This work is supported by ARRA NSF MRI Instrument Development grant #0923179 to FMF. We thank Aqua Asberry, Lab Coordinator for the Parker H. Petit Institute for Bioengineering and Biosciences Histology Core, for assistance with tissue sectioning.

1. Tissue Sectioning
<ol>
	<li>Store flash-frozen, whole tissue in -80 &deg;C freezer until ready for sectioning.</li>
	<li>Allow the tissue sample to reach the temperature in the cryomicrotome prior to sectioning (30 min). Set the blade and sample temperature to -30 &deg;C.</li>
	<li>Once tissue has reached the temperature, handling the specimen with tweezers, cut front or back of brain depending on what part of the brain is of interest for sufficient mounting surface (i.e. if the front of the brain is of pri...

<ol>
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Chem. 79, 8098-8106 (2007).</li><li>Nemes, P., Vertes, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Atmospheric-pressure+Molecular+Imaging+of+Biological+Tissues+and+Biofilms+by+LAESI+Mass+Spectrometry.">Atmospheric-pressure Molecular Imaging of Biological Tissues and Biofilms by LAESI Mass Spectrometry.</a> J. Vis. Exp. (43), e2097 (2010).</li><li>Van Berkel, G. J., Kertesz, V., Koeplinger, K. A., Vavrek, M., Kong, A. -. N. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Liquid+microjunction+surface+sampling+probe+electrospray+mass+spectrometry+for+detection+of+drugs+and+metabolites+in+thin+tissue+sections.">Liquid microjunction surface sampling probe electrospray mass spectrometry for detection of drugs and metabolites in thin tissue sections.</a> J. Mass Spectrom. 43, 500-508 (2008).</li><li>Tak&#225;ts, Z., Wiseman, J. M., Gologan, B., Cooks, R. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mass+Spectrometry+Sampling+Under+Ambient+Conditions+with+Desorption+Electrospray+Ionization.">Mass Spectrometry Sampling Under Ambient Conditions with Desorption Electrospray Ionization.</a> Science. 306, 471-473 (2004).</li><li>Venter, A., Sojka, P. E., Cooks, R. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Droplet+Dynamics+and+Ionization+Mechanisms+in+Desorption+Electrospray+Ionization+Mass+Spectrometry.">Droplet Dynamics and Ionization Mechanisms in Desorption Electrospray Ionization Mass Spectrometry.</a> Anal. Chem. 78, 8549-8555 (2006).</li><li>Costa, A. B., Cooks, R. 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S., Stokes, P., O'Connor, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+electrospray+solvent+composition+on+desorption+electrospray+ionisation+(DESI)+efficiency+and+spatial+resolution.">The effect of electrospray solvent composition on desorption electrospray ionisation (DESI) efficiency and spatial resolution.</a> Analyst. 135, 731-737 (2010).</li><li>Van Berkel, G. J., Kertesz, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+Sampling+and+Imaging+of+Analytes+Separated+on+Thin-Layer+Chromatography+Plates+Using+Desorption+Electrospray+Ionization+Mass+Spectrometry.">Automated Sampling and Imaging of Analytes Separated on Thin-Layer Chromatography Plates Using Desorption Electrospray Ionization Mass Spectrometry.</a> Anal. Chem. 78, 4938-4944 (2006).</li><li>Ifa, D. R., Wu, C., Ouyang, Z., Cooks, R. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Desorption+electrospray+ionization+and+other+ambient+ionization+methods:+current+progress+and+preview.">Desorption electrospray ionization and other ambient ionization methods: current progress and preview.</a> Analyst. 135, 669-681 (2010).</li><li>Eberlin, L. S., Ferreira, C. R., Dill, A. L., Ifa, D. R., Cooks, R. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Desorption+electrospray+ionization+mass+spectrometry+for+lipid+characterization+and+biological+tissue+imaging.">Desorption electrospray ionization mass spectrometry for lipid characterization and biological tissue imaging.</a> Biochim. Biophys. Acta. 1811, 946-960 (2011).</li><li>Wu, C., Ifa, D. R., Manicke, N. E., Cooks, R. 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S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Classifying+Human+Brain+Tumors+by+Lipid+Imaging+with+Mass+Spectrometry.">Classifying Human Brain Tumors by Lipid Imaging with Mass Spectrometry.</a> Cancer Res. 72, 645-654 (2012).</li><li>Wiseman, J. M., Ifa, D. R., Venter, A., Cooks, R. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ambient+molecular+imaging+by+desorption+electrospray+ionization+mass+spectrometry.">Ambient molecular imaging by desorption electrospray ionization mass spectrometry.</a> Nat. Protocols. 3, 517-524 (2008).</li><li>Van Berkel, G. J., Ford, M. J., Deibel, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thin-Layer+Chromatography+and+Mass+Spectrometry+Coupled+Using+Desorption+Electrospray+Ionization.">Thin-Layer Chromatography and Mass Spectrometry Coupled Using Desorption Electrospray Ionization.</a> Anal. Chem. 77, 1207-1215 (2005).</li><li>Lane, A. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Desorption+electrospray+ionization+mass+spectrometry+reveals+surface-mediated+antifungal+chemical+defense+of+a+tropical+seaweed.">Desorption electrospray ionization mass spectrometry reveals surface-mediated antifungal chemical defense of a tropical seaweed.</a> Proc. Natl. Acad. Sci. 106, 7314-7319 (2009).</li><li>Kertesz, V., Van Berkel, G. 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The optimization of the DESI source geometry is critical for successful MSI experiments. The multiple variables contributing to the alignment of the system directly affect sensitivity and image resolution. If during optimization, the experimenter has difficulties obtaining signal, we recommend using red Sharpie spot drawn on the slide as a benchmark; the dye, rhodamine 6G, m/z 443, produces a strong signal in the positive ion mode and can be used for initial optimization. Additionally, the solvent selection for DESI is c...

