Tools for the processing and analysis of single cell Proteoform imaging Mass Spectrometry (scPiMS)
- scores proteoforms against single cell ions
- optical single cell feature correlation
- scPiMS single cell feature extraction for downstream I2MS processing
- pathway-adjusted PAScore based on single sample GSVA analysis
- cluster analysis
- Heatmap
- PCA plots
- tSNE plot
- UMAP plot
Because the SingleCellApp.exe is a Windows Presentation Foundation (WPF) application that requires Windows 10 or later to run, all validation and processing were performed on Windows 10 22H2.
- No additional installation (.NET Runtime 8.0 packaged directly into application)
- The SingleCellApp.exe application is included in supplementary files in our publication located here:
IN SUBMISSION
- MATLAB 2023b or later
- Image Processing Toolbox
- Statistics and Machine Learning Toolbox
- Parallel Computing Toolbox
- Installs in less than 10 minutes on standard workstation
- R (v4.3.2+) and RStudio (2024.12.0+, Quarto v1.3.450)
- Installs in less than 10 minutes on standard workstation
- Download the 4 required demo files (Total_5272_cells.dmt, 165_proteoforms.fastp, pfr_annotation.xlsx, cell_annotation.xlsx) from MassIVE dataset MSV000096472
- Open the SingleCellApp.exe application and select 2 input demo files (using the
Select Envelope Filebutton to pick the .fastp and theSelect Single Cell Filebutton to pick the .dmt). Use the parameters shown in the screenshot of the SingleCellApp.exe in "MassIVE and GitHub Roadmap.docx" in Supplementary Data 5 (FDR: 0.10; Decoys/forward: 10; StoriGroupId: 1). - Click the
Run Single Cellbutton to process the demo dataset (should take approximately 30 minutes on a standard desktop) - Click the
Export score Matrixbutton to save the processed data as an XLSX file in the directory with the other demo files (use the standard name pca_input.xlsx as this is referenced in the QMD script) - Open the ss_GSVA.qmd R Markdown file using RStudio
- Install the prerequisite R packages by clicking
Installin RStudio
- Scroll down to the
Loading scPiMS datasection and update thedata_dirvariable to point to the directory where you saved the demo files. Be sure to include slashes at the end of the path.
- Run the QMD script by clicking the
Runmenu and selected theRun Alloption
- RStudio will prompt you in the Console (tab on bottom left side) to install any packages. Type
aand hit Enter to install.
- The figures from the processing will be displayed directly in the script window
You can use the SingleCellApp.exe application to process other datasets by following the same steps as the demo dataset. The inputs are a .dmt file (containing single cell ion data) and a .fastp (containing the proteoform data).