Name: large-scale top-down proteomics using capillary zone electrophoresis tandem mass spectrometry
Uploaded: 2018-10-24T15:00:00
Description: Watch this Scientific Journal Video about Large-scale Top-down Proteomics Using Capillary Zone Electrophoresis Tandem Mass Spectrometry at JoVE.com

作者感谢 Heedeok 在密歇根州立大学化学系的小组, 为实验提供大肠杆菌细胞. 作者感谢全国普通医学研究院、国立卫生研究院 (NIH) 通过授予 R01GM118470 (向 x. 刘) 和授予 R01GM125991 (对 l. 孙和 x. 刘) 的支持。

1. 分离毛细管内壁上的 LPA 涂层的制备
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毛细管的预处理
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<li>冲洗熔融石英毛细管 (长度为120厘米, 内径为50µm [内径], 360 µm 外径 [外径]), 连续500µL 1 m 氢氧化钠, 去离子水, 1 m 盐酸, 去离子水, 和 LC MS 级甲醇使用注射器泵。</li>
		<li>用氮气 (10 psi, ≥ 12 h) 干燥毛细管, 用注射器泵在甲醇中填充 50% (v/v) 3-(氧硅) 丙基丙烯酸酯。用硅胶密封毛细管两端, 并在室温下孵育至少24小时。...

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在这里, 我们提供了一个详细的协议, 使用 CZE 的高分辨率表征 proteoforms 在简单的蛋白质样本和大规模鉴定 proteoforms 在复杂的蛋白质组样品。图 1显示了 CZE-ESI/ms 系统的示意图。协议中有四关键步骤。首先, 在分离毛细管的内壁上制备高质量的 LPA 涂层非常重要。用 LPA 包覆分离毛细管可以减少毛细管中的 EOF, 拓宽 CZE 的分离窗, 减少其内壁19的蛋白质吸附。<em...

