Demonstrating voxel-by-voxel (v×v) single-point calibration in liver tissue by IR-MALDESI quantitative MSI
Data files
Sep 17, 2025 version files 3.37 GB
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0.03125_2to1_Solution.raw
46.68 MB
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0.0625_2to1_Solution.raw
56.40 MB
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0.125_2to1_Solution.raw
45.83 MB
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0.25_2to1_Solution.raw
43.09 MB
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0.5_2to1_Solution.raw
41.49 MB
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1.0_2to1_Solution.raw
37.79 MB
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Liver_031_1.raw
174.28 MB
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Liver_031_2.raw
154.51 MB
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Liver_031_3.raw
117.77 MB
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Liver_062_1.raw
159.87 MB
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Liver_062_2.raw
155.20 MB
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Liver_062_3.raw
114.97 MB
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Liver_1000_1.raw
150.19 MB
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Liver_1000_2.raw
145.19 MB
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Liver_1000_3.raw
109.67 MB
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Liver_125_1.raw
163.75 MB
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Liver_125_2.raw
150.57 MB
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Liver_125_3.raw
120.30 MB
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Liver_250_1.raw
162.46 MB
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Liver_250_2.raw
149.92 MB
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Liver_250_3.raw
113.40 MB
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Liver_500_1.raw
159.06 MB
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Liver_500_2.raw
146.33 MB
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Liver_500_3.raw
105.38 MB
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PRM_of_GSH_1.raw
42.88 MB
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PRM_of_GSH_2.raw
46.96 MB
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README.md
12.08 KB
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SIL_GSH_MS1_1.raw
127.89 MB
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SIL_GSH_MS1_2.raw
114.54 MB
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VxV_GSH_MS1_1.raw
90 MB
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VxV_GSH_MS1_2.raw
127.44 MB
Abstract
The quantification of glutathione (GSH) on a per-voxel basis was achieved by homogenously spraying a known amount of stable isotope labeled glutathione (SIL-GSH) on a microscope slide. Next, liver sections were mounted on top of the coated slides and imaged using IR-MALDESI MSI. Statistical analysis has shown high precision for V×V quantification over a wide concentration range, but the accuracy of this method is currently limited due to the sprayer configuration. Results support the feasibility of V×V quantification as evidence by concentration heatmaps. Additionally, voxel-based quantification provides benefits such as high specificity, straightforward sample preparation, and promising initial statistics analysis which will be advantageous for future qMSI studies.
Dataset DOI: 10.5061/dryad.b5mkkwhrk
Description of the data and file structure
Data collected for this manuscript. All files were converted to .mzML and then to .imzML format.
Files and variables
File: VxV_GSH_MS1_1.raw
Description: Replicate 1 of VxV quantification. Image of mouse liver at 7 μm where stable isotope labeled version glutathione (SIL-GSH) was sprayed underneath the liver section and served as a single point calibration at each voxel within the image. The concentration of endogenous GSH (NAT-GSH) was calculated using the Voxel-by-Voxel quantification tool in MSiReader.
File: VxV_GSH_MS1_2.raw
Description: Replicate 2 of VxV quantification. The same conditions from the first replicate of VxV quantification apply to replicate 2.
File: SIL_GSH_MS1_1.raw
Description: Replicate 1 of spatial curve quantification. Image of mouse liver at 7 μm where the SIL-GSH was spotted on top of the liver section at concentrations of 0, 0.03125, 0.625, 0.125, 0.25, 0.50, 1.0 mg/mL to output a calibration curve. The calibration curve was used to determine the average on tissue concentration of NAT-GSH for this liver section. The spatial calibration curve tool in MSiReader was utilized for the calibration curve output and calculations.
File: SIL_GSH_MS1_2.raw
Description: Replicate 2 of spatial curve quantification. The same conditions from the first replicate of the spatial calibration curve apply to replicate 2.
File: 0.03125_2to1_Solution.raw
Description: 2 to 1 solution with concentration of 0.03125 mg/mL. Image of sprayed glass slide with a 2 to 1 solution of NAT-GSH and SIL-GSH at a total analyte concentration of 0.03125 mg/mL. There were a total of 2704 scans in this image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios.
File: 0.0625_2to1_Solution.raw
Description: 2 to 1 solution with concentration of 0.0625 mg/mL. Image of sprayed glass slide with a 2 to 1 solution of NAT-GSH and SIL-GSH at a total analyte concentration of 0.0625 mg/mL. There were a total of 2704 scans in this image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios.
File: 0.125_2to1_Solution.raw
Description: 2 to 1 solution with concentration of 0.125 mg/mL. Image of sprayed glass slide with a 2 to 1 solution of NAT-GSH and SIL-GSH at a total analyte concentration of 0.125 mg/mL. There were a total of 2704 scans in this image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios.
File: 0.25_2to1_Solution.raw
Description: 2 to 1 solution with concentration of 0.25 mg/mL. Image of sprayed glass slide with a 2 to 1 solution of NAT-GSH and SIL-GSH at a total analyte concentration of 0.25 mg/mL. There were a total of 2704 scans in this image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios.
File: 0.5_2to1_Solution.raw
Description: 2 to 1 solution with concentration of 0.50 mg/mL. Image of sprayed glass slide with a 2 to 1 solution of NAT-GSH and SIL-GSH at a total analyte concentration of 0.50 mg/mL. There were a total of 2704 scans in this image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios.
