--- name: natural-isotope-abundance-calculation description: Use when when processing mass spectrometry imaging (MSI) data in positive ion mode where both [M+H]+ and [M+Na]+ adducts are present for the same lipid species, and you observe intensity overlap in [M+H]+ ion images caused by the isotopic fine structure of [M+Na]+ adducts. license: CC-BY-4.0 metadata: edam_operation: http://edamontology.org/operation_3629 edam_topics: - http://edamontology.org/topic_3520 - http://edamontology.org/topic_0121 tools: - LipidQMap - Cardinal techniques: - MS-imaging derived_from: - doi: 10.1101/2025.10.15.682422v1 title: LipidQMap evidence_spans: - LipidQMap writes MSI exports as HDF5 containers - LipidQMap writes MSI exports as HDF5 containers that follow the [`Cardinal::HDF5`](https://cardinalmsi.org) conventions. claims: [] provenance: collection: https://w3id.org/holobiomicslab/asb-skill/collection/metabolomics/v2 assembled_by: scripts/collect_metabolomics_collection.py sources: - build: coll_lipidqmap_cq doi: 10.1101/2025.10.15.682422v1 title: LipidQMap dedup_kept_from: coll_lipidqmap_cq schema_version: 0.2.0 attribution: generator: AgenticScienceBuilder original_doi: 10.1101/2025.10.15.682422v1 all_source_dois: - 10.1101/2025.10.15.682422v1 zenodo_doi: 10.5281/zenodo.20794027 curators: [] promoter: Louis-Félix Nothias sponsor: CNRS & Université Côte d'Azur --- # natural-isotope-abundance-calculation ## Summary Calculate the fractional contribution of sodium adducts ([M+Na]+) to protonated ions ([M+H]+) based on natural isotope abundance ratios, used to correct for isotopic overlap in mass spectrometry imaging. This calculation is the quantitative foundation for Type II isotopic correction in lipidomics. ## When to use When processing mass spectrometry imaging (MSI) data in positive ion mode where both [M+H]+ and [M+Na]+ adducts are present for the same lipid species, and you observe intensity overlap in [M+H]+ ion images caused by the isotopic fine structure of [M+Na]+ adducts. This is particularly critical in lipidomics workflows where accurate quantitation of lipid species requires removal of sodium-induced isotopic artifacts. ## When NOT to use - Input data contains only [M+H]+ adducts with no corresponding [M+Na]+ features — the correction will have no effect and is unnecessary. - Negative ion mode MSI data — the ~22 Da sodium mass shift and isotopic overlap pattern are specific to positive ion mode [M+H]+ vs [M+Na]+ competition. - Low-resolution or unit-resolution mass spectrometry where [M+H]+ and [M+Na]+ isotopic envelopes cannot be resolved and paired — the assumption that isotopic patterns can be calculated and matched breaks down. ## Inputs - Feature-by-pixel intensity matrix (float32, shape n_features × n_pixels) - Feature metadata with neutral lipid identifiers and m/z values - Paired feature assignments linking [M+H]+ and [M+Na]+ features - Elemental composition or natural isotope abundance lookup table ## Outputs - Fractional isotopic contribution factors (one scalar per [M+Na]+/[M+H]+ pair) - Corrected [M+H]+ intensity matrix (float32, same shape as input) ## How to apply After identifying paired [M+H]+ and [M+Na]+ features by matching neutral lipid identifiers and confirming the ~22 Da m/z difference (mass difference between Na and H substitution), calculate the fractional isotopic contribution of each [M+Na]+ feature to its paired [M+H]+ feature using natural isotope abundance ratios specific to the lipid's elemental composition. This fractional value is then used as a scaling factor to subtract the [M+Na]+ intensity from the [M+H]+ intensity across all pixels. The calculation is grounded in the abundance of heavy isotopes (primarily 13C) naturally present in both the sodium and protonated forms; the Type II correction model assumes that the isotopic pattern of [M+Na]+ overlaps predictably with [M+H]+ based on these natural abundances. Apply the correction pixel-by-pixel, clamping negative corrected values to zero to preserve physical intensity constraints. ## Related tools - **LipidQMap** (Implements Type II isotopic correction by calculating natural isotope abundance ratios and subtracting scaled [M+Na]+ intensities from [M+H]+ intensities across all pixels in ion images) — https://github.com/swinnenteam/LipidQMap - **Cardinal** (Provides HDF5 data container format and conventions for storing feature-by-pixel intensity matrices and metadata required for isotopic correction workflows) — https://cardinalmsi.org ## Evaluation signals - Calculated fractional contributions are physically valid (0 ≤ fraction ≤ 1) and reflect expected natural 13C/12C and other isotope ratios for the lipid composition. - After correction, [M+H]+ intensities are equal to or lower than before (no negative values after clamping); intensity decreases are largest at pixels where [M+Na]+ signal is highest. - Paired [M+Na]+ and [M+H]+ features are correctly matched by neutral lipid ID and m/z difference of ~22 Da (within mass tolerance, typically ≤ 5 ppm for TOF instruments). - The corrected intensity matrix preserves the HDF5 layout (spectraData/intensity dataset, dimension scales, pixelData and featureData groups unchanged) and output dimensions remain n_features × n_pixels. - Visual inspection: toggling between raw and isotope-corrected ion images shows reduction of [M+H]+ image intensity in spatial regions where [M+Na]+ signal is prominent, with smoother distribution and reduced edge artifacts. ## Limitations - Calculation assumes that natural isotope abundance ratios are constant across all lipid species and do not vary with elemental composition; in practice, the 13C contribution may vary slightly with chain length and unsaturation. - The fractional contribution model assumes that [M+Na]+ isotopic pattern overlaps linearly with [M+H]+ fine structure; this breaks down if the mass difference leads to non-overlapping isotopic envelopes or if resolution is insufficient to resolve individual isotopomers. - Correction is only valid when [M+H]+ and [M+Na]+ features are correctly paired; misidentification due to m/z tolerance being too loose or neutral formula mismatch will propagate incorrect fractional values. - Clamping negative values to zero introduces a small bias in corrected intensities for pixels where [M+Na]+ contribution is estimated to exceed [M+H]+ signal; this typically occurs at low signal-to-noise pixels. - No changelog or version history provided in the LipidQMap repository, making it unclear whether the natural isotope abundance model has been validated or updated across releases. ## Evidence - [other] LipidQMap performs Type II isotopic correction by correcting [M+H]+ adducts for isotopic overlap contributed by [M+Na]+ adducts in ion images.: "LipidQMap performs Type II isotopic correction by correcting [M+H]+ adducts for isotopic overlap contributed by [M+Na]+ adducts in ion images." - [other] For each [M+Na]+ feature, calculate its fractional contribution to the corresponding [M+H]+ feature based on natural isotope abundance ratios (Type II correction model).: "For each [M+Na]+ feature, calculate its fractional contribution to the corresponding [M+H]+ feature based on natural isotope abundance ratios (Type II correction model)." - [other] Identify paired [M+H]+ and [M+Na]+ features by matching neutral lipid identifiers and comparing their m/z values (accounting for mass difference of ~22 Da for Na vs H substitution).: "Identify paired [M+H]+ and [M+Na]+ features by matching neutral lipid identifiers and comparing their m/z values (accounting for mass difference of ~22 Da for Na vs H substitution)." - [other] Subtract the scaled [M+Na]+ intensities from the [M+H]+ intensities across all pixels, clamping negative values to zero.: "Subtract the scaled [M+Na]+ intensities from the [M+H]+ intensities across all pixels, clamping negative values to zero." - [readme] Can perform Type II isotopic correction, and can correct [M+H]+ adducts for isotopic overlap from [M+Na]+ adducts.: "Can perform Type II isotopic correction, and can correct [M+H]+ adducts for isotopic overlap from [M+Na]+ adducts."