自上而下的蛋白质组学 (TDP) 的目的是大规模表征 proteoforms 内的蛋白质。TDP 依赖于在电喷雾电离-串联质谱 (ESI ms/ms) 分析之前, 由于蛋白质组1、2 的高复杂性和大浓度动态范围而对完整蛋白进行有效液相分离 ,3,4,5。毛细管区电泳 (CZE) 是一种强大的技术, 以分离生物分子的大小-电荷比6。CZE 是相对简单的, 只需要一个开放管状熔融石英毛细管, 背景电解质 () 和电源。完整的蛋白质样品可以通过压力或电压加载到毛细管中, 并通过浸泡毛细管两端并施加高电压来启动分离。CZE 可接近超高分离效率 (> 100万个理论板), 用于分离生物分子7。CZE 的灵敏度比广泛使用的反相液相色谱 (RPLC)-ms 对完整蛋白8的分析有了极大的提高。虽然 CZE 对大规模自上而下蛋白质组学有很大的潜力, 但它在蛋白质组学中的广泛应用受到了一些问题的阻碍, 包括低采样量和窄分离窗口。CZE 中典型的样品加载量约为总毛细管体积的 1%, 通常对应于小于 100 nL9、10、11。由于强电渗流 (EOF)9、10, CZE 的分离窗口通常小于30分钟。这些问题限制了 CZE (ms), 用于识别来自复杂蛋白质组的大量 proteoforms 和低丰 proteoforms。
通过在线样品浓度法 (例如固相微萃取 [固相微萃取]12、13、场增强样品堆积 (CZE)9 , 提高了样品加载量。,11,14、动态 pH 结15、16、17、18)。和动态 ph 结比固相微萃取简单, 只需要在电导率和 pH 值之间有显著差异的样品缓冲液。在取样区域和毛细管中的的小区之间的边界上, 该样品缓冲液的电导率要低得多。动态 pH 结使用一个基本的样品塞 (例如, 50 毫米碳酸氢钠, ph 8) 和酸盐 (例如, 5% [v-/v] 醋酸, pH 2.4) 在样品插头的两侧。在毛细管的注射端施加高正压时, 会发生基本样品塞的滴定, 在进行 CZE 分离之前将分析物集中在紧密的插头上。最近, Sun 集团系统地比较了在完整的蛋白质在线堆叠的承认和动态 ph 结, 表明动态 ph 结可以产生更好的性能比坦白的在线浓度完整的蛋白质时样品注射量为总毛细管体积19的25%。
中立涂层分离毛细血管 (例如, 线性聚丙烯酰胺 [LPA]) 已被用于减少毛细管中的 EOF, 减慢 CZE 分离和扩大分离窗口20,21。最近, Dovichi 集团开发了一个简单的程序, 在毛细血管内壁上制备稳定的 LPA 涂层, 利用过硫酸铵 (ap) 作为引发剂和温度 (50 摄氏度) 自由基生产和聚合22.最近, Sun 集团采用了 LPA 涂层分离毛细管和动态 pH 结方法 CZE 分离完整的蛋白质, 达到微升的样品加载量和90分钟的分离窗口19。这个 CZE 系统打开门使用 CZE ms/ms 的大规模自上而下的蛋白质组学。
CZE 需要一个高度健壮和敏感的接口, 以耦合 CZE 到 MS。三 ce ms 接口在 ce 的历史中得到了很好的开发和商业化, 它们是共轴鞘流接口23, sheathless 接口使用多孔尖端作为 ESI 发射器24, 以及电动力学泵送护套流量接口25,26。电动力学泵鞘-流界面 CZE-ms 已达到低 zeptomole 肽检测限制9, 超过1万肽鉴定 (IDs) 从 HeLa 细胞蛋白质组在单一运行14, 快速表征完整的蛋白质11, 和高度稳定和可重现的生物分子分析26。最近, 采用 LPA 包覆分离毛细管、动态 pH 结法和电动力学泵鞘流界面, 用于大肠埃希氏杆菌 (大肠杆菌) 蛋白质组 19 的大尺度上向下蛋白分析 ,27。CZE 平台通过与尺寸排除色谱 (SEC)-RPLC 分馏27的耦合, 在单个运行19和近 6000 proteoform id 中接近 500 proteoform id。结果清楚地显示了 CZE 对大范围自上而下蛋白质组学的能力。
本文介绍了一种用于大规模自上而下蛋白质组学的 CZE/ms 的详细过程。CZE 系统采用了 LPA 涂层毛细管, 以减少毛细管中的 EOF, 动态 pH 结方法的蛋白质在线浓度, 电动力学泵鞘流接口 CZE 耦合到 MS, orbitrap 质量用于采集蛋白质的 ms 和 ms/ms 光谱的光谱仪, 以及通过数据库搜索 Proteoform ID 的 TopPIC (自上而下的质谱 Proteoform 识别和表征) 软件。

