Workflow diagram
The workflow diagram summarises the entire workflow and can help you make decisions about which tools are most relevant to your analysis based on the type of Mass Spectrometry you have used.
The diagram is split into sections which correspond to the stages of the guide (see sidebar menu). Click on an element to navigate the guide or use the menu to the left.
Open source\nlocal\nversion control with git
Open source\nonline\nsense checks
Stage 1 - Metabolite Extraction
Quenching, extraction & phase separation
Growth & isolation of cells
Stage 2 - Data Acquisition
Standardized GCMS workflow
Stage 3 - Data conversion to open format
Convert data\n Proteowizard > MSConvert\n * scanEvent\n * polarity
.mzML open format\n profile
Convert & compress data\n Proteowizard > MSConvert\n * Scan range\n * scanEvent\n * polarity
.mzML open format\n profile, selected scans
Convert & compress data\n Proteowizard > MSConvert\n * peakPicking\n * scanEvent\n * polarity
.mzML open format\n centroid
Stage 4 - Data preprocessing
MassUp or R to run\n MALDIquant\nbaseline correction, normalization, alignment, peak detection
XCMSonline or R to run\n XCMS\nPeak picking, RT correction, alignment, grouping
Stage 5 - Extract & format peak table & metadata
Download each sample's .csv\n and tidy in R
.csv peak table\n samples as rows, m.z as columns
Download results folder\n and tidy in R
.csv metadata\n sample, independent variable1 ...
Stage 6 - Multivariate analysis
Open source, online, \nlocal and/or proprietary?
pcaMethods & muma\n using R & RStudio
Proprietary software\n e.g. SIMCA by Umetrics
Exploratory analysis\n PCA, PERMANOVA & interpretation
Directed analysis where appropriate\n OPLS-DA & detailed stats
List of features of interest\nm/z or m/z with RT
Graphs & stats for reporting
Stage 7 - Putative metabolite identification
Use features of interest\n to search databases/repositories and literature\n e.g. METLIN, KEGG, PubChem, Cyc, MassBank
Putative identities of metabolites\n responsible for differences between classes
Further MS to confirm IDs\n targeted, tandem or NMR
Pathway analysis, enrichment analysis
Stage 8 - Data archiving and reporting
Prepare data for\n submission to repository
Submit data to MetaboLights
Remember to cite tools used