File: 1.0_2to1_Solution.raw
Description: 2 to 1 solution with concentration of 1.0 mg/mL. Image of sprayed glass slide with a 2 to 1 solution of NAT-GSH and SIL-GSH at a total analyte concentration of 1.0 mg/mL. There were a total of 2704 scans in this image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios.
File: PRM_of_GSH_1.raw
Description: Replicate 1 of PRM-qMSI. Image of mouse liver at 7 μm where SIL-GSH was sprayed underneath the liver section and served as a single point calibration at each voxel within the image. The collision energy was set to 20% while the isolation width was set to 4 m/z. By applying the collision energy, a fragmentation pattern for NAT-GSH and SIL-GSH were observed. A peak at 272.0888 was identified as the endogenous GSH quantifier peak while a peak at 275.0926 was classified as the SIL-GSH quantifier peak. Those m/z values were used in the VxV tool in MSiReader to propagate concentration heatmaps of our quantifier peak.
File: PRM_of_GSH_2.raw
Description: Replicate 2 of PRM-qMSI. The same conditions from the first replicate of the PRM imaging apply to replicate 2.
File: Liver_031_3.raw
Description: Replicate 3 of Liver and 0.03125 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_031_1.raw apply to this data file.
File: Liver_031_2.raw
Description: Replicate 2 of Liver and 0.03125 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_031_1.raw apply to this data file.
File: Liver_031_1.raw
Description: Replicate 1 of Liver and 0.03125 mg/mL sprayed SIL-GSH. Image of mouse liver at 7 μm where SIL-GSH was sprayed underneath the liver section at 0.03125 mg/mL and served as a single point calibration at each voxel within the image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios. Biological variability is introduced due to the liver section.
File: Liver_062_3.raw
Description: Replicate 3 of Liver and 0.0625 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_062_1.raw apply to this data file.
File: Liver_062_1.raw
Description: Replicate 1 of Liver and 0.0625 mg/mL sprayed SIL-GSH. Image of mouse liver at 7 μm where SIL-GSH was sprayed underneath the liver section at 0.0625 mg/mL and served as a single point calibration at each voxel within the image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios. Biological variability is introduced due to the liver section.
File: Liver_062_2.raw
Description: Replicate 2 of Liver and 0.0625 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_062_1.raw apply to this data file.
File: Liver_125_3.raw
Description: Replicate 3 of Liver and 0.125 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_125_1.raw apply to this data file.
File: Liver_125_2.raw
Description: Replicate 2 of Liver and 0.125 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_125_1.raw apply to this data file.
File: Liver_125_1.raw
Description: Replicate 1 of Liver and 0.125 mg/mL sprayed SIL-GSH. Image of mouse liver at 7 μm where SIL-GSH was sprayed underneath the liver section at 0.125 mg/mL and served as a single point calibration at each voxel within the image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios. Biological variability is introduced due to the liver section.
File: Liver_250_3.raw
Description: Replicate 3 of Liver and 0.25 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_250_1.raw apply to this data file.
File: Liver_500_3.raw
Description: Replicate 3 of Liver and 0.50 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_500_1.raw apply to this data file.
File: Liver_250_2.raw
Description: Replicate 2 of Liver and 0.25 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_250_1.raw apply to this data file.
File: Liver_1000_3.raw
Description: Replicate 3 of Liver and 1.0 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_1000_1.raw apply to this data file.
File: Liver_250_1.raw
Description: Replicate 1 of Liver and 0.25 mg/mL sprayed SIL-GSH. Image of mouse liver at 7 μm where SIL-GSH was sprayed underneath the liver section at 0.25 mg/mL and served as a single point calibration at each voxel within the image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios. Biological variability is introduced due to the liver section.
File: Liver_500_2.raw
Description: Replicate 2 of Liver and 0.50 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_500_1.raw apply to this data file.
File: Liver_500_1.raw
Description: Replicate 1 of Liver and 0.50 mg/mL sprayed SIL-GSH. Image of mouse liver at 7 μm where SIL-GSH was sprayed underneath the liver section at 0.5 mg/mL and served as a single point calibration at each voxel within the image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios. Biological variability is introduced due to the liver section.
File: Liver_1000_2.raw
Description: Replicate 2 of Liver and 1.0 mg/mL sprayed SIL-GSH. The same conditions apply from File: Liver_1000_1.raw apply to this data file.
File: Liver_1000_1.raw
Description: Replicate 1 of Liver and 1.0 mg/mL sprayed SIL-GSH. Image of mouse liver at 7 μm where SIL-GSH was sprayed underneath the liver section at 1.0 mg/mL and served as a single point calibration at each voxel within the image. There were 6 bioinformatic ROIs employed where the abundance of NAT-GSH and SIL-GSH were exported. For each bioinformatic ROI, we calculated the average abundance of SIL-GSH, the average abundance ratio between NAT-GSH and SIL-GSH, as well as the statistical variance of those ratios. Biological variability is introduced due to the liver section.
Code/software
- Freestyle 1.8 SP 2
- MSiReader v3.10 (QMSI Spatial Calibration Curve Tool & QMSI Voxel-by-Voxel Quantification Tool)
Access information
Other publicly accessible locations of the data:
Data was derived from the following sources: