MALDI FTICR IMS of Intact Proteins: Using Mass Accuracy to Link Protein Images with Proteomics Data

Abstract

MALDI imaging mass spectrometry is a highly sensitive and selective tool used to visualize biomolecules in tissue. However, identification of detected proteins remains a difficult task. Indirect identifications strategies have been limited by insufficient mass accuracy to confidently link ion images to proteomics data. Here we demonstrate the capabilities of MALDI FTICR MS for imaging intact proteins. MALDI FTICR IMS provides an unprecedented combination of mass resolving power (∼75,000 at m/z 5,000) and accuracy (<5 ppm) for proteins up to ∼12 kDa enabling identification based on correlation with LC-MS/MS proteomics data. Analysis of rat brain tissue was performed as a proof-of-concept highlighting the capabilities of this approach by imaging and identifying a number of proteins including N-terminally acetylated Thymosin β4 (m/z 4,963.502, 0.6 ppm) and ATP Synthase subunit ε (m/z 5,636.074, −2.3 ppm). MALDI FTICR IMS was also used to differentiate a series of oxidation products of S100A8 (m/z 10,164.03, −2.1 ppm), a subunit of the heterodimer calprotectin, in kidney tissue from mice infected with Staphylococcus aureus. S100A8 – M37O/C42O3 (m/z 10228.00, −2.6 ppm) was found to co-localize with bactierial microcolonies at the center of infectious foci. The ability of MALDI FTICR IMS to distinguish S100A8 modifications is critical to understanding calprotectin’s roll in nutritional immunity.

Dietary Manganese Promotes Staphylococcal Infection of the Heart

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Approaching cellular resolution and reliable identification in mass spectrometry imaging of tryptic peptides

“…Identification of intact proteins from tissue is difficult due to the limited mass resolution of time-of-flight mass spectrometers (TOF-MS), which is insufficient to resolve the complexity of these samples. A promising approach is the detection of intact proteins by Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) [18, 19]. A general limitation of direct analysis of intact proteins under imaging conditions is the limited mass range (typically up to 25 kDa), i.e., the majority of proteins cannot be detected.…”

Label-free molecular imaging of the kidney

“…Ions of interest are identified using one or a combination of several techniques, including accurate mass measurements 5,6 and tandem MS (MS/MS). 7–13 …”

“…13,115–117 The increasing public health threat of evolved antimicrobial resistance indicates that further study of hospital- and community-acquired infections caused by bacterial pathogens will be important for developing novel targets for therapeutic intervention. Kehl-Fie et al .…”