High-resolution tissue imaging on an orbitrap mass spectrometer by desorption electrospray ionization mass spectrometry.

The major methods for examining tissue samples include histology, autoradiography of radiolabeled compounds and fluorescence microscopy in conjunction with labeled antibodies. Mass spectrometry (MS) imaging has the potential to complement these methods and is advantageous for reasons that include the fact that more chemical information can be gleaned during a single experiment and that labeling is not required. Some of its limitations include ion suppression effects, the fact that targeted experiments are often more challenging, and, most importantly, that it is an invasive experiment. The principal methods for MS imaging are secondary ion mass spectrometry (SIMS),[1,2] which has the advantage of high spatial resolution, and matrix-assisted laser desorption/ionization (MALDI),[3,4] which allows for the detection of higher molecular weight species and has been used for high-resolution/highmass accuracy imaging of lipids from tissue. [5] More recently, attention has been given to imaging by ambient ionization methods, which allow for the examination of samples in situ, outside the vacuum of the mass spectrometer. Reports in the literature of ambient imaging include the use of desorption electrospray ionization mass spectrometry (DESIMS),[6] surface-sampling probe electrospray ionization[7] and laser ablation electrospray ionization (LAESI). [8] The first ambient imaging method, DESI-MS, analyzes condensed phase samples using a pneumatically assisted stream of charged microdroplets directed at the surface. [9] A thin liquid film collects on the surface, and the impact of incoming droplets on this thin film causes the ejection of small secondary droplets containing the dissolved analyte. [10] Reports in the literature include examples of imaging of thin tissue sections,[6,11,12] seaweed,[13] thin layer chromatography (TLC) plates,[14] ink on paper[15] and latent fingerprints. [16] One defining feature of ambient analysis is that, because samples are analyzed in their native state, little or no sample preparation is used. Any sample preparation or treatment is typically done concurrently with the analysis and not as a separate step. The principal benefits of ambient analysis, therefore, are a reduction in the time required for analysis and the ease of operation. One drawback is that untreated samples are often chemically complex, and this is particularly true in tissue imaging applications. High selectivity is therefore important for the successful implementation of ambient analysis. Increased selectivity in DESI over that available by unit resolution, single-stage MS, has been achieved by tandem MS, the use of in situ chemical reactions,[11] and by high-resolution MS.[17] High-resolution DESI experiments were reported in 2006 by Fernandez, Muddiman and their coworkers using Fourier transform ion cyclotron resonance (FT-ICR). Volny and coworkers recently extended this work by performing high m/z resolution and mass accuracy DESI imaging on an FT-ICR, but the spatial resolution was lower than previously reported for DESI imaging. [18] In this letter, we describe DESI-MS imaging of rat brain tissue sections using an orbitrap mass spectrometer (Thermo Scientific, San Jose, CA, USA), which has mass accuracy and resolving power in the same range as that of the FT-ICR. All the experiments were performed on an LTQ Orbitrap mass spectrometer (Thermo Scientific). Imaging experiments were done using the 30 k mass resolution setting, while the 100 k setting with lock masses was used for exact mass measurements to calculate molecular formulae. Mass standards were delivered directly to the mass spectrometer via an electrosonic spray ionization (ESSI) source concurrently with the DESI analysis. The [M + H]+ of reserpine and the [M + Na]+ ion of cyclosporine A, theoretical exact masses of 609.28121 and 1224.83114, respectively, were used as standards in the positive ion mode; the [M − H]− ion of taurocholate and the C15H15O6N3P3F20(C2F4)2 ultramark 1621 peak, with theoretical exact masses of 514.28385 and 1005.97266, respectively, were used in the negative ion mode. Frozen rat brain 720 760 800 840 880 920 m/z 0 20 40 60 80 100