OMAP-16 Organ Mapping Antibody Panel (OMAP) for Multiplexed Antibody-Based Imaging of Human Lung with CODEX, v1.2

OMAP-16 was developed for Multiplexed Immunofluorescent (MxIF) imaging of formalin-fixed paraffin-embedded (FFPE)<br>human lung sections using the Akoya Biosciences Phenocycler-Fusion platform (Purkerson, Pryhuber, Colon, et al. 2023).<br>This panel includes 34 antibodies and the nuclear marker DAPI for image alignment and nuclear segmentation. OMAP-16<br>provides spatial context for airway, vasculature, and alveolar features, and detects 35 of 72 epithelial, endothelial,<br>and immune cell types listed in the ASCT+B Lung v1.2 table (Pryhuber 2021). It does not capture Cap1 and Cap2<br>endothelial cells, eosinophils/basophils, fibroblasts/pericytes, neuroendocrine cells, ciliated cells, chondrocytes,<br>or NK cells, though antibody barcode conjugates for these are under development. Cell segmentation was performed using<br>the StarDist extension in QuPATH 0.3.2, based on DAPI-stained nuclei. Vascular smooth muscle and airway epithelial<br>structures were annotated using thresholded ACTA2 and PanCK signals with the Pixel Classifier in QuPATH, with<br>resulting object data exported as GeoJSON files. Antigen retrieval was carried out using AR9 Buffer (Akoya<br>Biosciences) at 110°C for 20 minutes. Barcode-conjugated antibodies were sourced from Akoya Biosciences or created<br>through custom conjugation using their antibody conjugation kit (Cat# 232195). For optimal preservation of structural<br>features, antibodies targeting extracellular matrix and vascular smooth muscle should be imaged early (within the<br>first five imaging cycles). The antibody validation protocol and full imaging workflow are documented in the<br>referenced protocols.<br><br>**Bibliography:**<br><br>* Pryhuber, Gloria. 2021. “Lung (v1.2) Graph Data, v1.2.” https://lod.humanatlas.io/asct-b/lung/v1.2/.<br>* Purkerson, Jeffrey, Gloria S. Pryhuber, Luis Colon, and Heidie Huyck. 2023. “813.1 Multiplexed Immunofluorescence Phenocycler-Fusion® Imaging of FFPE Lung Sections.” https://www.protocols.io/view/813-1-multiplexed-immunofluorescence-phenocycler-f-c4f5ytq6.<br>* Purkerson, Jeffrey, Gloria S. Pryhuber, Heidie Huyck, and Gail Deutsch. 2023. “812.1 Lung FFPE OMAP Multiplexed Immunofluorescence Phenocycler-Fusion® Antibody Validation Protocol.” https://www.protocols.io/view/812-1-lung-ffpe-omap-multiplexed-immunofluorescenc-c32ayqae.