A correction was made to <a href="http://www.jove.com/video/50575" target="_parent">Imaging of Biological Tissues by Desorption Electrospray Ionization Mass Spectrometry</a>. There was an error with an author's name. The author's surname was appended with a missing character:
Rachel V. Bennett
instead of:
Rachel V. Bennet

Erratum: Imaging of Biological Tissues by Desorption Electrospray Ionization Mass Spectrometry

Untargeted imaging by mass spectrometry facilitates the acquisition of chemical information for discovery and hypothesis-generating applications. Targeted imaging of a known chemical of interest, on the other hand, can facilitate increased sensitivity and selectivity through specific method development. Mass spectrometry imaging (MSI) is most commonly performed on tissues using MALDI,1 secondary ion mass spectrometry (SIMS),2 and ambient ionization techniques, including desorption electrospray ionization (DESI),3 laser ablation-electrospray ionization (LAESI),4,5 and liquid micro-junction-surface sampling probe (LMJ-SSP).6 In MALDI and SIMS, samples have to be physically removed from the specimen, and have to be flat and thin, as they are analyzed under high-vacuum. MALDI requires coating of the sample with a radiation-absorbing matrix, adding an additional and cumbersome step to the sample preparation. SIMS has the highest lateral resolution, but bombardment with highly energetic particles causes extensive molecular fragmentation. Therefore, MSI by ambient methods fill a niche where soft analysis with minimal sample preparation is desirable. However, to date, all methods are still limited by the requirement of flat sample surfaces.	
	DESI uses a pneumatically-assisted charged solvent spray directed at the sample surface to desorb and ionize analytes.7 The working model for desorption and subsequent ionization by DESI is known as the "droplet pick-up model".8-10 The charged primary droplets produced by the DESI probe collide with the surface, wetting it and forming a thin film into which the analyte is dissolved by a solid-liquid microextraction mechanism8 Subsequent droplet collisions result in momentum transfer and takeoff of secondary droplets containing the material extracted from the surface.9,10 Ultimately, gas phase ions are believed to be produced through ESI-like processes following the ion evaporation, charge residue models or other models,11 however the precise ion formation process in DESI has yet to be experimentally proven.12 DESI sensitivity is strongly dependent upon the solubility of the analyte in the spray solvent, as desorption relies on the localized microextraction.13 	
	When used with a software-controlled sample stage, the sample is scanned unidirectionally with lane stepping underneath the DESI ionization probe, and through the time domain, m/z information is correlated with the chemical species' spatial distribution (Figure 1). Since the first proof of principle DESI-MSI experiment reported by Van Berkel and Kertesz in 2006,14 the technique has matured considerably,15 with reported applications in the analysis of lipids,3,16 drug metabolites,17,18 disease biomarkers,19 brain tissue,3,18,20 lung tissue,18 kidney tissue,18 testis tissue,18 adrenal glands,17 thin layer chromatography plates,21 and algae surfaces.22 The routine resolution of images obtained by DESI-MSI is 100-200 &mu;m, which is ultimately determined by the effective surface area extracted by the spray, but resolutions as low as 40 &mu;m have been reported.23-25 Such resolution and ease of analysis makes DESI-MSI appropriate for the rapid and simple analysis of biological tissue samples with surface areas in the 0.5-5 cm2 range, enabling the acquisition of valuable spatial information to better understand biological processes26. Here, as an example of a typical DESI-MSI application, we review the procedural details of conducting a successful experiment involving imaging of lipids in rat brain tissues. The two most critical steps in the protocol are the tissue preparation27 and DESI ion source optimization, as described below.

