OMAP-21: Organ Mapping Antibody Panel (OMAP) for Multiplexed Antibody-Based Imaging of Palatine Tonsil (FFPE) with MICS on MACSima, v1.1

OMAP-21 was designed for MICS (MACSima imaging cyclic staining) imaging of FFPE human tonsil samples. <br>Tissue fixation and antigen retrieval is described in (Spatial protein and RNA analysis on the same tissue section<br>using MICS technology). The MACSima technology is described in detail in the following publication (MACSima imaging<br>cyclic staining (MICS) technology reveals combinatorial target pairs for CAR T cell treatment of solid tumors). Most,<br>but not all, antibodies in this panel are recombinant antibodies with a mutated human IgG1 constant region. The<br>described mutation removes the Fc receptor binding capacity of human IgG1, eliminating the need for additional<br>blocking steps and reducing non-specific binding of human antibodies on human tissues. Highly multiplexed imaging is<br>achieved through cycles of immunolabeling with FITC, PE, and APC conjugated antibodies and photobleaching to<br>eliminate fluorescence signal between imaging cycles. The panel contains 47 antibodies and the nuclear marker DAPI<br>for image alignment and nuclear segmentation. This OMAP provides a spatial context for all anatomical structures and<br>most cell types present in the human palatine tonsil. OMAP-21 follows closely OMAP-1 described for human lymph nodes<br>(omap-1-human-lymph-node-ibex) and OMAP-10 (omap-10-palatine-tonsil-macsima). All reagents were obtained from<br>Miltenyi Biotec and have been rigorously tested through an internal quality control system to have minimal variation<br>between lots. For this reason, lot information is not included in this table. Analysis was performed by an<br>accompanied software package MACSIQ View Analysis also described in the MACSima publication<br>(https://doi.org/10.1038/s41598-022-05841-4). The result of the analysis is included in the uploaded dataset. In<br>brief, the software processes the raw images of the MACSima run, generates stitched images, and then allows<br>downstream analysis including cell segmentation, cell gating, data normalization, dimension reduction plots (tSNE,<br>UMAP), heat maps, distance analyses and cluster analyses, all of which are interactively linked together. The MACSima<br>system is continuously evolving, this is the second OMAP for the MACSima system. v1.1 updates UniProt IDs for CD45<br>splice variants: P08575-9 CD45RB, P08575-8 CD45RA, P08575-4 CD45R0 vs P08575 CD45.