<table border="0" cellpadding="0" cellspacing="0" style="width:905px;" width="903">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:287px;">Fused silica capillary</td>
			<td style="width:117px;">Polymicro Technologies</td>
			<td style="width:147px;">1068150017</td>
			<td style="width:355px;">50 &#181;m i.d. 360 &#181;m o.d.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:287px;">Sodium hydroxide pellets</td>
			<td style="width:117px;">Macron Fine Chemicals</td>
			<td style="width:147px;">7708-10</td>
			<td>Corrosive</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">LC-MS grade water</td>
			<td style="width:117px;">Fisher Scientific</td>
			<td>W6-1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Hydrochloric acid</td>
			<td style="width:117px;">Fisher Scientific</td>
			<td style="width:147px;">SA48-1</td>
			<td>Corrosive</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Methanol</td>
			<td style="width:117px;">Fisher Scientific</td>
			<td style="width:147px;">A456-4</td>
			<td>Toxic, Health Hazard</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">3-(Trimethoxysilyl)propyl methacrylate</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">M6514</td>
			<td>Moisture and heat sensitive</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Hydrofluoric acid</td>
			<td style="width:117px;">Acros Organics</td>
			<td style="width:147px;">423805000</td>
			<td>Highy toxic</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Acrylamide</td>
			<td style="width:117px;">Acros Organics</td>
			<td style="width:147px;">164855000</td>
			<td>Toxic, health hazard</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Ammonium persulfate</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">A3678</td>
			<td>Health hazard, Oxidizer</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">lysozyme</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">L6876</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Cytochrome C</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">C7752</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Myoglobin</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">M1882</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">&#223;-casein</td>
			<td style="width:117px;">Sgma-Aldrich</td>
			<td style="width:147px;">C6905</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Carbonic anhydrase</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">C3934</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Bovine serum albumin</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">A2153</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Urea</td>
			<td style="width:117px;">Alfa Aesar</td>
			<td style="width:147px;">36428-36</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">DL-Dithiothreitol</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">D0632</td>
			<td>Health Hazard</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Iodoacetamide</td>
			<td style="width:117px;">Fisher Scientific</td>
			<td style="width:147px;">AC122270250</td>
			<td>Health Hazard</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Formic Acid</td>
			<td style="width:117px;">Fisher Scientific</td>
			<td style="width:147px;">A117-50</td>
			<td>Corrosive, Health Hazard</td>
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		<tr height="42">
			<td height="42" style="height:42px;width:287px;">C4 trap column</td>
			<td style="width:117px;">Sepax Technologies</td>
			<td style="width:147px;">110043-4001C</td>
			<td>3 &#181;m particles, 300 &#197; pores, 4.0 mm i.d. 10 mm long</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Acetonitrile</td>
			<td style="width:117px;">Fisher Scientific</td>
			<td style="width:147px;">A998SK-4</td>
			<td>Toxic, Oxidizer</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Ammonium bicarbonate</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">1066-33-7</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:287px;">Nalgene rapid-flow filters</td>
			<td style="width:117px;">Thermo Scientific</td>
			<td style="width:147px;">126-0020</td>
			<td>0.2 &#181;m CN membrane, and 50 mm diameter</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">E. coli cells</td>
			<td style="width:117px;"></td>
			<td style="width:147px;"></td>
			<td>K-12 MG1655</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Dulbecco&#39;s phosphate-buffered saline</td>
			<td style="width:117px;">Sigma-Aldrich</td>
			<td style="width:147px;">D8537</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:287px;">BCA assay</td>
			<td style="width:117px;">Thermo Scientific</td>
			<td style="width:147px;">23250</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Acetone</td>
			<td style="width:117px;">Fisher Scientific</td>
			<td style="width:147px;">A11-1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">HPLC system for protein desalting</td>
			<td style="width:117px;">Agilient</td>
			<td style="width:147px;">1260 Infinity II</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Acetic Acid</td>
			<td style="width:117px;">Fisher Scientific</td>
			<td style="width:147px;">A38-212</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">CE autosampler</td>
			<td style="width:117px;">CMP Scientific</td>
			<td style="width:147px;">ECE-001</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:287px;">Electro-kinetically pumped sheath flow interface</td>
			<td style="width:117px;">CMP Scientific</td>
			<td style="width:147px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:287px;">Q Exactive HF Hybrid Quadrupole-Orbitrap Mass Spectrometer</td>
			<td style="width:117px;">Thermo Fisher Scientific</td>
			<td style="width:147px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:287px;">Sutter flaming/brown micropipette puller</td>
			<td style="width:117px;">Sutter Instruments</td>
			<td style="width:147px;">P-1000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:287px;">Ultrasonic cell disruptor for cell lysis</td>
			<td style="width:117px;">Branson</td>
			<td align="right" style="width:147px;">101063196</td>
			<td>Model S-250A</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:287px;">Vaccum concentrator</td>
			<td style="width:117px;">Thermo Fisher Scientific</td>
			<td style="width:147px;">SPD131DDA-115</td>
			<td></td>
		</tr>
	</tbody>
</table>

large-scale top-down proteomics using capillary zone electrophoresis tandem mass spectrometry

