Name: Visualization of Ambient Mass Spectrometry with the Use of Schlieren Photography (Video) | JoVE Journal
Uploaded: 2016-06-20T15:00:00.000+00:00
Duration: 6 min 49 s
Description: Learn to visualize gas streams of ambient ionization sources using schlieren photography for mass spectrometry optimization. This method reveals physical phenomena affecting ionization efficiency and source performance.

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

1. Schlieren Photography
<ol>
	<li>Establishment of the Test Region 
	Note: The test region exists directly in front of the mirror.
	<ol>
		<li>Clamp a spherical concave mirror (150 mm diameter, focal length 1,500 mm) in a ring stand clamp large enough to support the mirror. Attach the ring stand clamp with the mirror to a ring stand perpendicular to the floor. The current study used a 3 foot ring stand, but any height can be used as long as it is tall enough to be able to center the mirror in the viewing window of the source.</li>
		<li>Place the ring stand and mirror to the side of the mass spectrometer source. Make the face of the mirror parallel to, and at the same height, as the source.</li>
		<li>Position the mirror so its center is aligned with the center source region of the mass spectrometer. Some overlap of the instrument will occur.</li>
	</ol>
	</li>
	<li>Cutoff, Camera and Light Source
	<ol>
		<li>Cutoff
		<ol>
			<li>Attach a metal plate to the top of the tripod. The plate will serve as a platform to hold both the razor blade and the light source. The razor blade acts as what is known as the &#34;cutoff&#34; in schlieren photography.</li>
			<li>Attach the razor blade to the metal plate using a magnet so that the sharp edge is vertical.</li>
			<li>Place the tripod in line with the mirror at twice the focal length of the mirror, 3,000 mm. Align the razor blade orthogonal to the path of light reflected from the mirror.</li>
			<li>Manually adjust the height of the tripod so that the sharp edge of the razor blade is approximately aligned with the center of the mirror. 
			NOTE: Fine adjustment will happen later.</li>
		</ol>
		</li>
		<li>Camera
		<ol>
			<li>Mount a digital camera with 300 mm telephoto lens on a separate tripod.</li>
			<li>Position the camera so the lens (when at full zoom) is 4 cm directly behind and at the same height as the razor blade. Do not remove the lens cap at this time.</li>
		</ol>
		</li>
		<li>Optional Monitor
		<ol>
			<li>Connect the video output of the camera to a computer monitor or TV to easily view the schlieren phenomenon in real time. 
			NOTE: This is a recommended process. This procedure may vary depending on the type of camera used.</li>
		</ol>
		</li>
		<li>Pinhole Light Source
		<ol>
			<li>Drill a small hole (approximately 0.6 mm in diameter) into the center of a cover (in this case, a vial cap the same diameter of the flashlight was used) which can be attached/taped to the light source. Ensure that the cover has sufficient diameter to completely cover the flashlight lens.</li>
			<li>Attach the cover over a 200 lumen LED flashlight using foil tape. 
			NOTE: The flashlight will get warm and a high temperature tape is recommended.</li>
		</ol>
		</li>
		<li>Light Source Positioning
		<ol>
			<li>First use a laser pointer to align the light source with the mirror, razor blade, and camera, to ensure proper positioning of the light source.</li>
			<li>Place the laser pointer on the metal plate next to the razor blade.</li>
