OMAP-21: Organ Mapping Antibody Panel (OMAP) for Multiplexed Antibody-Based Imaging of Palatine Tonsil (FFPE) with MICS on MACSima, v1.2

OMAP-21 was developed for MICS (MACSima Imaging Cyclic Staining) imaging of human formalin-fixed, paraffin-embedded<br>(FFPE) palatine tonsil samples. The tissue fixation and antigen retrieval protocol is detailed in Neil et al. (2023).<br>This panel employs the MACSima platform, described in Kinkhabwala et al. (2022), which enables highly multiplexed<br>imaging through iterative immunolabeling and photobleaching cycles using FITC, PE, and APC-conjugated antibodies. The<br>panel consists of 47 antibodies and the nuclear marker DAPI for alignment and segmentation. Most antibodies are<br>recombinant and incorporate a modified human IgG1 constant region to prevent Fc receptor binding and reduce<br>non-specific staining, thus eliminating the need for additional blocking. OMAP-21 provides spatial resolution of all<br>anatomical structures and most cell types in the human tonsil and closely mirrors previous OMAPs, including OMAP-1 for<br>human lymph node (Radtke 2024) and OMAP-10 for palatine tonsil (Müller, Rüberg, and Bosio 2023). Image analysis was<br>conducted using MACSIQ View Analysis software, which processes raw images from the MACSima run, generates stitched<br>composites, and supports downstream analytical functions such as cell segmentation, gating, normalization, tSNE/UMAP<br>dimensionality reduction, heat maps, distance, and cluster analysis—all interactively linked. The dataset and results<br>are available via Zenodo (Müller et al. 2024). OMAP-21 is the second MICS-based panel released, and v1.1 specifically<br>updates UniProt IDs for CD45 isoforms to reflect CD45RB (P08575-9), CD45RA (P08575-8), and CD45RO (P08575-4).<br><br>**Bibliography:**<br><br>* Kinkhabwala, Ali, Christoph Herbel, Jennifer Pankratz, Dmytro A. Yushchenko, Silvia Rüberg, Paurush Praveen, Sandy Reiß, et al. 2022. “MACSima Imaging Cyclic Staining (MICS) Technology Reveals Combinatorial Target Pairs for CAR T Cell Treatment of Solid Tumors.” *Scientific Reports* 12 (1): 1911. https://doi.org/10.1038/s41598-022-05841-4.<br>* Müller, Werner, Julia Femel, Silvia Rüberg, and Andreas Bosio. 2024. “OMAP-21: Organ Mapping Antibody Panel (OMAP) for Multiplexed Antibody-Based Imaging of Human Palatine Tonsil with MICS on MACSima 1.5.” Zenodo. https://zenodo.org/records/13132639.<br>* Müller, Werner, Silvia Rüberg, and Andreas Bosio. 2023. “10-Palatine-Tonsil-Mics (v1.0) Graph Data, v1.0.” 2023. https://lod.humanatlas.io/omap/10-palatine-tonsil-mics/v1.0/.<br>* Neil, Emily, Dongju Park, Rebecca C. Hennessey, Eric C. DiBiasio, Michael DiBuono, Hanna Lafayette, Erica Lloyd, et al. 2023. “Spatial Protein and RNA Analysis on the Same Tissue Section Using MICS Technology.” bioRxiv. https://doi.org/10.1101/2023.10.27.564191.<br>* Radtke, Andrea J. 2024. “1-Human-Lymph-Node-Ibex (v1.4) Graph Data, v1.4.” 2024. https://lod.humanatlas.io/omap/1-human-lymph-node-ibex/v1.4/.