of the dissolved analyte species from the liquid to the gas phase. The released analytes are immediately transferred to the atmospheric pressure chemical ionization interface of a triple quadrupole mass spectrometer. The ionizable analyte species give rise to mass spectrometric signals in the time domain. Because the release of the analyte species occurs over short periods of time (a few seconds), the temporal signals have high amplitudes and high signal-to-noise ratios. The amplitudes and areas of the temporal peaks can then be correlated with concentrations of the analytes in the liquid samples subjected to fizzy extraction, which enables quantitative analysis. The advantages of fizzy extraction include: simplicity, speed, and limited use of chemicals (solvents).

Fizzy extraction is a new laboratory technique for analysis of volatile and semivolatile compounds. A carrier gas is dissolved in the liquid sample by applying overpressure and stirring the sample. The sample chamber is then decompressed. The analyte species are liberated to the gas phase due to effervescence.

挥发性有机化合物与大气压化学电离四极杆质谱联用

Chemical analysis of volatile and semivolatile compounds dissolved in liquid samples can be challenging. The dissolved components need to be brought to ...

Synthesis and Mass Spectrometry Analysis of Oligo-peptoids

Peptoids are sequence-controlled peptide-mimicking oligomers consisting of N-alkylated glycine units. Among many potential applications, peptoids have been thought of as a type of molecular information storage. Mass spectrometry analysis has been considered the method of choice for sequencing peptoids. Peptoids can be synthesized via solid phase chemistry using a repeating two-step reaction cycle. Here we present a method to manually synthesize oligo-peptoids and to analyze the sequence of the peptoids using tandem mass spectrometry (MS/MS) techniques. The sample peptoid is a nonamer consisting of alternating N-(2-methyloxyethyl)glycine (Nme) and N-(2-phenylethyl)glycine (Npe), as well as an N-(2-aminoethyl)glycine (Nae) at the N-terminus. The sequence formula of the peptoid is Ac-Nae-(Npe-Nme)4-NH2, where Ac is the acetyl group. The synthesis takes place in a commercially available solid-phase reaction vessel. The rink amide resin is used as the solid support to yield the peptoid with an amide group at the C-terminus. The resulting peptoid product is subjected to sequence analysis using a triple-quadrupole mass spectrometer coupled to an electrospray ionization source. The MS/MS measurement produces a spectrum of fragment ions resulting from the dissociation of charged peptoid. The fragment ions are sorted out based on the values of their mass-to-charge ratio (m/z). The m/z values of the fragment ions are compared against the nominal masses of theoretically predicted fragment ions, according to the scheme of peptoid fragmentation. The analysis generates a fragmentation pattern of the charged peptoid. The fragmentation pattern is correlated to the monomer sequence of the neutral peptoid. In this regard, MS analysis reads out the sequence information of the peptoids.

A protocol is described for the manual synthesis of oligo-peptoids followed by sequence analysis by mass spectrometry.

寡 peptoids 的合成及质谱分析

Peptoids are sequence-controlled peptide-mimicking oligomers consisting of N-alkylated glycine units. Among many potential applications, peptoids have ...

Imaging Corrosion at the Metal-Paint Interface Using Time-of-Flight Secondary Ion Mass Spectrometry

Corrosion developed at the paint and aluminum (Al) metal-paint interface of an aluminum alloy is analyzed using time-of-flight secondary ion mass spectrometry (ToF-SIMS), illustrating that SIMS is a suitable technique to study the chemical distribution at a metal-paint interface. The painted Al alloy coupons are immersed in a salt solution or exposed to air only. SIMS provides chemical mapping and 2D molecular imaging of the interface, allowing direct visualization of the morphology of the corrosion products formed at the metal-paint interface and mapping of the chemical after corrosion occurs. The experimental procedure of this method is presented to provide technical details to facilitate similar research and highlight pitfalls that may be encountered during such experiments.

Time-of-flight secondary ion mass spectrometry is applied to demonstrate the chemical mapping and corrosion morphology at the metal-paint interface of an aluminum alloy after being exposed to a salt solution compared with a specimen exposed to air.

