Streamlined Multimodal DESI and MALDI Mass Spectrometry Imaging on a Singular Dual-Source FT-ICR Mass Spectrometer

Abstract

The study of biological specimens by mass spectrometry imaging (MSI) has had a profound influence in the various forms of spatial-omics over the past two decades including applications for the identification of clinical biomarker analysis; the metabolic fingerprinting of disease states; treatment with therapeutics; and the profiling of lipids, peptides and proteins. No singular approach is able to globally map all biomolecular classes simultaneously. This led to the development of many complementary multimodal imaging approaches to solve analytical problems: fusing multiple ionization techniques, imaging microscopy or spectroscopy, or local extractions into robust multimodal imaging methods. However, each fusion typically requires the melding of analytical information from multiple commercial platforms, and the tandem utilization of multiple commercial or third-party software platforms-even in some cases requiring computer coding. Herein, we report the use of matrix-assisted laser desorption/ionization (MALDI) in tandem with desorption electrospray ionization (DESI) imaging in the positive ion mode on a singular commercial orthogonal dual-source Fourier transform ion cyclotron resonance (FT-ICR) instrument for the complementary detection of multiple analyte classes by MSI from tissue. The DESI source was 3D printed and the commercial Bruker Daltonics software suite was used to generate mass spectrometry images in tandem with the commercial MALDI source. This approach allows for the generation of multiple modes of mass spectrometry images without the need for third-party software and a customizable platform for ambient ionization imaging. Highlighted is the streamlined workflow needed to obtain phospholipid profiles, as well as increased depth of coverage of both annotated phospholipid, cardiolipin, and ganglioside species from rat brain with both high spatial and mass resolution.

Keywords: 3D printed/printing; Fourier transform ion cyclotron resonance (FT-ICR); desorption electrospray ionization (DESI); lipidomics; mass spectrometry imaging (MSI); matrix-assisted laser desorption/ionization (MALDI); multimodal imaging.

Figure 1

(A) is a digitized scan of the sections profiled by DESI-MSI, with corresponding images for the following m/z windows: (B) (820.4522 ± 0.0075), (C) (820.5050 ± 0.0075), (D) (820.5203 ± 0.0075), and (E) (820.5357 ± 0.0075). Images are created from inputting the transient of a DESI serial acquisition into the MALDI-MSI data file, reprocessing in FTMS Processing 2.2.0 (Bruker Daltonics, Bremen, Germany), and proper alignment of pixels in the sequence in FlexImaging 5.0 (Bruker Daltonics, Bremen, Germany). Highlighted to the right of the images in (B–E) are the artifacts produced from continuously scanning during the fly back and step. Line scans are aligned post-manual triggering of acquisition in FTMS Control 2.1.150 and subsequently sending the file of ASC II commands to the positioners. (F) represents an H&E-stained serial section. In the images, the highest ion intensities are white and the lowest ion intensities are black.

Figure 2

(A) is an exported average spectrum of every pixel from MALDI-MSI from FlexImaging 5.0 (Bruker Daltonics, Bremen, Germany). (B) is a zoomed view of the phospholipid profile from m/z 700 to 900. (C) is a zoomed view highlighting the matrix clusters formed from 1,5-DAN in the positive ion mode from m/z 200 to 725 marked by asterisks. (D) is a zoomed view of the putative cardiolipin and ganglioside profile from m/z 1425 to 1675.

Figure 3

(A) is an exported average spectrum of every pixel from DESI-MSI from FlexImaging 5.0 (Bruker Daltonics, Bremen, Germany), (B) is a zoomed view of the phospholipid profile from m/z 700 to 900, (C) is a zoomed view highlighting the matrix cluster formed from 1,5-DAN in the positive ion mode from m/z 200 to 725, and (D) is a zoomed view of the putative cardiolipin and ganglioside profile from m/z 1425 to 1675.

Figure 4

(A) is a digitized scan of the left section profiled by DESI-MSI, (D) is a completed H&E-stained serial section collected 220 µm from the DESI sampled tissue, with (B) (1570.2060 ± 0.05), (C) (1568.2100 ± 0.05), (E) (1520.2360 ± 0.05), (F) (1542.2190 ± 0.05), and (G) (1558.1860 ± 0.05) corresponding to the top five images in a m/z window from 1400 to 1600. In the images, the highest ion intensities are white and the lowest ion intensities are black.

Figure 5

Individual zoomed views from the averaged spectra of all pixels of the potassiated adducts of PC 32:0 and PC 36:4 from m/z ranges of 794.42 to 798.61 and 820.43 to 820.57, respectively. This area of 210 mDa contains several species annotated with varying resolution from MALDI (top, red) and DESI (bottom, blue). The PC main adduct is marked with an asterisk for each respective peak.

Figure 6

Photograph of the sourced mounted on the mass spectrometer, highlighting the Agilis LS-25-27 array and UC2 controller (A), the capillary extension and nanospray gas diverter (B), the removed endplate and capillary nub (C), and the standard sprayer electrospray emitter (D).