			<li>Manually move the laser pointer so the beam is hitting the center of the mirror. Adjust as necessary to ensure the reflected beam intersects orthogonally to the razor blade so that approximately half of the beam is blocked.</li>
			<li>Manually adjust the mirror&#39;s position to aim the beam of the laser pointer directly at the razor blade if the beam alignment was not achieved in 1.2.5.3. 
			CAUTION! Do not look directly into the laser pointer or the reflected beam.</li>
			<li>Ensure that the laser beam is centered on the lens while keeping the lens cap on the camera.</li>
			<li>Replace the laser pointer with the covered flashlight while everything is aligned. Ensure that the flashlight is in the same orientation as the laser pointer.</li>
			<li>Turn on the flashlight and, using a piece of white paper, observe the reflected light at the cutoff. Ensure that the beam is a small focused spot at the cutoff.</li>
			<li>Make any vertical adjustments necessary to block approximately half of the reflected light beam with the cutoff.</li>
			<li>Remove the lens cap on the camera and focus on the mirror. 
			NOTE: It is recommended that the camera/lens be used in manual focusing mode.</li>
		</ol>
		</li>
	</ol>
	</li>
</ol>
2. Example Test Object: Mass Spectrometry Ionization Source
<ol>
	<li>Manually align the mass spectrometry ion source within the test region, with a distance of 10 mm between the end of the nozzle and the inlet of the mass spectrometer.</li>
	<li>Manually open the needle valve to the ambient source allowing nitrogen to flow through the source.</li>
	<li>Open the software used to control the mass spectrometer. For this study, the software used was &#34;SQ driver&#34;. Click on file -open- then select the appropriate tune file.</li>
	<li>Apply all voltages and temperatures to the ambient source once the manual tune is opened. Each mass spectrometer will have its own software for this step. For the current study, once the manual tune is open, click the button &#34;Source Voltage is off&#34; and the button &#34;All gas and heaters are off&#34; to perform this task.</li>
	<li>Observe the appearance of the flow exiting the nozzle with the schlieren apparatus on the view screen of the digital camera as the temperature increases. Observe the gas stream (see description in &#34;Results&#34; section) coming out of the end of nozzle. The gas stream can be viewed on the back of the camera, or it can be viewed directly on a LCD monitor.</li>
	<li>Collect the image by either recording a video from the camera, or taking a picture of the gas stream, once desired images are visualized live on the camera.</li>
	<li>Transfer the picture(s) collected to a computer with the camera memory card or USB connection and view the image with software of your choosing.</li>
</ol>
3. Determination of Spray Half Angle from a Collected Image
<ol>
	<li>Open the collected image using an image viewing software and print out the collected image(s).</li>
	<li>Draw a line on the printed image(s) defining the center axis of the gas stream parallel to the direction of flow using a ruler.</li>
	<li>Draw a line along the edge of the visualized gas stream on the printed image(s) using a ruler. This can be visualized better from a recorded video due to a shimmer that is present in video format; use this to help identify the edge in the printed images. Mark the outer edges of the gas streams to obtain a range for the spray half angle.</li>
	<li>Measure the angle produced between the center axis and the line drawn in 3.2 using a protractor.</li>
</ol>