利用飞行时二次离子质谱法成像金属颜料界面的腐蚀

Corrosion developed at the paint and aluminum (Al) metal-paint interface of an aluminum alloy is analyzed using time-of-flight secondary ion mass ...

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. ...

Capillary Electrophoresis Mass Spectrometry Approaches for Characterization of the Protein and Metabolite Corona Acquired by Nanomaterials

The adsorption of biomolecules from surrounding biological matrices to the surface of nanomaterials (NMs) to form the corona has been of interest for the past decade. Interest in the bio-nano interface arises from the fact that the biomolecular corona confers a biological identity to NMs and thus causes the body to identify them as &#8220;self&#8221;. For example, previous studies have demonstrated that the proteins in the corona are capable of interacting with membrane receptors to influence cellular uptake and established that the corona is responsible for cellular trafficking of NMs and their eventual toxicity. To date, most research has focused upon the protein corona and overlooked the possible impacts of the metabolites included in the corona or synergistic effects between components in the complete biomolecular corona. As such, this work demonstrates methodologies to characterize both the protein and metabolite components of the biomolecular corona using bottom-up proteomics and metabolomics approaches in parallel. This includes an on-particle digest of the protein corona with a surfactant used to increase protein recovery, and a passive characterization of the metabolite corona by analyzing metabolite matrices before and after NM exposures. This work introduces capillary electrophoresis &#8211; mass spectrometry (CESI-MS) as a new technique for NM corona characterization. The protocols outlined here demonstrate how CESI-MS can be used for the reliable characterization of both the protein and metabolite corona acquired by NMs. The move to CESI-MS greatly decreases the volume of sample required (compared to traditional liquid chromatography &#8211; mass spectrometry (LC-MS) approaches) with multiple injections possible from as little as 5 &#181;L of sample, making it ideal for volume limited samples. Furthermore, the environmental consequences of analysis are reduced with respect to LC-MS due to the low flow rates (&#60;20 nL/min) in CESI-MS, and the use of aqueous electrolytes which eliminates the need for organic solvents.

Here we present a protocol to characterize the complete biomolecular corona, proteins, and metabolites, acquired by nanomaterials from biofluids using a capillary electrophoresis &#8211; mass spectrometry approach.

纳米材料获得的蛋白质和代谢物科罗纳的毛细电电量谱方法

The adsorption of biomolecules from surrounding biological matrices to the surface of nanomaterials (NMs) to form the corona has been of interest for the ...

Gas Chromatography-Mass Spectrometry Paired with Total Vaporization Solid-Phase Microextraction as a Forensic Tool

Gas Chromatography &#8211; Mass Spectrometry (GC-MS) is a frequently used technique for the analysis of numerous analytes of forensic interest, including controlled substances, ignitable liquids, and explosives. GC-MS can be coupled with Solid-Phase Microextraction (SPME), in which a fiber with a sorptive coating is placed into the headspace above a sample or immersed in a liquid sample. Analytes are sorbed onto the fiber which is then placed inside the heated GC inlet for desorption. Total Vaporization Solid-Phase Microextraction (TV-SPME) utilizes the same technique as immersion SPME but immerses the fiber into a completely vaporized sample extract. This complete vaporization results in a partition between only the vapor phase and the SPME fiber without interference from a liquid phase or any insoluble materials. Depending upon the boiling point of the solvent used, TV-SPME allows for large sample volumes (e.g., up to hundreds of microliters). On-fiber derivatization may also be performed using TV-SPME. TV-SPME has been used to analyze drugs and their metabolites in hair, urine, and saliva. This simple technique has also been applied to street drugs, lipids, fuel samples, post-blast explosive residues, and pollutants in water. This paper highlights the use of TV-SPME to identify illegal adulterants in very small samples (microliter quantities) of alcoholic beverages. Both gamma-hydroxybutyrate (GHB) and gamma-butyrolactone (GBL) were identified at levels that would be found in spiked drinks. Derivatization by a trimethylsilyl agent allowed for conversion of the aqueous matrix and GHB into their TMS derivatives. Overall, TV-SPME is quick, easy, and requires no sample preparation aside from placing the sample into a headspace vial.

