OMAP-17: Organ Mapping Antibody Panel (OMAP) for Multiplexed Antibody-Based Imaging of Thymus with IBEX, v1.0

OMAP-17 was designed for IBEX (Iterative Bleaching Extends multi-pleXity) imaging of fixed frozen human pediatric<br>thymus samples (https://doi.org/10.1038/s41596-921-00644-9; https://doi.org/10.1073/pnas.2018488117). The panel<br>consists of 9 cycles of 43 primary antibodies, 6 secondary antibodies, and the nuclear label Hoechst for image<br>alignment and nuclear segmentation. This OMAP provides a spatial context for 7 anatomical structures and more than 30<br>cell types and cell states (dividing, immature, precommitted) present in the ASCT+B thymus table<br>(https://doi.org/10.48539/HBM668.QPSF.678). This panel was specifically designed to phenotype unique, and often rare,<br>populations of thymic epithelial cells (TECs) of the medulla, e.g., myo-TECs and neuro-TECs. Marker selection was<br>guided by companion studies using advanced sequencing methods (scRNA-seq and CITE-seq) as well as expert review.<br>OMAP-17 is the first OMAP for the human thymus but targets several stromal and immune cell types included in OMAPs for<br>other lymphoid organs. OMAP-1 (lymph node; https://doi.org/10.48539/HBM458.NSWR.957), OMAP-11 (spleen;<br>https://doi.org/10.48539/HBM957.RHVB.545), and OMAP-10 (tonsil; (https://doi.org/10.48539/HBM663.JJMJ.628). This OMAP<br>does not contain markers for phenotyping plasma cells or ionocytes/tuft TECs. CD1a, a critical marker for developing<br>thymocytes, was removed from OMAP-17 due to steric hindrance of T cell markers in later cycles. Several antibodies<br>required custom conjugation by commercial suppliers or antibody conjugation kits (e.g., AF532 and iF594 conjugates).<br>This work is included in a spatial thymus atlas constructed by the Human Cell Atlas. Additional details on image<br>analysis, data integration, and the establishment of a common coordinate framework for the thymus can be found in the<br>accompanying paper (https://doi.org/10.1101/2023.10.25.562925). Note that three (3) of the antibodies have been<br>discontinued since this OMAP-17 was created: Novus Biologicals (catalog #:NBP2-54402PE) KRT10, Novus Biologicals<br>(catalog #: NBP2-75467) LY75 and this secondary antibody Thermo Fisher Scientific Z25303 AF532 conjugate.