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

1. 슐 리렌 사진 <ol><li> 테스트 지역의 설립 참고 : 시험 영역은 직접 거울 앞에 존재한다. <ol><li> 거울을지지하기에 충분히 큰 링 스탠드 클램프 구형 오목 면경 (직경 150mm, 초점 길이 1,500mm)을 조인다. 바닥에 수직으로 서 링에 미러 링 스탠드 클램프를 연결합니다. 현재의 연구는 3 피트 링 스탠드를 사용하지만, 임의의 높이로는, 소스보기 창에서 미러를 중심 수있을만큼 높이로 사용될 수있다. </li><li> 질량 분석기 소스의 측면에 링 스탠드와 거울을 배치합니다. 소스로서, 상기 미러에 평행 한면을하고, 같은 높이. </li><li> 그 중심이 질량 분석의 중심 소스 영역과 정렬되도록 미러를 배치. 악기의 일부 중복이 발생합니다. </li></ol></li><li> 컷오프, 카메라 및 광원 <ol><li> 절단 <ol><li> 삼각대의 상단에 금속판을 부착합니다. 플레이트는 면도날 광원을 모두 보유하는 플랫폼 역할을합니다. 면도날은 슐 리렌 사진에서 &quot;차단&quot;으로 알려진 역할을합니다. </li><li> 날카로운 에지가 수직이되도록 자석을 이용하여 금속판 면도날 첨부. </li><li> 거울, 3,000mm의 두 배 초점 거리에있는 거울에 맞춰 삼각대를 놓습니다. 미러로부터 반사 된 광의 경로에 직교하는 면도날 정렬. </li><li> 면도날의 날카로운 모서리가 약 거울의 중심과 정렬되도록 수동으로 삼각대의 높이를 조정합니다. 참고 : 미세 조정 후에 발생합니다. </li></ol></li><li> 카메라 <ol><li> 별도의 삼각대에 300mm의 망원 렌즈 디지털 카메라를 탑재합니다. </li><li> (전체 줌에서) 렌즈 뒤에 같은 평에 4cm 직접 그래서 카메라의 위치를면도날로 GHT. 이 때 렌즈 캡을 제거하지 마십시오. </li></ol></li><li> 옵션 모니터 <ol><li> 쉽게 실시간 슐 리렌 현상을 확인하기 위해 컴퓨터 모니터 또는 TV 카메라의 영상 출력을 연결한다. 참고 :이 추천하는 과정입니다. 이 절차를 사용하는 카메라의 종류에 따라 달라질 수있다. </li></ol></li><li> 핀홀 광원 <ol><li> 표지의 중심에 미세 구멍 (직경 약 0.6 mm)을 뚫는다 (이 경우에, 바이알을 캡 플래쉬 동일한 직경을 사용 하였다) 부착 될 수있는 / 광원에 녹화. 덮개가 완전히 손전등 렌즈를 커버하기에 충분한 직경을 가지고 있는지 확인하십시오. </li><li> 호일 테이프를 사용하여 손전등을 LED 200 루멘을 통해 커버를 부착합니다. 참고 : 손전등 따뜻한 얻을 것이다 고온 테이프를 권장합니다. </li></ol></li><li> 광원의 위치 <ol><li> 먼저 라를 사용SER 포인터 광원의 적절한 포지셔닝을 보장하기 위해 미러, 면도날, 그리고 카메라와 광원을 정렬한다. </li><li> 면도날 옆에 금속 접시에 레이저 포인터를 놓습니다. </li><li> 빔은 거울의 중심을 타격되도록 수동으로 레이저 포인터를 이동합니다. 빔의 약 절반이 차단되도록 상기 반사 된 빔이 면도날 직교 교차되도록 필요한 조정한다. </li><li> 수동 빔 정렬이 1.2.5.3으로 달성되지 않은 경우 면도날에 직접 레이저 포인터 빔을 목표로하는 거울의 위치를​​ 조정합니다. 주의! 레이저 포인터 또는 반사 된 광선을 직접 보지 마십시오. </li><li> 카메라에 렌즈 캡을 유지하면서 레이저 빔이 렌즈의 중앙에 있는지 확인. </li><li> 모든 것이 정렬하는 동안 적용 손전등 레이저 포인터를 교체합니다. 손전등은 레이저 포인터와 같은 방향에 있는지 확인하십시오. </li><li> 손전등을 켜고, 흰 종이를 사용하여, 컷오프에서 반사 된 빛을 관찰합니다. 빔이 컷오프에 작은 초점을 맞춘 자리가 있는지 확인합니다. </li><li> 컷오프와 반사 된 광선의 약 절반을 차단하기 위해 모든 수직 조정이 필요합니다. </li><li> 카메라의 렌즈 캡을 제거하고 거울에 초점을 맞 춥니 다. 주 : 카메라 / 렌즈는 수동 초점 모드를 사용하는 것이 좋습니다. </li></ol></li></ol></li></ol> 2. 예 시험 대상 : 질량 분석 이온화 소스 <ol><li> 수동 노즐의 단부와 질량 분석기의 입구 사이에 10mm의 거리를두고, 테스트 영역 내의 질량 스펙트럼 이온 소스를 정렬. </li><li> 수동 질소 소스를 통해 흐르도록 주변 소스 니들 밸브를 연다. </li><li> 질량 분석 장치를 제어하는​​ 데 사용되는 소프트웨어를 연다. 이 연구를 위해 사용 된 소프트웨어는 &quot;SQ 드라이버 &#39;이었다. Fi를 클릭합니다제작 : 다음 적절한 조정 파일을 선택 -open-. </li><li> 수동 조정이 개방되면 주변 소스에 모든 전압 및 온도를 적용합니다. 각 질량 분석기는이 단계에 대한 자신의 소프트웨어를해야합니다. 수동 조정이 개방되면 현재의 연구에서, 버튼 &quot;소스 전압이 꺼져&quot;하고 버튼 &quot;모든 가스 히터가 꺼져&quot;이 작업을 수행합니다. </li><li> 온도가 증가함에 따라 상기 디지털 카메라의 뷰 화면의 슐 리렌 장치와 상기 노즐로부터 방출되는 흐름의 외관을 관찰한다. 노즐의 끝에서 나오는 가스 스트림 ( &quot;결과&quot;절에 설명 참조) 준수하십시오. 가스 스트림은 카메라의 뒷면에서 볼 수 있으며, 혹은 LCD 모니터에 직접 볼 수있다. </li><li> 카메라에서 영상을 기록하거나, 가스 스트림의 사진을 찍는 중 하나에 의해 이미지를 수집 일단 원하는 이미지가 카메라에서 실시간으로 시각화된다. </li><li> 가 camer으로 컴퓨터에 수집 된 화상 (들)을 전송메모리 카드 또는 USB 연결하고 사용자가 선택한 소프트웨어로 이미지를 볼 수 있습니다. </li></ol> 수집 된 이미지에서 스프레이 절반 각도 3. 결정 <ol><li> 이미지보기 소프트웨어를 사용하여 수집 된 이미지를 열고 수집 된 이미지 (들)을 인쇄. </li><li> 자를 사용하여 흐름 방향으로 가스 흐름과 평행 한 중심 축선을 정의하는 인쇄 된 이미지 (들)의 선을 그린다. </li><li> 눈금자를 사용하여 인쇄 된 이미지 (들)에서 시각화 가스 스트림의 가장자리를 따라 선을 그립니다. 이는 비디오 포맷에 존재하는 반짝임에 기록 된 비디오에서 잘 시각화 될 수있다; 인쇄 된 이미지의 가장자리를 식별하는 데 도움이 사용합니다. 분무 반 화각의 범위를 획득하기 위해 상기 가스 흐름의 바깥 쪽 가장자리를 표시한다. </li><li> 중심 축 및 각도기를 사용하여 3.2으로 그린​​ 선 사이에 생성 된 각도를 측정한다. </li></ol>