<table border="1" fo:keep-together.within-page="always">
		<tbody>
		 <tr>
			<td>Reagents</td>
			<td></td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Tissue-Tek O.C.T. Compound</td>
			<td>Sakura-Finetek</td>
			<td>4583</td>
			<td><a href="http://www.sakuraeu.com/products/showitem.asp?cat=11&amp;subcat=48" target="_blank">http://www.sakuraeu.com/products/showitem.asp?cat=11&amp;subcat=48</a></td>
		 </tr>
		 <tr>
			<td>Acetonitrile</td>
			<td>EMD</td>
			<td>AX0156-6</td>
			<td>OmniSolv, LC-MS Grade</td>
		 </tr>
		 <tr>
			<td>Acetic Acid</td>
			<td>Sigma Aldrich</td>
			<td>695092-500 ml</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Equipment</td>
			<td></td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Cryostat microtome</td>
			<td>Thermo Scientific</td>
			<td>CryoStar* NX70</td>
			<td>Any available microtome can be used for tissue sectioning <a href=" http://www.thermoscientific.com/ecomm/servlet/productsdetail?productId=13958375&amp;groupType=PRODUCT&amp;searchType=0&amp;storeId=11152&amp;from=search&amp;ca=cryostar" target="_blank"> http://www.thermoscientific.com/ecomm/servlet/productsdetail?productId=13958375&amp;groupType=PRODUCT&amp;searchType=0&amp;storeId=11152&amp;from=search&amp;ca=cryostar</a></td>
		 </tr>
		 <tr>
			<td>Omni Spray;DESI Spray Head</td>
			<td>Prosolia Inc.</td>
			<td></td>
			<td>Can also use the 2-D Omni Spray; Source kit instead of assembling components of imaging experiment <a href=" http://www.prosolia.com/sources.php" target="_blank"> http://www.prosolia.com/sources.php</a></td>
		 </tr>
		 <tr>
			<td>High Voltage Power Supply</td>
			<td>Stanford Research Systems, Inc.</td>
			<td>PS350/5000V-25W</td>
			<td><a href="http://www.thinksrs.com/products/PS300.htm" target="_blank">http://www.thinksrs.com/products/PS300.htm</a></td>
		 </tr>
		 <tr>
			<td>Rope heater, RTD, controller</td>
			<td>Omega</td>
			<td></td>
			<td><a href="http://www.omega.com/toc_asp/subsectionSC.asp?subsection=M02&amp;book=Heaters" target="_blank">http://www.omega.com/toc_asp/subsectionSC.asp?subsection=M02&amp;book=Heaters</a></td>
		 </tr>
		 <tr>
			<td>Labview</td>
			<td>National Instruments</td>
			<td>Version 7.1 </td>
			<td></td>
		 </tr>
		 <tr>
			<td>Translational stage</td>
			<td>Prior Scientific</td>
			<td>Optiscan II</td>
			<td><a href="http://www.prior.com/productinfo_auto_motorized_optiscan.html" target="_blank">http://www.prior.com/productinfo_auto_motorized_optiscan.html</a></td>
		 </tr>
		 <tr>
			<td>AccuTOF Mass Spectrometer</td>
			<td>JEOL</td>
			<td>JMS-T100LC</td>
			<td>Can use any mass spectrometer equipped with an extended capillary atmospheric pressure interface</td>
		 </tr>
		</tbody>
 </table>

imaging of biological tissues by desorption electrospray ionization mass spectrometry

Mass spectrometry imaging (MSI) provides untargeted molecular information with the highest specificity and spatial resolution for investigating biological tissues at the hundreds to tens of microns scale. When performed under ambient conditions, sample pre-treatment becomes unnecessary, thus simplifying the protocol while maintaining the high quality of information obtained. Desorption electrospray ionization (DESI) is a spray-based ambient MSI technique that allows for the direct sampling of surfaces in the open air, even in vivo. When used with a software-controlled sample stage, the sample is rastered underneath the DESI ionization probe, and through the time domain, m/z information is correlated with the chemical species' spatial distribution. The fidelity of the DESI-MSI output depends on the source orientation and positioning with respect to the sample surface and mass spectrometer inlet. Herein, we review how to prepare tissue sections for DESI imaging and additional experimental conditions that directly affect image quality. Specifically, we describe the protocol for the imaging of rat brain tissue sections by DESI-MSI.

Figure 3 shows a representative spectrum obtained from an untreated rat brain section. In the positive mode, the mass spectrum is dominated by phosphatidylcholines due to their high ionization efficiencies (attributed to the positively charged quaternary ammonium group). The total ion image of the tissue section is also shown in Figure 3, showing abundant signal across the entire brain section. Key lipids detected are identified in Table 1 through literature comparisons....

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Imaging of Biological Tissues by Desorption Electrospray Ionization Mass Spectrometry

Desorption electrospray ionization mass spectrometry (DESI-MS) is an ambient method by which samples, including biological tissues, can be imaged with minimal sample preparation. By rastering the sample below the ionization probe, this spray-based technique provides sufficient spatial resolution to discern molecular features of interest within tissue sections.

Mass spectrometry imaging (MSI) provides untargeted molecular information with the highest specificity and spatial resolution for investigating biological ...

imaging-biological-tissues-desorption-electrospray-ionization-mass

Research

JoVE Journal

Bioengineering

18.4K Views.  Georgia Institute of Technology. Desorption electrospray ionization mass spectrometry (DESI-MS) is an ambient method by which samples, including biological tissues, can be imaged with minimal sample preparation. By rastering the sample below the ionization probe, this spray-based technique provides sufficient spatial resolution to discern molecular features of interest within tissue sections.