<ol><li>Settles, G. S. <a target="_blank" href="http://scholar.google.com/scholar?hl=en&safe=off&q=author%3aSettles%2C+GS+%22Schlieren+and+Shadowgraph+Techniques%3A+Visualization+Phenomena+in+Transparent+Media%22">Schlieren and Shadowgraph Techniques: Visualization Phenomena in Transparent Media</a>. , 1st, Springer-Verlag. Germany. (2001).</li>
<li>Strawa, A. W., Chapman, G. T., Arnold, J. O., Canning, T. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Ballistic+range+and+aerothermodynamic+testing%5BTitle%5D)+AND+%22J.+Aircraft%22%5BJournal%5D)">Ballistic range and aerothermodynamic testing.</a> J. Aircraft. 28 (7), 443-449 (1991).</li>
<li>Settles, G. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Imaging+gas+leaks+by+using+schlieren+optics%5BTitle%5D)+AND+%22Pipeline+%26+Gas+Journal%22%5BJournal%5D)">Imaging gas leaks by using schlieren optics.</a> Pipeline & Gas Journal. 226 (9), 28-30 (1999).</li>
<li>Takagi, T., Kubota, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+application+of+schlieren+optics+for+detection+of+protein+bands+and+other+phenomena+in+polyacrylamide+gel+electrophoresis%5BTitle%5D)+AND+%22Electrophoresis%22%5BJournal%5D)">The application of schlieren optics for detection of protein bands and other phenomena in polyacrylamide gel electrophoresis.</a> Electrophoresis. 11 (5), 361-366 (1990).</li>
<li>Clark, I. G., Cruz, J. R., Huges, M. F., Ware, J. S., Madlangbayan, A., Braun, R. D. Aerodynamic and Aeroelastic Characteristics of a Tension Cone Inflatable Aerodynamic Decelerator. Proceedings from the 20th AIAA Aerodynamic Decelerator Systems Technology Conference and, May 7, 2009, Seattle, , (2009).</li>
<li>Froome, K. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+Refractive+Indices+of+Water+Vapour%2C+Air%2C+Oxygen%2C+Nitrogen+and+Argon+at+72+kMc%2Fs%5BTitle%5D)+AND+%22Proc.+Phys.+Soc.+B%22%5BJournal%5D)">The Refractive Indices of Water Vapour, Air, Oxygen, Nitrogen and Argon at 72 kMc/s.</a> Proc. Phys. Soc. B. 68, 833-835 (1955).</li>
<li>Winter, G. T., Wilhide, J. A., LaCourse, W. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Characterization+of+a+Direct+Sample+Analysis+%28DSA%29+Ambient+Ionization%5BTitle%5D)+AND+%22J.+Am.+Soc.+Mass+Spectrom%22%5BJournal%5D)">Characterization of a Direct Sample Analysis (DSA) Ambient Ionization.</a> J. Am. Soc. Mass Spectrom. 26 (9), 1502-1507 (2015).</li>
<li>Pfeuffer, K. P., Schaper, J. N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Halo-Shaped+Flowing+Atmospheric+Pressure+Afterglow%3A+A+Heavenly+Design+for+Simplified+Sample+Introduction+and+Improved+Ionization+in+Ambient+Mass+Spectrometry%5BTitle%5D)+AND+%22Anal.+Chem%22%5BJournal%5D)">Halo-Shaped Flowing Atmospheric Pressure Afterglow: A Heavenly Design for Simplified Sample Introduction and Improved Ionization in Ambient Mass Spectrometry.</a> Anal. Chem. , 7512-7518 (2013).</li>
<li>Pfeuffer, K. P., Shelley, J. T., Ray, S. J., Hieftje, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Visualization+of+Mass+Transport+and+Heat+Transfer+in+the+FAPA+Ambient+Ionization+Source%5BTitle%5D)+AND+%22J.+Anal.+At.+Spectrom%22%5BJournal%5D)">Visualization of Mass Transport and Heat Transfer in the FAPA Ambient Ionization Source.</a> J. Anal. At. Spectrom. 28 (379-387), 379-387 (2013).</li>
<li>Pfeuffer, K. P., Ray, S. J., Hieftje, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Measurement+and+Visualization+of+Mass+Transport+for+the+Flowing+Atmospheric+Pressure+Afterglow+%28FAPA%29+Ambient+Mass-Spectrometry+Source%5BTitle%5D)+AND+%22J.+Am.+Soc.+Mass+Spectrom%22%5BJournal%5D)">Measurement and Visualization of Mass Transport for the Flowing Atmospheric Pressure Afterglow (FAPA) Ambient Mass-Spectrometry Source.</a> J. Am. Soc. Mass Spectrom. 25 (5), 800-808 (2014).</li>
<li>Keelor, J. D., Dwivedi, P., Fernández, F. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((An+Effective+Approach+for+Coupling+Direct+Analysis+in+Real+Time+with+Atmospheric+Pressure+Drift+Tube+Ion+Mobility+Spectrometry%5BTitle%5D)+AND+%22J.+Am.+Soc.+Mass+Spectrom%22%5BJournal%5D)">An Effective Approach for Coupling Direct Analysis in Real Time with Atmospheric Pressure Drift Tube Ion Mobility Spectrometry.</a> J. Am. Soc. Mass Spectrom. 25 (9), 1538-1548 (2014).</li>
</ol>

