IntegrAlign: a comprehensive tool for multi-immunofluorescence panel integration through image alignment.

Hermet L, Laoubi L, Scavino M, Doffin AC, Gazeu A, Berthet J, Pillat B, Tissot S, Rusakiewicz S, Michallet MC, Bendriss-Vermare N, Valladeau-Guilemond J, Hausser J, Caux C, Hubert M

Bioinformatics 41 (11) - [2025-11-01; online 2025-10-16]

Tyramide-based multiplex-immunofluorescence (mIF) enables the simultaneous analysis of up to seven protein markers on a whole slide, providing a comprehensive approach to study the tumor microenvironment. Integrating multiple mIF panels through image alignment of serial slide significantly expands the number of cell populations analysed in a single space. IntegrAlign was developed to optimize this integration on serial whole slides, enhancing the value and applicability of -mIF for comprehensive spatial analyses and enabling biomarker discovery at scale. IntegrAlign, leveraging the SimpleITK toolkit, applies a two-step alignment using rigid and B-spline transformations to integrate serial mIF whole slides. Validation on simulated and real datasets demonstrated alignment accuracy below the diameter of a cell nucleus (∼6 µm), outperforming existing methods. This precision enhances spatial analyses by combining extended phenotypic data, supporting novel insights into tissue architecture and cellular interactions. IntegrAlign is open-source, implemented in Python, and freely available under the MIT license at https://github.com/CAUXlab/IntegrAlign.

PubMed 41100186

DOI 10.1093/bioinformatics/btaf567

Crossref 10.1093/bioinformatics/btaf567

pmc: PMC12582367
pii: 8287766