A Caffeic Acid Matrix Improves In Situ Detection and Imaging of Proteins with High Molecular Weight Close to 200,000 Da in Tissues by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging

Abstract

To our knowledge, this was the first study in which caffeic acid (CA) was successfully evaluated as a matrix to enhance the in situ detection and imaging of endogenous proteins in three biological tissue sections (i.e., a rat brain and Capparis masaikai and germinating soybean seeds) by matrix-assisted laser desorption/ ionization mass spectrometry imaging (MALDI-MSI). Our results show several properties of CA, including strong ultraviolet absorption, a super-wide MS detection mass range close to 200,000 Da, micrometer-sized matrix crystals, uniform matrix deposition, and high ionization efficiency. More high-molecularweight (HMW) protein ion signals (m/z > 30,000) could be clearly detected in biological tissues with the use of CA, compared to two commonly used MALDI matrices, i.e., sinapinic acid (SA) and ferulic acid (FA). Notably, CA shows excellent performance for HMW protein in situ detection from biological tissues in the mass range m/z > 80,000, compared to the use of SA and FA. Furthermore, the use of a CA matrix also significantly enhanced the imaging of proteins on the surface of selected biological tissue sections. Three HMW protein ion signals (m/z 50,419, m/z 65,874, and m/z 191,872) from a rat brain, two sweet proteins (mabinlin-2 and mabinlin-4) from a Capparis masaikai seed, and three HMW protein ion signals (m/z 94,838, m/z 134,204, and m/z 198,738) from a germinating soybean seed were successfully imaged for the first time. Our study proves that CA has the potential to become a standard organic acid matrix for enhanced tissue imaging of HMW proteins by MALDI-MSI in both animal and plant tissues.

The application of mass spectrometry imaging in traditional Chinese medicine: a review

“…Then in 2010, MALDI imaging exceeding 50 kDa was accomplished using a high mass detector [ 82 ]. More recently, with the application of a caffeic acid matrix, proteins with high molecular weight close to 200 kDa was successfully imaged by MALDI-MSI [ 83 ].…”

Highly multiplexed, label-free proteoform imaging of tissues by individual ion mass spectrometry

“…Modern MS instrumentation has reached the sensitivity for spatially resolved top-down proteomics suitable for imaging experiments (17,18). Matrix-assisted laser desorption/ionization (MALDI) is widely used for protein imaging (19,20) because of the broad mass range of proteome sampling (21)(22)(23). However, MALDI predominantly generates singly charged ions, which gives limited fragment information for direct top-down identification of intact proteins (24).…”

Combined Quantitative (Phospho)proteomics and Mass Spectrometry Imaging Reveal Temporal and Spatial Protein Changes in Human Intestinal Ischemia–Reperfusion

“…Recent studies have shown imaging of proteins up to 200 kDa. 13 , 14 MSI of trypsinized proteins can therefore—at least theoretically—increase the coverage of the proteome beyond this sensitivity limit. Moreover, the on-tissue digestion approach allows better integration of MSI data with liquid chromatography-tandem mass spectrometry (LC-MS/MS) of the same or an adjacent tissue section to enable the identification of the observed local peptide signals, especially when using high-mass resolution MSI instrumentation.…”

“…Intact proteins can be detected by MSI; however, their detection is limited by low ionization efficiency that limits sensitivity in the high mass range of the instrumentation. Recent studies have shown imaging of proteins up to 200 kDa. , MSI of trypsinized proteins can thereforeat least theoreticallyincrease the coverage of the proteome beyond this sensitivity limit. Moreover, the on-tissue digestion approach allows better integration of MSI data with liquid chromatography-tandem mass spectrometry (LC-MS/MS) of the same or an adjacent tissue section to enable the identification of the observed local peptide signals, especially when using high-mass resolution MSI instrumentation. , However, these high-mass resolution MSI methods are limited by a long acquisition time and consequently suffer from a low throughput.…”