Total Vaporization Solid Phase Microextraction (TV-SPME) completely vaporizes a liquid sample whilst analytes are sorbed onto a SPME fiber. This allows for partitioning of the analyte between only the solvent vapor and the SPME fiber coating.

气相色谱-质量光谱学与总蒸发固体相微切成一对，作为取证工具

Gas Chromatography &#8211; Mass Spectrometry (GC-MS) is a frequently used technique for the analysis of numerous analytes of forensic interest, including ...

Covalent Labeling with Diethylpyrocarbonate for Studying Protein Higher-Order Structure by Mass Spectrometry

Characterizing a protein&#39;s higher-order structure is essential for understanding its function. Mass spectrometry (MS) has emerged as a powerful tool for this purpose, especially for protein systems that are difficult to study by traditional methods. To study a protein&#39;s structure by MS, specific chemical reactions are performed in solution that encode a protein&#39;s structural information into its mass. One particularly effective approach is to use reagents that covalently modify solvent accessible amino acid side chains. These reactions lead to mass increases that can be localized with residue-level resolution when combined with proteolytic digestion and tandem mass spectrometry. Here, we describe the protocols associated with use of diethylpyrocarbonate (DEPC) as a covalent labeling reagent together with MS detection. DEPC is a highly electrophilic molecule capable of labeling up to 30% of the residues in the average protein, thereby providing excellent structural resolution. DEPC has been successfully used together with MS to obtain structural information for small single-domain proteins, such as &#946;2-microglobulin, to large multi-domain proteins, such as monoclonal antibodies.

The experimental procedures for performing diethylpyrocarbonate-based covalent labeling with mass spectrometric detection are described. Diethylpyrocarbonate is simply mixed with the protein or protein complex of interest, leading to the modification of solvent accessible amino acid residues. The modified residues can be identified after proteolytic digestion and liquid chromatography/mass spectrometry analysis.

通过质谱学研究蛋白质高阶结构，与二乙基碳酸酯共价标签

Characterizing a protein&#39;s higher-order structure is essential for understanding its function. Mass spectrometry (MS) has emerged as a powerful tool ...

Capillary Electrophoresis-based Hydrogen/Deuterium Exchange for Conformational Characterization of Proteins with Top-down Mass Spectrometry

Resolving conformational heterogeneity of multiple protein states that coexist in solution remains one of the main obstacles in the characterization of protein therapeutics and the determination of the conformational transition pathways critical for biological functions, ranging from molecular recognition to enzymatic catalysis. Hydrogen/deuterium exchange (HDX) reaction coupled with top-down mass spectrometric (MS) analysis provides a means to characterize protein higher-order structures and dynamics in a conformer-specific manner. The conformational resolving power of this technique is highly dependent on the efficiencies of separating protein states at the intact protein level and minimizing the residual non-deuterated protic content during the HDX reactions. 
Here we describe a capillary electrophoresis (CE)-based variant of the HDX MS approach that aims to improve the conformational resolution. In this approach, proteins undergo HDX reactions while migrating through a deuterated background electrolyte solution (BGE) during the capillary electrophoretic separation. Different protein states or proteoforms that coexist in solution can be efficiently separated based on their differing charge-to-size ratios. The difference in electrophoretic mobility between proteins and protic solvent molecules minimizes the residual non-deuterated solvent, resulting in a nearly complete deuterating environment during the HDX process. The flow-through microvial CE-MS interface allows efficient electrospray ionization of the eluted protein species following a rapid mixing with the quenching and denaturing modifier solution at the outlet of the sprayer. The online top-down MS analysis measures the global deuteration level of the eluted intact protein species, and subsequently, the deuteration of their gas-phase fragments. This paper demonstrates this approach in differential HDX for systems, including the natural protein variants coexisting in milk.

Presented here is a protocol for a capillary electrophoresis-based hydrogen/deuterium exchange (HDX) approach coupled with top-down mass spectrometry. This approach characterizes the difference in higher-order structures between different protein species, including proteins in different states and different proteoforms, by conducting concurrent differential HDX and electrophoretic separation.

基于毛细管电泳的氢/氘交换，用于自上而下的质谱法对蛋白质进行构象表征

Resolving conformational heterogeneity of multiple protein states that coexist in solution remains one of the main obstacles in the characterization of ...