毛细管区电泳-电喷雾电离-串联质谱 (CZE) 已被公认为自上而下蛋白质组学的有用工具, 旨在表征复杂蛋白质组中的 proteoforms。然而, CZE 在大范围自上而下蛋白质组学中的应用由于 CZE 的样品加载能力低, 分离窗口狭窄而受阻。在这里, 使用 CZE 的协议描述了一个微升的样品加载量和一个90分钟的分离窗口, 用于大规模自上而下的蛋白质组学。CZE 平台基于线性聚丙烯酰胺 (LPA) 涂层分离毛细管, 具有极低的电渗流, 一种基于动态 pH 结的在线样品浓缩方法, 具有高效率的蛋白质堆积,电动力学泵鞘流 CE 接口具有极高的灵敏度, 以及具有高质量分辨率和扫描速度的离子阱质谱仪。该平台可用于简单完整蛋白样品的高分辨率表征和各种复杂蛋白质组中 proteoforms 的大尺度表征。例如, 我们演示了一种高效分离标准蛋白混合物和使用该平台对许多杂质进行高度灵敏检测的方法。作为另一个例子, 这个平台可以产生超过 500 proteoform 和190蛋白质鉴定从大肠杆菌蛋白质组在一个单一的 CZE ms/ms 运行。

图 1显示了在实验中使用的基于动态 pH 结的 CZE 系统的示意图。在基本缓冲液中, 样品的长插头被注射到带有酸性物质的 LPA 涂层分离毛细管中。在应用高电压 I 和 II 后, 样品区中的分析物将通过动态 pH 结法集中。为了评估 CZE 系统的性能, 通常分析标准蛋白混合物 (细胞色素 c、溶菌酶、β酪蛋白、肌红蛋白、钙和 BSA)。标准蛋白混合物的代表电...

Watch this Scientific Journal Video about Large-scale Top-down Proteomics Using Capillary Zone Electrophoresis Tandem Mass Spectrometry at JoVE.com

Large-scale Top-down Proteomics Using Capillary Zone Electrophoresis Tandem Mass Spectrometry

利用毛细管区电泳串联质谱技术进行大尺度自上而下蛋白质组学

通过毛细管区电泳-电喷雾-串联质谱 (CZE-proteoforms), 描述了蛋白质样品中的分离、鉴定和表征的详细协议。该协议可用于简单蛋白样品中 proteoforms 的高分辨率表征和复杂蛋白质组样本中 proteoforms 的大尺度鉴定。

Capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry (CZE-ESI-MS/MS) has been recognized as a useful tool for top-down ...