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There are no competing financial interests with this article.

이 프로토콜을 시도하기 전에 해결해야 할 몇 가지 고려 사항이 있습니다. 소스 및 미러 질량 분석기 주위 공간에 더하여, 충분한 개방 공간은 거울의 두 배의 초점 거리를 수용 할 수 있어야. 또한, 미러의 크기는 궁극적 연구중인 소스의 크기에 의해 결정된다. 미러가 너무 작 으면, 소스가 완전히 가시화되지 않는다. 이는 원본 커버가 슐 리렌 포토 이미징 기술을 구현하기 위해 제거되어야의 전...

질량 분석, 분자량 식별에 사용할 수있는 분석 도구는 현재까지 가장 강력한 분석 기술 중 하나가되고있다. 지난 10 년간 새로운 주변 이온화 소스의 모든 호스트는 질량 분석 검출을 위하여 이용 될 수있다. 이 논문에서 수집 한 데이터의 경우, 직접 시료 분석 (DSA) 소스가 이용되었다. 이러한 소스는 매우 다양한 있지만, 실제 이온화 프로세스의보다 상세한 기술은 그 목적 및 최적화의 연장이 필요하다. 이 실험의 목적은 슐 리렌 그래피이라는 기술을 사용하는 장치에서 질소 기류 가시화 통해 주변 소스에서 이온화 과정의 더 나은 이해를 얻을 수있다. 과학 연구는 종종 연구의 목적은 육안으로 투명 경우 어려운 관찰을 시작합니다. 슐 리렌 사진은 눈에 보이지 않는 수 있도록하는 기술이다투명 용지 (1) 내의 굴절률의 변화에 의존을 통해 볼 수있게합니다. 굴절률의 불균일성 시각화를 허용 빛의 왜곡을 야기한다. 슐 리렌 기법은 통상적으로 전기 영동 2-5 단백질 밴드를 시각화 탄도 모델링, 항공 우주 공학, 일반적인 가스 검출 및 모니터링 흐름을 포함한 특수 다양한 분야에 때때로 사용된다. 대부분의 주변 이온화 소스는 이온화를 촉진하기 위해 기류를 사용한다. 다양한 조건 그러나이 실험의 파라미터는 주위 실험실 공기와 상이한 굴절률 가스의 이용을 포함하며, 소스 옵션 존재할 수있다. 이 특정 연구는 뜨거운 질소를 사용합니다. 굴절률 단지 작은 차이는 주로 때문에, RT (6)에서의 가스 흐름 및 공기 순수한 질소 사이에 관찰되는 것을 주목해야한다적외선은 대부분 질소로 구성되어있다. 이러한 문제로 인해 가스가 관찰 할 굴절률 상당한 정도로 변화를 일으키는 기류 순수한 질소의 고온 이때 극복된다. 리얼 타임 (DART)에 대기압 잔광 (FAPA) 8-10, 및 직접 분석을 흐르는 이러한 탈착 대기 화학 이온화 (DAPCI) 7과 같은 다른 질량 분석 원은 11 이온화 소스는 슐 리렌 사진을 사용했다. 이 프로토콜의 목적은 기본 슐 리렌 사진 구성을 사용하여 주변의 이온화를 공부하는 방법에 대해 설명하는 것입니다. 이 기술은, 그러나, 가스 흐름을 포함하는 다른 분석 기법의 임의의 수에 적용 할 수있다.

<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>
		</tr>
		<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>
		</tr>
		<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>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Laser Pointer</td>
			<td>Apollo</td>
			<td>MP1200</td>
			<td>or equvilent&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">razor blade</td>
			<td>Blue Hawk</td>
			<td>34112</td>
			<td>or equvilent&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">small drill bit #73</td>
			<td>CML Supply</td>
			<td>503-273</td>
			<td>or equvilent&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Protractor</td>
			<td>Sterling&#160;</td>
			<td>582</td>
			<td>or equvilent&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Hose Clamp</td>
			<td>Trident</td>
			<td>720-6000L</td>
			<td>or equvilent&#160;</td>
		</tr>
	</tbody>
</table>

visualization of ambient mass spectrometry with the use of schlieren photography

이 원고는 슐 리렌 사진을 사용하여 질량 분석 주변 이온화 소스를 시각화하는 방법에 대해 설명합니다. 적절 질량 분석을 최적화하기 위하여, 특성화 및 소스의 물리적 원리를 이해하는 것이 필요하다. 대부분의 상용 주변 이온화 소스는 분석 물질의 이온화를 촉진하기 위해, 질소, 헬륨, 또는 대기 공기 제트를 이용한다. 결과적으로 슐 리렌 그래피 실시간으로 시각화 스트림과 주변 공기 사이의 굴절률의 차이를 이용하여 기체 스트림을 시각화 할 수있다. 기본 설정은 카메라, 거울, 손전등, 그리고 면도날이 필요합니다. 적절히 구성된 경우, 소스의 실시간 이미지는 반사를 보면서 관찰된다. 이는 소스 작용 기전에 대한 통찰력을 허용하고, 그 최적화 경로가 설명 될 수있다. 라이트는 달리 눈에 보이지 않는 상황에 흘려된다.

질량 분석기의 이온화 소스를 포함 슐 리렌 설정의 개략적 인 모든 슐 리렌 구성 요소가 제대로 정렬되어 그림 1.에서 찾아 볼 수있다, 시험 영역 내의 가스는 어둡고 밝은 영역을 대조로 볼 수있다. (2)이 대비 될 수있는 방법을 보여줍니다 그림 질량 스펙트럼 소스 변경되는 질소 제트류의 형상은 노즐 크기가 감소함에 따라 방법을 ?...

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.

슐 리렌 사진의 사용과 주변 질량 분석의 시각화

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

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JoVE Journal

Chemistry

8.2K 조회수.  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.

비디오: 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.

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

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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.

전자 분무 이온화 질량 분석에 의해 Dicysteinyl Tetrapeptides와 수은 (II)의 복합체 형성에 관한 연구

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.

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.

합성 및 올리고 peptoids의 질량 분석 분석

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.

비행 시간 보조 이온 질량 분석기를 사용 하 여 금속 페인트 인터페이스의 이미징 부식

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.

단일 현탁액 세포에서 약물 및 대사 산물의 질량 분석 분석을 위한 단일 프로브와 결합된 통합 셀 조작 플랫폼

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.

나노 물질에 의해 획득 된 단백질과 대사 산물 코로나특성화를위한 모세관 전기 전구 질량 분광법 접근법

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.

가스 크로마토그래피 질량 분광법은 토탈 기화 고체-위상 미세 추출과 결합하여 법의학 도구로 사용됩니다.

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.

질량 분광법에 의한 단백질 고차 구조를 연구하기 위한 디틸피로카보네이트와 공유 라벨링

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.

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