Considerations for MALDI-Based Quantitative Mass Spectrometry Imaging Studies

Abstract

Significant advances in mass spectrometry imaging (MSI) have pushed the boundaries in obtaining spatial information and quantification in biological samples. Quantitative MSI (qMSI) has typically been challenging to achieve because of matrix and tissue heterogeneity, inefficient analyte extraction, and ion suppression effects, but recent studies have demonstrated approaches to obtain highly robust methods and reproducible results. In this perspective, we share our insights into sample preparation, how the choice of matrix influences sensitivity, construction of calibration curves, signal normalization, and visualization of MSI data. We hope that by articulating these guidelines that qMSI can be routinely conducted while retaining the analytical merits of other mass spectrometry modalities.

MS imaging of multicellular tumor spheroids and organoids as an emerging tool for personalized medicine and drug discovery

“…There are several commonly used normalization methods such as total ion count (TIC), median normalization or normalization to a manually selected m/z peak. 82 Among all these methods, TIC is the most commonly used one, where all mass spectra are divided by their TIC. This approach works well in homogenous samples.…”

“…96 Therefore, more considerations should be taken to achieve MALDI quantitative MSI (qMSI), including sample preparation protocols, the influence of sensitivity by matrix selection, construction of calibration curves, signal normalization and visualization of MSI data. 82 Although difficult, both absolute 97 and relative 98 quantitative studies have been completed using MALDI-MSI on tissue samples, mostly evaluating the concentration of small molecules 99 , with relative quantification studying concentration differences from experimental conditions. Absolute quantification experiments typically calculate the specific molecular concentration directly.…”

“…These steps help to reduce experimental variance and extract meaningful information based on different research purposes so that they ensure the accuracy and efficiency of data analysis. There are several commonly used normalization methods, such as TIC, median normalization, or normalization to a manually selected m/z peak ( 74 ). Among all these methods, TIC is the most commonly used one, where all mass spectra are divided by their TIC.…”

Natural Products, the Fourth Industrial Revolution, and the Quintuple Helix

“…Historically, quantitation of AMS data has proven to be challenging due to reproducibility, especially the uniform introduction of standards on to tissue surfaces and matrix ion suppression effects, 188,189 although improved techniques have been reported recently. 190,191 For the future, advances in database metabolite identification, the ability to quantify the data, and particularly to view drug distribution and metabolic profile changes over time in a continuous manner, would provide important information regarding catabolic and biosynthetic pathways, as well as the mechanisms and timing of metabolite translocation in plant and animal tissues. Enhancement in spatial resolution requires a corresponding increase in sensitivity.…”

Emerging applications in mass spectrometry imaging; enablers and roadblocks

“…Due to the well-documented issues with ion suppression analyte extraction and matrix effects, the fundamental aspects of MSI need to be considered in order to produce fully validated MSI methodology for quantitative analysis. [63][64][65] In Q-MSI, a calibration curve is necessary for the analyte of interest (AOI). Choosing a method to include standards into the experiment in order to construct calibration curves is dependent on the nature of the study, i.e.…”