该方法有助于改善蛋白质异构形式和细胞转化后蛋白质的大规模和高分辨率表征。该技术的主要优点是毛细管区电泳质谱仪提供高效分离和高度灵敏的检测完整蛋白质。要预处理毛细管，用氮气在 10 psi 下干燥至少 12 小时。然后，使用注射器泵将毛细管中 50% 体积的 3 丙基甲基丙烯酸酯填充到甲醇中。用硅橡胶密封毛细管的两端。在室温下孵育至少24小时后，继续在分离毛细管内壁上涂上线性聚丙烯酰胺涂层，如文本协议所述。使用柔和的火焰燃烧毛细管的一部分，以去除离毛细管一端约四厘米的聚酰胺外涂层。通过擦拭，轻轻清洁毛细管的烧伤部分，以完全去除聚酰胺涂层。在 200 微升管的末端钻一个小孔，其尺寸与毛细管外径相同，一旦毛细管穿过此孔，就足以将毛细管保持到位。螺纹毛细管的末端，靠近烧伤部分通过孔，直到烧伤部分在管中。然后，在 200 微升管中加入大约 150 微升 HF，使 HF 溶液大约是毛细管上烧伤部分的一半。在室温下孵育HF溶液中的毛细管90至100分钟。准备标准蛋白质混合物和大肠杆菌样品，如文本协议中所述。要设置毛细管区电泳或 CZE 系统，请将样品、背景电解质和分离毛细管加载到 CZE 自动采样器中。在 CZE 自动采样器的手动控制页中选择样品负载。将背景电解质小瓶加载到自动采样器的缓冲盘中，注意其在缓冲盘中的位置。然后，将样品小瓶装入自动取样器的样品托盘中，注意其在样品托盘中的位置。使用手动控制将自动采样器置于对齐位置。现在，使用毛细管的非蚀刻端将分离毛细管加载到 CZE 自动采样器中。如果样品体积小于 50 微升，请调整毛细管喷射端的高度。使用手动控制将自动采样器切换到待机状态。在系统中键入样品位置，然后将毛细管移动到样品中。将毛细管的喷射端进一步向下推，以到达样品瓶的底部。继续使用手动控制，使用背景电解质冲洗毛细管 20 分钟，压力为 20 psi。现在，在质谱仪的前面安装一个商用电镀泵护套流接口。在 LCMS 级水中，用含有 10% 体积甲醇和 0.2% 体积甲酸的缓冲液填充护套缓冲液库。通过使用注射器手动施加压力，用护套缓冲液冲洗界面中的 T。通过套管螺纹电喷雾发射器的一毫米外径端，并通过接头将发射器与 T 的一个端口连接。只需用注射器手动再次冲洗 T，即可用护套缓冲液填充发射器。在接口附带的摄像机的帮助下，将发射器的孔与质谱仪入口之间的距离调整到大约两毫米。在护套缓冲器小瓶上施加 2 到 2.2 千伏电压进行电喷。然后，调整喷雾电压以达到稳定的电喷。在此过程期间，在毛细管的喷射端施加低压，以确保毛细管中没有气泡。关闭喷雾电压，通过 T 轻轻将分离毛细管蚀刻端螺纹入发射器，直到无法进一步推。在停止毛细管喷射端的低压后，用护套缓冲器冲洗发射器一点。再次，施加 2 到 2.2 千伏的喷雾电压来测试喷雾。在主仪器屏幕上的文件下拉菜单下选择新方法。在那里，选择入口作为起始样品小瓶，结束样品小瓶，托盘作为样品，压力为5 psi，千伏为零，持续时间为95秒的样品注射。然后，将进气作为进气瓶，托盘设置为缓冲液，压力为零 psi，千伏为30，持续时间为4，200秒，用于分离标准蛋白混合物样品。对于大肠杆菌蛋白蛋白组样品的分离，将进气作为入口小瓶，托盘设置为缓冲器，压力为零 psi，千伏为 20，持续时间为 6，600 秒。最后，选择入口作为入口小瓶，托盘作为缓冲器，压力为10 psi，千伏为30，持续时间为600秒毛细管冲洗。现在，使用四离子陷阱掩膜光谱仪设置MS和MS/MS参数，用于完整的蛋白质分析。要调整调谐文件设置，请打开完整的蛋白质模式并使用 0.2 的陷印压力。将离子传输毛细管温度设置为 320 摄氏度，将 S-镜头 RF 电平设置为 55。要执行 CZE MS/MS 实验，请首先选择运行序列，在质谱仪计算机上启动数据采集方法。然后，在毛细管电泳自动采样器计算机上启动毛细管电泳序列。在 TopPIC 套件和 TopPIC 图形用户界面中使用 TopPIC 执行数据库搜索。单击数据库文件并选择适当的数据库进行搜索。单击频谱文件并选择生成的 ms2。作为输入生成的 msalign 文件。选择 MS1 要素文件并选择生成的要素文件。将半胱氨酸碳基C57设置为固定修饰，由于碘多乙酰胺治疗。选择诱饵数据库功能和截止设置下，在频谱级别旁边的下拉菜单中选择错误发现率。在频谱级别将错误发现率设置为 0.01。保持生成函数未选中，将误差容差设置为百万分之 15。将错误发现率设置为 0.05 的蛋白质形式级别。在高级参数下，为下拉菜单中的最大质量班次数选择两个。将未知修改的最大质量移位设置为 500 道尔顿。将所有其他参数保留为默认值，然后单击图形用户界面右下角的"开始"按钮。显示了标准蛋白质混合物的代表性电图。标准蛋白质混合物通常至少重复运行，以评估系统的分离效率和可重复性。分离效率可以用某些蛋白质的理论板数来评价。可重复性可以通过蛋白质强度和迁移时间的相对标准偏差来评价。CZE MS/MS 平台可用于在各种复杂蛋白质组中大规模表征蛋白体，在一次运行中以高置信度识别出 500 多个蛋白质和 190 种蛋白质。在这里，电图的放大视图可用于评估系统的分离窗口。极低的E值和光谱FDR表明蛋白质组符号ID的高置信度。 大量匹配的片段离子进一步表明ID的高置信度。稳定的同位素标记或无标签方法可以纳入CZE MS程序，以确定细胞在各种条件下的蛋白质形式丰度如何变化。该技术开发后，为自上而下的蛋白质组学领域的研究人员探索蛋白质组在调节细胞中各种生物过程方面的作用铺平了道路。不要忘记，使用氢氟酸可能极其危险，执行此程序时应始终采取预防措施，如适当的个人防护设备和应急程序。