There are no competing financial interests with this article.

There are several considerations which must be addressed prior to attempting this protocol. In addition to the space around the mass spectrometer for the source and mirror, enough open space must be available to accommodate the distance of twice the focal point of the mirror. Furthermore, the size of the mirror is ultimately decided by the size of the source that is under study. If the mirror is too small, the source will not be fully visualized. It is important to note that some, if not all, of the source covers must be...

Mass Spectrometry, an analytical tool available for molecular mass identification, has become one of the most powerful analytical techniques to date. Over the last decade a whole host of new ambient ionization sources have become available for mass spectrometry detection. For the data collected in this manuscript, the Direct Sample Analysis (DSA) source was utilized. Although these sources are extremely versatile, a more detailed knowledge of the physical ionization process is needed for its optimization and extension of purpose. The aim of this experiment is to gain a better understanding of the ionization process within the ambient sources through visualization of the nitrogen stream on the device using a technique called schlieren photography.
Scientific study often initiates through observation, which is difficult if the object of study is transparent to the naked eye. Schlieren photography is a technique that allows the invisible to become visible through relying on changes in the refractive index within transparent media1. The inhomogeneity of the refractive indices causes a distortion of the light allowing for visualization. The schlieren technique has been routinely used in a variety of specialty fields including ballistics modeling, aerospace engineering, general gas detection and flow monitoring, and at times to visualize protein bands in gel electrophoresis2-5.
Most ambient ionization sources use a stream of gas in order to facilitate the ionization. A wide range of conditions can exist for source options, however the parameters of this experiment must involve the utilization of a gas with a refractive index that differs from the surrounding lab air. This specific study utilizes hot nitrogen. It should be noted that only a small difference in refractive index is observed between pure nitrogen from the gas stream and air at RT6, mainly because air is composed mostly of nitrogen. This issue is overcome in this instance due to the high temperatures of the pure nitrogen in the gas stream which produces a significant enough change in refractive index for the gas to be observed.
Other mass spectrometry sources such as a Desorption Atmospheric Chemical Ionization (DAPCI)7, Flowing Atmospheric Pressure Afterglow (FAPA)8-10, and Direct Analysis in Real Time (DART)11 ionization sources have used schlieren photography. The intention of this protocol is to discuss how to study ambient ionization using a basic schlieren photography configuration. This technique, however, is applicable to any number of different analytical techniques that involve gaseous streams.

<table><tbody><tr><td>Flashlight</td><td>EAGTAC</td><td>D25A Ti</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>Spherical Concave Mirror</td><td>Anchor Optics</td><td>27633</td><td></td></tr><tr><td>Rebel EOS T2i</td><td>Canon</td><td>4462B001</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>300 mm telephoto lens</td><td>Canon</td><td>6473A003</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>Direct Sample Analysis (DSA) Ionization Source</td><td>PerkinElmer</td><td>MZ300560</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>Sq 300 MS with SQ Driver Software</td><td>PerkinElmer</td><td>N2910801</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>Ring Stand</td><td>Fisher Scientific</td><td>11-474-207</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>Laser Pointer</td><td>Apollo</td><td>MP1200</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>razor blade</td><td>Blue Hawk</td><td>34112</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>small drill bit #73</td><td>CML Supply</td><td>503-273</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>Protractor</td><td>Sterling&amp;nbsp;</td><td>582</td><td>or equivalent&amp;nbsp;</td></tr><tr><td>Hose Clamp</td><td>Trident</td><td>720-6000L</td><td>or equivalent&amp;nbsp;</td></tr></tbody></table>

visualization of ambient mass spectrometry with the use of schlieren photography

Visualization of Ambient Mass Spectrometry with the Use of Schlieren Photography

This manuscript outlines how to visualize mass spectrometry ambient ionization sources using schlieren photography. In order to properly optimize the mass spectrometer, it is necessary to characterize and understand the physical principles of the source. Most commercial ambient ionization sources utilize jets of nitrogen, helium, or atmospheric air to facilitate the ionization of the analyte. As a consequence, schlieren photography can be used to visualize the gas streams by exploiting the differences in refractive index between the streams and ambient air for visualization in real time. The basic setup requires a camera, mirror, flashlight, and razor blade. When properly configured, a real time image of the source is observed by watching its reflection. This allows for insight into the mechanism of action in the source, and pathways to its optimization can be elucidated. Light is shed on an otherwise invisible situation.

A schematic of the schlieren setup including the mass spectrometry ionization source can be found in Figure 1. When all schlieren components are properly aligned, gases within the test region can be seen as contrasting dark and light regions. Figure 2 illustrates how this contrast can be used to observe how the shape of the nitrogen jet flow from the mass spectrometry source changes as nozzle size decreases.
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This paper presents a protocol for the visualization of gaseous streams of an ambient ionization source using schlieren photography and mass spectrometry.

