MALDI-Based Multiomics and Spatial Molecular Profiling—Principles, Integrated Workflows, Biomedical Applications, and Future Perspectives: A Systematic Review

Abstract

Although expression-based omics has greatly contributed to biological research, the biochemical state of cells or tissues cannot be fully explained by transcript or gene expression data alone. This systematic review investigated when matrix-assisted laser desorption/ionisation (MALDI)-based approaches can support the integration of multiple molecular layers and when they remain limited to single-class molecular mapping. PubMed was searched between 1 February and 8 June 2026. English-language peer-reviewed publications relevant to the principles, methods, molecular classes, or biomedical applications of matrix-assisted laser desorption/ionisation-based multiomics were eligible. Publications outside this scope were excluded. Records were independently screened by two reviewers, and disagreements were resolved through discussion. The findings were narratively synthesised based on the analytical platform, molecular layer, integrated spatial multiomics, spatial application, and biomedical use. The methodological quality and risk of bias were not formally assessed, which limits the certainty and strength of the conclusions drawn from this narrative synthesis. Following the reapplication of the relevance criteria, 111 publications from 1633 identified records were included in the final review. The evidence was reclassified and presented separately as true multilayer MALDI studies, MALDI integrated with orthogonal modalities, and background uniomics or spatial molecular profiling studies. These publications included human, animal, cellular, tissue-based, and methodological studies. These publications cover many molecular classes and biomedical applications. As many publications did not involve human participants, a single pooled participant total was not applicable to this study. Matrix-assisted laser desorption/ionisation-based techniques were used to analyse metabolites, lipids, glycans, peptides, and intact proteins in the reviewed literature. The literature also shows that spatial information can be retained using matrix-assisted laser desorption/ionisation imaging mass spectrometry (MALDI-IMS). Its ability to map molecular signals back to specific tissue regions or areas associated with disease supports its potential clinical utility. Specific areas of application, such as cancer, neurological, and infectious disease research, are also reviewed. The use of matrix-assisted laser desorption/ionisation-based techniques for biomarker discovery has been evaluated. Small sample sizes, proof-of-concept designs, inconsistent outcome reporting, and limited external validation have limited the evidence. Technical limitations include ion suppression, limited quantification, sample preparation variability, and differing analyte sensitivity. Emerging developments, such as single-cell imaging and AI analysis, have also been explored. Matrix-assisted laser desorption/ionisation enables multiomics when two or more complementary molecular layers are analysed and integrated within the same biological system. When only a single molecular class is analysed, the approach remains spatial molecular mapping rather than multiomic analysis. However, before matrix-assisted laser desorption/ionisation-based approaches can be applied in routine clinical practice, standardised workflows and better quantitative methods are needed. This review received no external funding and was not registered.