Research

JoVE Journal

Chemistry

Large Scale Non-targeted Metabolomic Profiling of Serum by Ultra Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS)

Large Scale Non-targeted Metabolomic Profiling of Serum by Ultra Performance Liquid Chromatography-Mass Spectrometry UPLC-MS

规模以上非靶向代谢组学剖析血清超高效液相色谱 - 质谱（UPLC-MS）

Non-targeted metabolite profiling by ultra performance liquid chromatography coupled with mass spectrometry (UPLC-MS) is a powerful technique to ...

科研

化学

Analyzing Protein Dynamics Using Hydrogen Exchange Mass Spectrometry

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 ...

CN-GELFrEE - Clear Native Gel-eluted Liquid Fraction Entrapment Electrophoresis

CN-GELFrEE  - 透明天然凝胶洗脱液体馏分偷天陷阱电泳

Protein complexes perform an array of crucial cellular functions. Elucidating their non-covalent interactions and dynamics is paramount for understanding ...

Real-time Breath Analysis by Using Secondary Nanoelectrospray Ionization Coupled to High Resolution Mass Spectrometry

利用二次 Nanoelectrospray 电离耦合高分辨质谱的实时呼吸分析

Exhaled volatile organic compounds (VOCs) have aroused considerable interest, since they can serve as biomarkers for disease diagnosis and environmental ...

Microprobe Capillary Electrophoresis Mass Spectrometry for Single-cell Metabolomics in Live Frog (Xenopus laevis) Embryos

Microprobe Capillary Electrophoresis Mass Spectrometry for Single-cell Metabolomics in Live Frog Xenopus laevis Embryos

活蛙 (爪蟾) 胚胎单细胞代谢的微探针毛细管电泳质谱

The quantification of small molecules in single cells raises new potentials for better understanding the basic processes that underlie embryonic ...

High-throughput and Comprehensive Drug Surveillance Using Multisegment Injection-Capillary Electrophoresis-Mass Spectrometry

New analytical methods are urgently needed to enable high-throughput, yet comprehensive drug screening, given an alarming opioid and prescription drug ...

Setup of Capillary Electrophoresis-Inductively Coupled Plasma Mass Spectrometry (CE-ICP-MS) for Quantification of Iron Redox Species (Fe(II), Fe(III))

Setup of Capillary Electrophoresis-Inductively Coupled Plasma Mass Spectrometry CE-ICP-MS for Quantification of Iron Redox Species FeII, FeIII

为铁氧化还原物种（Fe（II）、Fe（III）定量设置毛细管电泳-电感耦合等离子体质谱仪（CE-ICP-MS）

Dyshomeostasis of iron metabolism is accounted in the pathophysiological framework of numerous diseases, including cancer and several neurodegenerative ...

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

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 ...

Identification of Antibacterial Immunity Proteins in Escherichia coli using MALDI-TOF-TOF-MS/MS and Top-Down Proteomic Analysis

Identification of Antibacterial Immunity Proteins in Escherichia coli using MALDI-TOF-TOF-MS/MS and Top-Down Proteomic Analysis

使用MALDI-TOF-TOF-MS/MS和自上而下的蛋白质组学分析鉴定 大肠杆菌 中的抗菌免疫蛋白

This protocol identifies the immunity proteins of the bactericidal enzymes: colicin E3 and bacteriocin, produced by a pathogenic Escherichia coli strain ...

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

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 ...