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

The overall goal of this procedure is to visualize the gas stream of an ambient ionization source used with mass spectrometry, for the purpose of understanding and optimizing the ionization source. This method can help answer key questions in the ambient ionization mass spectrometry field, such as ionization efficiency versus source position. The main advantage of this technique is using visualized gas streams with an ambient ionization source to better understand the physical phenomenon associated with the detection technique.Though this method can provide insight into ambient mass spectrometry, it can also be applied to other systems, such as physical phenomenon, which involve gas streams. An example would be the aerodynamics field. Generally, individuals new to this method will struggle, because initially it's difficult to figure out how to manually adjust the system to achieve proper alignment of the camera.First, clamp a spherical concave mirror in a ring stand clamp large enough to support the mirror. Attach the ring stand clamp with the mirror to a three foot ring stand perpendicular to the floor. Have the ring stand and mirror placed to the side of the mass spectrometer source.Adjust the face of the mirror so that it is parallel to, and at the same height, as the source. Then, position the mirror so that it's center is aligned with the center source region of the mass spectrometer. Next, attach a metal plate to top of the tripod.Attach a razor blade, which is known as the cut off, to the metal plate using a magnet, so that the sharp edge is vertical. Place the tripod in line with the mirror, at twice the focal length of the mirror. Then, align the razor blade orthogonal to the path of light reflected from the mirror.Manually adjust the height of the tripod so that the sharp edge of the razor blade is approximately aligned with the center of the mirror. Following this, mount a digital camera, with a 300 millimeter telephoto lens on a separate tripod. Position the camera so the lens is four centimeters directly behind, and at the same height, as the razor blade.Connect the video output of the camera to a computer monitor to easily view the Schlieren phenomenon in real time. Drill a small hole approximately zero point six millimeters in diameter into the center of a vial cap in order to attach it to a flashlight. Affix the vial cap over a 200 lumen LED flashlight, using high temperature foil tape.The experiment will only work if alignment of all the components is correct. Please take care to ensure proper positioning. To ensure proper positioning of the flashlight, place a laser pointer on the metal plate next to the razor blade.Manually move the laser pointer so the bean is hitting the center of the mirror. Adjust as necessary to ensure the reflected beam intersects orthogonally to the razor blade, so that approximately half of the beam is blocked. If the beam is not aligned, manually adjust the mirror's position to aim the beam of the laser pointer directly at the razor blade.After ensuring that the laser beam is centered on the lens, replace the laser pointer with a covered flashlight while everything is aligned. Next, turn on the flashlight. Using a piece of white paper, observe the reflected light at the cut off, ensuring that the beam is a small, focused spot.Make any vertical adjustments necessary to block approximately half of the reflected light beam with the cutoff. Then, remove the lens cap on the camera and focus on the mirror. Manually align the mass spectrometry ion source within the test region, with a distance of 10 millimeters between the end of the nozzle, and the inlet of the mass spectrometer.Following this, manually open the needle valve to the ambient source, allowing nitrogen to flow through it. Open the software used to control the mass spectrometer. Click on File, and select Open Tune.Then, select the appropriate Tune file. After opening the manual Tune, apply all voltages and temperatures to the ambient source. Observe the appearance of flow exiting the nozzle with the Schlieren apparatus, on the view screen of the digital camera at the temperature increases.Once the desired image is visualized on the camera, collect it by taking a picture of the gas stream. At this point, open the acquired image using image viewing software, and print it. Manually draw a line using a pencil and a ruler on the printed image, defining the center axis of the gas stream parallel to the direction of flow.Using the ruler, manually draw a line along the edge of the visualized gas stream on the printed image. Mark the outer edges of the gas stream to obtain a range for the spray half angle. Finally, measure the angle produced between the center axis and the line using a protractor.When all components in the Schlieren set up are properly aligned, gasses within the test region can be seen as contrasting dark and light regions. This contrast can be used to observe how the shape of the nitrogen jet flow from the mass spectrometry source changes as nozzle size decreases. A full, uncropped, Schlieren image of the source, and gas flow, illustrates the orientation of the test objects relative to the mirror, and shows what should be expected when the proper amount of light is cut off.If the cut off is either too high, or too low, poor images will result. With constant nozzle size the half angle increases accordingly with an increase in pressure, signifying an overall size increase of the gas stream. The half angle increases with larger nozzle diameter at constant pressure, which signifies an overall scaling increase in size of the nitrogen jet, coming out of the source as the nozzle diameter is increased.Once mastered, this technique can be done in one to two hours, if it is performed properly. After watching this video, you should have a good understanding of how to visualize the gas stream of an ambient ionization source. Don't forget that working with laser pointers can be extremely hazardous, and protective eyewear should always be worn while performing this procedure.

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Learn to visualize gas streams of ambient ionization sources using schlieren photography for mass spectrometry optimization. This method reveals physical phenomena affecting ionization efficiency and source performance.

Visualization of Ambient Mass Spectrometry with the Use of Schlieren Photography

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